Histone modifications: Unveiling the epigenetic enigma of degenerative skeletal diseases

Abstract

Degenerative skeletal diseases, including osteoporosis, osteoarthritis, and intervertebral disc degeneration, are prevalent age-related conditions characterized by progressive tissue degeneration and functional decline. Histone modifications are covalent modifications of histone residues, catalyzed by specific enzymes, that modulate chromatin architecture and transcriptional activity. Accumulating evidence highlights the critical involvement of histone modifications in orchestrating disease-associated transcriptional programs. In osteoporosis, histone modifications regulate osteoblast and osteoclast differentiation, thereby disrupting bone homeostasis. In osteoarthritis, they drive the expression of matrix-degrading enzymes in chondrocytes, contributing to cartilage degradation. In intervertebral disc degeneration, they are implicated in nucleus pulposus cell senescence, apoptosis, and extracellular matrix degradation. This review summarizes the distinct mechanistic roles of histone modifications across these conditions and explores the therapeutic potential of targeting histone-modifying enzymes, underscoring epigenetic regulation as a promising strategy for precision intervention in degenerative skeletal diseases.

The translational potential of this article: This review comprehensively explores the role of histone modifications in degenerative skeletal diseases and evaluates the potential of histone-modifying enzyme inhibitors as therapeutic targets. These insights provide new strategies and directions for the treatment of degenerative skeletal diseases.

Graphical abstract

1. Introduction

A class of age-related disorders known as degenerative skeletal diseases, including osteoporosis (OP), osteoarthritis (OA), and intervertebral disc degeneration (IDD), cause pain and motor difficulties as a result of persistent, long-term deterioration of bone and cartilage [[1], [2], [3]]. Statistics show that the global prevalence of OA increased from approximately 256 million cases in 1990 to 606.9 million cases in 2021, representing a 137 % rise [4]. In China, OA cases reached 152.85 million in 2021, representing a significant increase of approximately 186.5 % over the three decades since 1990 [5]. Low back pain (LBP), one of the leading causes of disability, affected over 619 million people globally and 78.49 million in China in 2021, with global prevalence projected to rise to 890 million by 2050 [6,7]. Degenerative skeletal diseases have become a leading cause of disability worldwide, posing a huge challenge to healthcare systems worldwide and placing a heavy burden on people's health and national economies [[8], [9], [10]]. However, its treatment is still mainly limited to conservative or surgical treatments for symptomatic relief [2]. Among them, conservative drug treatment may cause drug dependence and limited therapeutic effects, while surgical treatment may have serious complications [[11], [12], [13]]. Therefore, there is a lack of efficient therapeutic methods and medications to address the etiology. Insufficient comprehension of its pathogenesis is the primary cause of this. Consequently, it is crucial to investigate the pathophysiology of degenerative skeletal diseases and identify potential targets for treatment.

Epigenetic modifications refer to reversible and heritable covalent modifications of nucleic acids and histones that modulate gene expression without altering the DNA sequence [14]. They mostly consist of DNA methylation, histone modifications, RNA methylation, and non-coding RNA regulation [15]. Increasing evidence has suggested that these epigenetic layers do not function in isolation but rather engage in dynamic crosstalk. For instance, histone modifications can influence DNA methylation patterns, while non-coding RNAs can recruit or inhibit histone-modifying enzymes, thereby jointly regulating chromatin accessibility and gene transcription [16]. Such interactions collectively shape the epigenetic landscape of degenerative skeletal diseases [17]. Although various epigenetic processes contribute to disease progression, this review focuses on histone modifications.

Histones are abundant in positively charged basic amino acids and serve as the fundamental structural proteins of eukaryotic chromatin [18]. They constitute the core histone octamer, which interacts with DNA containing negatively charged phosphate groups to create nucleosomes, the chromatin's fundamental structural unit [19]. Various post-translational modifications of histone, mainly occurring on its terminal tails and globular core domains, including methylation, acetylation, phosphorylation, and ubiquitination [19,20]. By changing chromatin condition and affecting transcription factors' ability to bind to DNA promoters, these modifications have an impact on gene expression [21]. Numerous research has demonstrated the close relationship between histone modifications and the skeletal system, as well as their involvement in the regulation of various bone physiological processes, including osteogenesis, osteoclast differentiation, and chondrocyte function [[22], [23], [24]]. These findings implicate histone modifications in the development of various orthopedic disorders, particularly degenerative skeletal diseases. In addition, there is growing proof that histone modifications contribute significantly to the pathogenesis of degenerative skeletal diseases by regulating relevant gene expression [[25], [26], [27], [28], [29]]. Therefore, elucidating the molecular functions of histone modifications in degenerative skeletal diseases is essential for understanding their pathogenesis and identifying viable therapeutic targets. However, a comprehensive summary of this topic remains lacking.

This review attempts to broadly summarize the recent progress of histone modifications in degenerative skeletal diseases, mainly OP, OA, and IDD, and discuss its potential application value in clinical treatment. To ensure comprehensive and relevant coverage, we conducted a literature search in PubMed, Web of Science, and Scopus databases using keywords such as “histone modification,” “epigenetics,” “osteoporosis,” “osteoarthritis,” and “degenerative skeletal diseases.” The search covered publications from 2000 to 2025, and only peer-reviewed English-language studies were included.

2. Classification of histone modifications

Since the initial discovery of histone modifications, their diversity has expanded significantly. According to current literature, more than 20 distinct types of histone post-translational modifications (PTMs) have been identified to date [19,30] (Table 1). In addition to the four classical modifications—acetylation, methylation, phosphorylation, and ubiquitination—numerous novel acylation marks have been characterized, such as lactylation, crotonylation, and succinylation [31,32]. Moreover, other modifications with limited investigation but experimental validation have also been reported, including citrullination, SUMOylation, ADP-ribosylation, and biotinylation [32] (Fig. 1). Among these, acetylation and methylation remain the most extensively studied, primarily due to their central roles in gene regulation. These modifications are dynamically regulated by three major classes of chromatin-associated proteins. Writers, such as histone acetyltransferases (HATs), histone methyltransferases (HMTs), serine/threonine kinases and E3 ubiquitin ligases, act as chromatin-modifying enzymes that catalyze the covalent addition of specific functional groups to histone tails. In contrast, Erasers, including histone deacetylases (HDACs), histone demethylases (HDMs), phosphatases and deubiquitinating enzymes (DUBs), remove those same marks to reverse epigenetic states [33]. Furthermore, Readers, which comprise non-catalytic recognition factors such as bromodomain-, chromodomain-, Tudor domain- or PHD finger-containing proteins, selectively bind to modified histones and recruit downstream effectors to execute a wide range of biological outcomes [34].

Table 1.

The overview of histone modifications.

Histone PTMs		Discovery Year	Writers	Erasers	Readers	References
Acylations	Acetylation	1964	HATs(GNATs, MYSTs, P300/CBP)	HDACs(HDAC1–11, SIRT1–7)	Bromodomain protein(BRD4, PCAF)	[44,[47], [48], [49]]
	Propionylation	2007	P300/CBP; GNATs; MYSTs	SIRT1-3	Bromodomain; YEATS domain	[[67], [68], [69], [70]]
	Butyrylation	2007	P300/CBP; MYSTs	SIRT1-3	Bromodomain (BRD4,BPTF); CECR 2; AF1	[67,69,70]
	Formylation	2008	NA	NA	NA	[71]
	Crotonylation	2011	P300/CBP; GNATs; MYSTs	HDAC3; SIRT1-3	YEATS domain (AF9; ENL)	[[72], [73], [74], [75]]
	Succinylation	2012	KAT2A; HAT1; P300/CBP	SIRT5; SIRT7	NA	[[76], [77], [78]]
	Malonylation	2012	KAT2A	SIRT5	NA	[[78], [79], [80]]
	β-hydroxyisobutyrylation	2014	P300/CBP; MYSTs	HDAC1-3; SIRT1-3	NA	[[81], [82], [83]]
	β-hydroxybutyrylation	2016	P300/CBP	SIRT3; HDAC1-2	NA	[84,85]
	Benzoylation	2018	NA	SIRT2	YEATS domain (YEATS2; AF9)	[86,87]
	Glutarylation	2019	KAT2A	SIRT7	NA	[88]
	Lactylation	2019	P300/CBP	HDAC1-3	Brg1	[57,60,89]
Ubiquitin-like	Ubiquitination	1980	RNF20/40; PRC1(Ring1B); BRCA1/BARD1	USP16, 22; PR-DUB(BAP1)	PRC2(Jarid2, AEBP2); RYBP; 53BP1; DNMT1	[[90], [91], [92], [93]]
	SUMOylation	2003	SAE1/SAE2, UBC9, PIAS	SENP1-3, 5–7; DeSI-1,2; USPL1	NA	[[94], [95], [96]]
	Ufmylation	2019	UFL1	UFSP2	STK38	[97,98]
Others	Methylation	1964	SET domain (SUV39H1/2, SETDB1/2, EZH1/2, etc.); Non-SET (Dot1L)	FAD-dependent (LSD1/2); JmjC domain (KDM2-8)	Chromodomain; MBT and Tudor domain; PHD finger	[33,40,44,99]
	Phosphorylation	1966	Kinases (MSK, AMPK, WSTF, CK2, etc.)	Phosphatase (PP1, PP2A, PP4, etc.)	MDC1, Survivin, 14-3-3	[50,[100], [101], [102], [103]]
	ADP-ribosylation	1977	ARTDs (ARTD1-17); ARTCs (ARTC1-5); ADPRT1a; ADPRT2; Sirtuins	PARG; TARG1; ARH1; ARH3; MacroD1; MacroD2	PBM; Macrodomain; WWE domain; PBZ domain	[[104], [105], [106]]
	O-GlcNAcylation	1984	OGT	OGA	NA	[[107], [108], [109]]
	Biotinylation	2001	HCS	NA	NA	[110,111]
	Citrullination	2002	PAD2; PAD4	NA	NA	[75,112,113]
	Serotonylation	2019	TGM2	TGM2	WDR5	[[114], [115], [116]]
	Dopaminylation	2020	TGM2	NA	NA	[117]

Fig. 1.

The classification of histone modifications. Histone modifications are highly diverse, including a wide range of types such as methylation, acetylation, phosphorylation, ubiquitination, and lactylation.

2.1. Histone methylation

The discovery of histone methylation dates back to the 1960s [35]. But it wasn't until 2000 that the first histone methyltransferase, SUV39H1, was identified [36]. It was announced in 2004 that LSD1, the first histone demethylase, exists [37].

Lysine (K) and arginine (R) residues are the primary sites of histone methylation, with the former undergoing mono-(me1), di-(me2), and trimethylation(me3), and the latter undergoing mono-(me1), symmetric-dimethylation(me2s), and asymmetric-dimethylation(me2a) [38]. Instead of changing the net charge, histone methylation increases hydrophobicity and alters intra- or intermolecular interactions to influence transcription [39]. In general, methylation of H3K4, H3K36, H3K79, and H3R17 is thought to promote transcription, whereas methylation of H3K9, H3K27, and H4K20 is thought to repress transcription [40].

HMTs catalyze the methylation of amino acid residues via S-adenosylmethionine (SAM) [41]. According to the catalytic domain sequence, lysine methyltransferases (KMTs) are divided into two groups: SET-containing and non-SET-containing; the former includes SUV, SET1, SET2, EZH, RIZ, and other protein families, while the latter is less common and mainly includes DOT1L [42,43].

HDMs are catalytic enzymes that mediate histone demethylation reactions. There are two kinds of lysine demethylases (KDMs): the lysine-specific demethylases (LSD) family, which mediate mono- and di-demethylation of lysine residues; and the Jumonji C (JmjC) domain-containing protein family, which mediate mono-, di-, or tri-demethylation of lysine residues [41].

2.2. Histone acetylation

The first evidence of the function of histone acetylation in the control of gene expression was shown in 1964 [44]. Subsequently, in 1996, the first histone acetyltransferase and histone deacetylase were identified [45].

Histone acetylation occurs mainly at the lysine residues in the N-terminal of histones. Histone acetylation often results in transcriptional activation because it neutralizes the positive charge of histones, which reduces the contact between DNA and histones and facilitates DNA binding to transcription factors [46].

HATs catalyze the transfer of acetyl groups from acetyl-CoA to histone lysine residues [47]. In humans, HATs are classified into three families: the p300/CBP family, the MYST family, and the GNAT family [48]. HDACs catalyze the deacetylation reaction by hydrolyzing acetyl groups on lysine residues. HDACs comprise four families: Classes I, II, and IV are Zn²⁺-dependent, while Class III/Sirtuins are NAD-dependent [49].

2.3. Other types of histone modifications

Histone phosphorylation is regulated by protein kinases and phosphatases through phosphorylation and dephosphorylation of histone tails [50]. Similar to acetylation, histone phosphorylation acts by increasing the negative charge to neutralize the positive charge of histones, attenuating DNA-histone interactions and affecting chromatin structure [51]. However, unlike the previous two, histone phosphorylation participates in histone modification crosstalk by affecting the binding of modifier and effector proteins to histone modification sites [51,52].

Histone ubiquitination modification adds a small-molecule protein called ubiquitin (Ub), which comprises 76 amino acids and depends on E1 activating, E2 conjugating, and E3 ligase enzymes, to the lysine residues in histone H2A and H2B tails [53]. Its catabolic release is mediated by DUBs [53]. Since ubiquitin itself has seven lysine residue sites, it allows histone-linked monoubiquitin to be further modified to form polyubiquitin chains [54]. Mostly found in heterochromatin, H2A monoubiquitination is linked to transcriptional repression, while mostly found in euchromatin, H2B monoubiquitination is linked to transcriptional activation [55].

Histone lactylation has rapidly become a key focus in epigenetic research since its discovery in 2019 [56]. As a novel epigenetic modification, it reveals a direct link between cellular energy metabolism—glycolysis—and epigenetic regulation. Lactate, the end product of glycolysis, is the critical substrate for histone lactylation, and its concentration closely correlates with lactylation levels [57]. By adding lactyl groups to lysine residues on histones, histone lactylation primarily activates gene expression, likely by loosening chromatin structure to promote transcription [58]. Current research on the specific writers, erasers, and readers of histone lactylation remains quite limited, with existing evidence suggesting that CBP/p300, HDAC1-3, and SIRT1-3 play crucial roles in the addition and removal of histone lactyl groups [57,59,60]. Given the abnormal glycolytic activity in tumor cells, histone lactylation's role in tumorigenesis has drawn significant attention [[61], [62], [63], [64]]. While studies on histone lactylation in orthopedics are just beginning, early findings suggest it plays a regulatory role in disease development [65,66].

3. The roles of histone modifications in degenerative skeletal diseases

Degenerative skeletal diseases are a prevalent and serious group of conditions that impose a substantial burden on individual health and socioeconomic systems. Among these, OP, OA, and IDD are three representative diseases. Histone modifications are vital epigenetic mechanisms that influence gene expression, resulting in alterations to cellular functions. Detailed exploration of histone modifications in the context of each disease is presented in the following sections (Table 2) (Table 3).

Table 2.

The roles of histone modifications in degenerative skeletal diseases.

	Proteins	Histone modification	Target genes	Cells	Biological functions	References
Osteoporosis	LSD1↑	H3K4me2↓	Wnt7b, BMP2↓	BMSCs	Inhibits osteogenic differentiation	[137]
	GCN5↓	H3K9ac↓	Wnt1, 6, 10a, 10b↓			[138]
	SETDB1↓	H3K9me3↓	OTX2↑			[139]
	KDM3a↓	H3K9me2↑	Erk2↓			[140]
	HDAC8↑	H3K9ac↓	Runx2↓			[129]
	HDAC6↑	H3K9/K14ac, H4K12ac↓	Runx2↓			[131]
	NAP1L2↑	H3K14ac↓	osteogenic gene↓			[132]
	KDM6b↓	H3K27me3↑	Runx2↓			[133]
	PKM2↓	H3K18la↓	osteogenic genes↓			[66]
	CS↓	H3K9ac↓	Runx2↓			[144]
	EZH2↓	H3K27me3↓	P16↑			[147,148]
	SETD2↓	H3K36me3↓	Lbp↓	BMSCs	Promotes adipogenic differentiation	[151]
	Ash1l↓	H3K4me3↓	Creb↓, PPARγ↑			[152]
	KDM7a↑	H3K9me2, H3K27me2↓	C/EBPα, Sfrp1↑			[153]
	KDM4a↑	H3K9me3↓	C/EBPα, Sfrp4↑			[154]
	BRD4↑	H3K9ac↑	Foxp1↑			[155]
	DOT1L↓, KAT2b↑	H3K79me2↓, H3K27ac↑	miR-181↓, SRSF1↑	BMMs	Enhances osteoclast differentiation	[157]
	PCAF↑	H3K27ac↑	CXCL12↑			[159]
	RNF40↑	H2BK120ub1↑	TNFSF11/RANKL↑			[160]
Osteoarthritis	HPIP↑	H3K56ac↑	Wnt target genes↑	Human chondrocytes	Enhances catabolism	[173]
	HOTAIR↑	H3K27me3↑	WIF-1↓	SW1353		[174]
	DOT1L↓	H3K79me2↓	Wnt target genes↑	Human chondrocytes		[175]
	Brd3, Brd4↑	H4K5/8/12ac↑	Matrix-degrading enzyme genes↑	SW1353		[179]
	JMJD3↑	H3K27me3↓	ZEB1↑	Human chondrocytes		[180]
	KAT6a↑	H3K9ac↑	IHH↑	Human chondrocytes, SW1353, C28/I2 cells		[182]
	ACLY↑	H3K9/27ac↑	iNOS, MMP3, MMP13↑	Human and mouse chondrocytes		[183]
	LDHA↑	H3K18la↑	TPI1↑	Mouse chondrocytes		[65]
	EZH2↑	H3K27me3↑	miR-138↓	Human chondrocytes		[184]
	/	H3K9me3↓	SOX9,Col2a1↓	ATDC5 cells	Inhibits anabolism	[187]
	UTX, PRC2↑	H3K27me3↑	Igf2↓	Mouse chondrocytes		[188]
	NSD1↓	H3K36me2↓	Osr2↓	Mouse chondrocytes		[189]
	JMJD3↑	H3K27me3↓	NR4A1↑	Mouse chondrocytes		[190]
	HDAC4↓	H3K9ac↑	ATF4↑	Human chondrocytes	Promotes apoptosis	[196]
	EZH2↑	H3K27me3↑	miR-142-3p↓	Human chondrocytes	Promotes pyroptosis	[201]
	NSD1↓	H3K36me2↓	SOX9↓	Mouse chondrocytes	Promotes ferroptosis	[205]
	LDHB↑	H3K18la↑	ACSL4↑	Mouse chondrocytes		[206]
Intervertebral disc degeneration	RREB1↓	H3K4me2/3↑, H3K9/27me3↓	ADAMTS-5↑	Human NP cells	Mediates ECM degradation	[228]
	SIRT6↓	H3K9ac↑	NF-κB target genes↑	Human NP cells		[231]
	KMT2D↑	H3K4me1↑	Mmp3/9/13↑	Human NP cells		[234]
	EZH2↑	H3K27me3↑	Sox-9↓	Primary rat endplate chondrocytes		[265]
	EZH2↑	H3K27me3↑	miR-129-5p↓	Human NP cells	Mediates NPC senescence	[240]
	SIRT6↓	H3K9ac↑	p-NF-κB p65↑	Human NP cells		[241]
	EZH2↓	H3K27me3↓	NOX4↑	Primary rat NP cells		[245]
	KDM4a↑	H3K9me3↓	ALKBH5↑	Human NP cells		[248]
	KDM5a↓	H3K4me3↑	WTAP↑	Human NP cells		[246]
	KMT2D↑	H3K4me2↑	miR-133a-5p↑	Primary mouse NP cells	Promotes apoptosis	[254]
	EZH2↑	H3K27me3↑	DKK1↓	Primary rat NP cells	Promotes pyroptosis	[256]
	GLS1↓	H3K9/K27ac↓	NFS1↓	Human NP cells	Promotes ferroptosis	[259]
	LDHA↑	H3K18la↑	ACSL4↑	Human NP cells		[260]

Table 3.

Classification of histone modification-related proteins and metabolites in degenerative skeletal diseases.

Category		Enzymes	Histone Modification	Target Genes	Diseases	References
Writers	Histone acetyltransferases	GCN5	H3K9ac	Wnt1, 6, 10a, 10b	Osteoporosis	[138]
		KAT2b/PCAF	H3K27ac	SRSF1, CXCL12	Osteoporosis	[157,159]
		KAT6a	H3K9ac	IHH	Osteoarthritis	[182]
	Histone methyltransferases	SETDB1	H3K9me3	OTX2	Osteoporosis	[139]
		EZH2	H3K27me3	P16, miR-138, miR-142-3p, Sox-9, miR-129-5p, NOX4, DKK1	Osteoarthritis/Intervertebral Disc Degeneration	[148,184,187,201,240,245,256,265]
		SETD2	H3K36me3	Lbp	Osteoporosis	[151]
		Ash1l	H3K4me3	Creb, PPARγ	Osteoporosis	[152]
		DOT1L	H3K79me2	miR-181, Wnt target genes	Osteoporosis/Osteoarthritis	[157,175]
		PRC2	H3K27me3	Igf2	Osteoarthritis	[188]
		NSD1	H3K36me2	Osr2, SOX9	Osteoarthritis	[189,205]
		KMT2D	H3K4me1/2	Mmp3/9/13, miR-133a-5p	Intervertebral Disc Degeneration	[234,254]
Erasers	Histone demethylases	LSD1	H3K4me2	Wnt7b, BMP2	Osteoporosis	[137]
		JMJD3	H3K27me3	ZEB1, NR4A1	Osteoarthritis	[180,190]
		UTX	H3K27me3	Igf2	Osteoarthritis	[188]
		KDM3a	H3K9me2	Erk2	Osteoporosis	[140]
		KDM4a	H3K9me3	C/EBPα, Sfrp4, ALKBH5	Osteoporosis/Intervertebral Disc Degeneration	[154,248]
		KDM5a	H3K4me3	WTAP	Intervertebral Disc Degeneration	[246]
		KDM6b	H3K27me3	Runx2	Osteoporosis	[133]
		KDM7a	H3K9me2, H3K27me2	C/EBPα, Sfrp1	Osteoporosis	[153]
	Histone deacetylases	HDAC4	H3K9ac	ATF4	Osteoarthritis	[196]
		HDAC6	H3K9/K14ac, H4K12ac	Runx2	Osteoporosis	[131]
		HDAC8	H3K9ac	Runx2	Osteoporosis	[129]
		SIRT6	H3K9ac	NF-κB targets genes	Intervertebral Disc Degeneration	[231,241]
Readers	Histone acetylation reader	BRD3	H4K5/8/12ac	Matrix-degrading enzymes	Osteoarthritis	[155,179]
		BRD4	H3K9ac	Foxp1	Osteoporosis	[179]
Metabolic precursors	Acetyl-CoA	CS	H3K9ac	Runx2	Osteoporosis	[144]
		ACLY	H3K9/27ac	iNOS, MMP3, MMP13	Osteoarthritis	[183]
		GLS1	H3K9/K27ac	NFS1	Intervertebral Disc Degeneration	[259]
	Lactate	PKM2	H3K18la	Osteogenic genes	Osteoporosis	[66]
		LDHA	H3K18la	TPI1	Osteoarthritis	[65,260]
		LDHB	H3K18la	ACSL4	Osteoarthritis/Intervertebral Disc Degeneration	[206]
Others	E3 ubiquitin ligase	RNF40	H2BK120ub1	TNFSF11/RANKL	Osteoporosis	[160]
	Histone chaperone	NAP1L2	H3K14ac	Osteogenic genes	Osteoporosis	[132]

3.1. Histone modifications and osteoporosis

Osteoporosis, a degenerative metabolic bone disease, is characterized by low bone mass, increased bone fragility, and microstructural destruction of bone tissue, which greatly increases the incidence of fragility fracture and brings tremendous pressure on the healthcare system and economy [[118], [119], [120]]. An investigation conducted in 2019 found that OP affects as many as 6.46 % and 29.13 % of Chinese men and women 50 years of age and older [121]. It is predicted that 4.83 million occurrences of osteoporotic fractures will occur annually by 2035, costing 19.92 billion dollars [122].

Bone is in a continuous process of renewal and metabolism called bone remodeling [123]. During this process, osteoblast-mediated bone formation and osteoclast-mediated bone resorption play a dominant role [124]. Between them, a dynamic equilibrium known as bone homeostasis is maintained, maintaining the skeleton's integrity [125,126]. The majority of research now available indicates that one of the main pathways leading to osteoporosis is the imbalance of bone homeostasis caused by multiple pathogenic factors [118,126,127]. Through altering the expression of important genes during bone formation and resorption, histone modifications contribute to the disruption of bone homeostasis and hence play a role in the progression of OP (Fig. 2).

Fig. 2.

Histone modifications-induced bone homeostasis dysregulation in OP pathogenesis. Histone modifications disrupt bone homeostasis and contribute to the occurrence and development of OP by inhibiting osteogenic differentiation, promoting adipogenic differentiation, and enhancing osteoclast differentiation.

3.1.1. Histone modifications inhibit osteogenic differentiation

3.1.1.1. Histone modifications mediate disruption of osteogenic regulatory networks

Runx2 is a major transcription factor during osteogenic differentiation and binds to cis-acting elements to transactivate major osteogenic matrix protein genes, which promote osteogenic differentiation and bone formation [128]. Several studies have demonstrated that Runx2 is regulated by histone acetylation, whereas multiple histone deacetylases, including HDAC4, 6, and 8, decrease histone acetylation level on the Runx2 promoter, inhibit its expression, limit BMSCs' capacity for osteogenic differentiation, and contribute to the development of OP [[129], [130], [131]].

Hu et al. found that elevated levels of the histone chaperone NAP1L2 suppressed osteogenic gene expression, including Runx2, by recruiting SIRT1 to decrease H3K14ac levels at their promoters, thereby inhibiting osteogenic differentiation [132]. According to Behera et al., histone methylation plays a part in the regulation of Runx2, which is connected to the mechanism via which senescence-associated mitochondrial dysfunction inhibits osteogenic differentiation [133]. They found that the release of mt-DNA from dysfunctional mitochondria in senescent BMSCs inhibited the production of the histone demethylase KDM6b, which resulted in the increased level of H3K27me3 in the Runx2 promoter region and the decreased expression of Runx2. These outcomes ultimately hindered BMSCs' capacity for osteogenic differentiation and reduced bone density [133].

The Wnt/β-catenin signaling pathway is a pivotal regulator across multiple degenerative skeletal diseases, governing the fate and phenotype of bone-related cells, including bone marrow mesenchymal stem cells (BMSCs), osteoblasts, chondrocytes, and osteoclasts [[134], [135], [136]]. Wnt signaling is essential for bone homeostasis, promoting osteogenic differentiation of BMSCs while suppressing adipogenic and chondrogenic commitment [136]. Dysregulation of Wnt signaling leads to disrupted bone formation and OP, with histone modifications playing a critical role.

Sun et al. demonstrated that the histone demethylase LSD1 repressed BMSC osteogenic differentiation by reducing H3K4me2 levels at the promoters of Wnt7b and Bmp2, thereby downregulating their expression and bone formation [137]. Furthermore, Jing et al. discovered that the expression of histone acetyltransferase GCN5 was reduced in BMSCs from ovariectomy (OVX) rats. This resulted in a lower level of H3K9ac in the Wnt gene promoter region, which inhibited Wnt protein expression, prevented osteogenic differentiation, and reduced bone mass [138]. According to Hu et al., osteoporotic femoral tissues had downregulated expression of histone methyltransferase SETDB1, which had an impact on osteogenic differentiation [139]. Mechanistically, the OTX2 promoter region is modified by SETDB1 through H3K9me3, which inhibits OTX2 expression. This, in turn, promotes the BMP-Smad and Wnt/β-catenin pathways, facilitating osteogenic differentiation. Reduced expression of SETDB1 causes osteogenic differentiation to be inhibited and OP to develop [139]. Wu et al. found that in BMSCs from OVX rats, the H3K9me2 demethylase KDM3a was inhibited by increased miR-199a-3p, which resulted in elevated histone methylation levels and reduced Erk2 expression, inhibiting osteogenic differentiation and leading to OP [140].

3.1.1.2. Histone modifications mediate metabolism-associated osteogenic dysfunction

Recent research focuses on the crucial role of cellular metabolism in osteogenic differentiation of BMSCs and osteoblasts [141,142]. Glucose metabolism participates in histone modifications by providing intermediates like SAM and acetyl-CoA, which serve as substrates for histone methylation and acetylation, thereby affecting gene expression during bone formation [143]. Da et al. found that BMSCs from OVX mice exhibit an abnormal tricarboxylic acid cycle in which citrate depletion impairs acetyl-CoA synthesis, reducing histone acetylation of osteogenic genes and suppressing their differentiation potential [144].

In addition to acetylation, histone lactylation, a novel modification derived from lactate produced during glycolysis, has emerged as an important epigenetic regulator. Increasing evidence indicates that lactate-induced histone lactylation enhances chromatin accessibility and activates the gene transcription [57]. Nian et al. and Minami et al. highlighted the regulatory potential of histone lactylation in osteoblast differentiation. They reported that LDHA-driven lactate production promotes H3K18 lactylation (H3K18la) and activates osteogenic gene transcription, thereby enhancing osteoblast differentiation and bone formation [145,146]. Wu et al. further demonstrated the involvement of histone lactylation in OP. They reported that serum lactate levels were reduced in patients with OP, and BMSCs isolated from these patients exhibited decreased H3K18la levels along with reduced expression of osteogenic genes [66]. They revealed that in OVX mice, reduced expression of vascular endothelium pyruvate kinase M2 (PKM2), a key glycolytic enzyme, lowered serum lactate levels and diminished H3K18la levels on osteogenic genes such as COL1A2, COMP, and ENPP1. This epigenetic alteration suppressed their transcription, impaired osteogenic differentiation, and ultimately accelerated OP progression [66].

In addition, by triggering the senescence signaling pathway, histone modification accelerates the cellular senescence of BMSCs, which decreases their osteogenic differentiation and proliferation abilities, promotes apoptosis, and disrupts bone homeostasis [147,148].

3.1.2. Histone modifications mediate osteogenic-adipogenic differentiation imbalance

Although BMSCs are multipotent stem cells with three-lineage differentiation potential of osteogenesis, adipogenesis, and chondrogenesis, many investigations have revealed that the differentiation of BMSCs into osteogenic and adipogenic lineages is negatively correlated [149]. Consequently, an imbalance in differentiation is one of the main factors contributing to OP, maintaining the osteogenic-adipogenic differentiation balance of BMSCs is crucial for bone homeostasis and remodeling [149,150]. Some studies have reported the regulatory mechanism of histone modifications in the imbalance of osteogenic-adipogenic differentiation of BMSCs.

Histone methyltransferases regulate osteogenic-adipogenic differentiation. Wang et al. found that lipopolysaccharide-binding protein (Lbp) regulated by SETD2-mediated H3K36me3, controlled the equilibrium between BMSCs' osteogenic and adipogenic differentiation [151]. The deficiency of SETD2 inhibits the expression of Lbp, which causes BMSCs to differentiate into adipocytes but not osteoblasts. Yin et al. discovered that histone methyltransferase Ash1l prevents bone mass loss and that it was dramatically downregulated in the bone tissue of OP model mice [152]. Further research revealed that Ash1l directly regulated the level of H3K4me3 of osteogenesis-related genes and, through modulating the repressor Creb, indirectly regulated the expression of PPARγ, an adipogenic transcription factor [152]. Therefore, down-regulating Ash1l expression promotes adipogenic differentiation while inhibiting osteogenic differentiation.

Histone demethylases regulate osteogenic-adipogenic differentiation. Wang et al. observed that by decreasing H3K9me2 and H3K27me2 levels at the promoters of C/EBPα and Sfrp1, KDM7a upregulated their expression. This, in turn, enhanced adipogenic differentiation and suppressed osteogenic differentiation by interacting with PPARγ and blocking Wnt signaling [153]. They also found a similar regulatory mechanism on KDM4a [154].

Lian et al. also reported a mechanism by which histone acetylation Readers regulate osteogenic-adipogenic differentiation. This study revealed that BMSCs from the glucocorticoid-induced OP model had an elevated expression of BRD4, a histone acetylation reader that promoted Foxp1 expression by binding to H3K9ac at the Foxp1 promoter. As a transcription factor, Foxp1 promotes the transcription of PPARγ2 and enhances adipogenic differentiation [155].

3.1.3. Histone modifications enhance osteoclast differentiation

The mechanism by which histone methylation regulates osteoclast differentiation has been reported in several studies [[156], [157], [158]]. Wang et al. reported the mechanism by which the DOT1L-miR-181-KAT2b-SRSF1 axis regulates osteoclast differentiation [157]. Low expression of histone methyltransferase DOT1L in OVX mice and OP patients reduces the level of H3K79me2 of the miR-181 promoter and its expression, which leads to high expression of KAT2b and SRSF1, which ultimately enhances osteoclast differentiation from bone marrow mononuclear macrophages (BMMs) and promotes OP progression [157]. The regulatory role of histone acetylation has also been reported. Lian et al. found that histone acetyltransferases PCAF-mediated H3K27ac regulate the osteoclast regulatory factor CXCL12, whereas the loss of estrogen reduces miR-29a inhibition of PCAF, which in turn leads to H3K27 hyperacetylation and up-regulation of CXCL12 expression and promotes osteoclast differentiation and OP development [159]. Najafova et al. also reported a mechanism by which H2B monoubiquitination regulates the TNFSF11 (encoding RANKL) gene to enhance osteoclast differentiation [160]. In response to stimulus factors, transcription factors (TF) are recruited to enhancers that recruit the H2B ubiquitin ligase RNF40 to target genes, thereby elevating H2BK120ub1 levels and promoting H3K4me3 levels near the transcription start site (TSS), which in turn promotes transcription of TNFSF11, elevates RANKL expression, and promotes osteoclast differentiation [160].

In conclusion, histone modifications lead to the imbalance of bone homeostasis by suppressing osteogenic differentiation, mediating the disproportion between osteogenic and adipogenic differentiation, enhancing osteoclast differentiation, and ultimately promoting the progression of OP.

3.2. Histone modifications and osteoarthritis

Osteoarthritis is a degenerative joint disease characterized by the gradual breakdown of articular cartilage, secondary synovitis, and bone remodeling [161,162]. Its prevalence and disability are rising annually, placing a significant financial burden and medical pressure on the affected population [9,163]. According to a growing body of research conducted in recent years, histone modifications have a significant impact on the progression of OA [24,164,165] (Fig. 3).

Fig. 3.

Histone modifications disrupt chondrocyte function and contribute to OA development. They promote OA progression by enhancing catabolic processes, inhibiting anabolic activities, and inducing chondrocyte death.

3.2.1. Histone modifications regulate extracellular matrix

Articular cartilage mainly refers to hyaline cartilage, which consists of chondrocytes, extracellular matrix (ECM), and water, and is a non-vascularized tissue that enables joint movement by providing a smooth surface with minimal friction [166]. Cartilage homeostasis, is largely dependent on chondrocytes for the proper synthesis and breakdown of ECM components [167,168]. One of the important mechanisms in the development of OA is the ECM degradation and disrupted cartilage homeostasis caused by an imbalance between anabolic and catabolic gene expression in chondrocytes [169]. By modulating the balance between catabolic and anabolic processes at the gene level, histone modifications disrupt the integrity of ECM, thereby impairing cartilage homeostasis and facilitating the progression of OA.

3.2.1.1. Histone modifications enhance catabolism

Wnt signaling, which is epigenetically suppressed in OP to inhibit osteogenesis, is abnormally hyperactivated in OA, driving the expression of matrix-degrading enzymes and disrupting cartilage homeostasis [[170], [171], [172]]. Histone acetylation has been demonstrated by Ji et al. to regulate the Wnt pathway [173]. They found that Hematopoietic PBX-interacting protein (HPIP) was elevated in OA cartilage, and formed a complex with LEF1 to recruit histone acetyltransferase p300. This resulted in an increase in H3K56ac in the promoters of Wnt target genes, activated Wnt signaling, and induced the production of catabolic factors, all of which disrupted cartilage homeostasis and accelerated the progression of OA [173]. Histone methylation has also been demonstrated by Yang et al. to regulate the Wnt pathway [174]. They found that, induced by increased lncRNA HOTAIR in OA, histone methyltransferase PRC2 raised the level of H3K27me3 and reduced the expression of Wnt inhibitory factor 1 (WIF-1), which activated the Wnt signaling pathway, increased the production of matrix-degrading proteins, and accelerated the progression of OA [174]. However, not all histone modifications positively regulate the Wnt pathway. Monteagudo's team reported the ability of histone methyltransferase DOT1L to block Wnt pathway overactivation to maintain cartilage homeostasis by inhibiting the function of SIRT1 after activation of Wnt signaling [175]. In contrast, decreased DOT1L activity in OA results in the loss of this inhibitory capacity, which leads to Wnt overactivation, inducing the expression of catabolic genes and accelerating OA progression [175,176].

It has been demonstrated that the development of OA is linked to persistent, low-grade inflammation. Inflammatory factors cause the production of catabolic factors MMPs and ADAMTS to be upregulated, which disturbs cartilage homeostasis and accelerates the progression of OA [177,178]. According to a study by Dai et al., the increased expression of catabolic genes induced by pro-inflammatory cytokines is mediated by histone acetylation [179]. Histone acetylation readers Brd3 and Brd4 are recruited to the MMP1, 3, 13, and ADAMTS4 promoters by IL-1β and TNF-α. They recognize histone acetylation modifications and mediate transcriptional elongation of DNA, thus facilitating the expression of these catabolic genes [179]. Histone modifications also mediate inflammatory damage in chondrocytes, a mechanism reported by Ai et al. [180]. IL-1β-induced upregulation of HMGA1 expression increases the expression of histone H3K27 demethylase JMJD3, which in turn mediates H3K27 demethylation of the ZEB1 promoter to promote ZEB1 transcription, exacerbating inflammatory damage and oxidative stress in chondrocytes [180].

Senescent chondrocytes secrete senescence-associated secretory phenotype (SASP), including matrix-degrading protease MMPs, ADAMTSs, and inflammatory cytokines IL-1, TNF-α, which induce OA inflammation and ECM degradation, disrupt cartilage homeostasis, and ultimately result in the development of OA [181]. Ji et al. discovered the mechanism of histone modification to promote chondrocyte senescence and OA progression [182]. They found that lncRNA ELDR mediates the recruitment of KAT6a and hnRNPL to the IHH promoter, enhancing H3K9ac and H3K4me3 modifications. This chromatin remodeling facilitates the binding of the transcription factor NRF1 to the IHH promoter, which initiates IHH expression, triggers hedgehog signaling, and accelerates chondrocyte senescence [182]. As a result, there is increased expression of SASPs and catabolism, which promotes OA progression.

Emerging evidence suggests that metabolic reprogramming in OA not only affects energy supply but also shapes the epigenetic landscape of chondrocytes, thereby influencing gene expression and cartilage homeostasis. Chen et al. reported a mechanism by which altered glucose metabolism in OA affects histone modifications and thus regulates matrix metabolism [183]. They found that the action of ATP citrate lyase (ACLY) was enhanced in OA chondrocytes, leading to an increase in acetyl-CoA levels. This raised the level of H3K9ac and H3K27ac in the promoter regions of catabolic factors, thereby enhancing their expression and accelerating matrix degradation and OA [183]. Similarly, Xia et al. reported that OA mouse models exhibit abnormally elevated glycolysis and increased histone lactylation levels [65]. They further demonstrated that LDHA-induced H3K18 lactylation enhances TPI1 transcription, which upregulates catabolic factors such as MMP13, disrupting cartilage homeostasis and accelerating cartilage loss. These findings reveal a critical link between metabolic intermediates and chromatin regulation in OA pathogenesis. Wang et al. found that the upregulation of histone methyltransferase EZH2 was associated with OA, and that inhibiting EZH2 alleviated cartilage deterioration and OA symptoms [184]. Mechanistically, EZH2 suppresses miR-138 expression by elevating H3K27me3 levels in its promoter region, thereby upregulating its target gene SDC1, promoting the production of catabolic factors, and disrupting cartilage homeostasis [184].

3.2.1.2. Histone modifications inhibit anabolism

Major components of ECM include type II collagen and aggrecan, whose gene expression is controlled by transcription factor SOX9, a major cartilage regulatory factor [185]. Nham's team and UKITA's team have reported on the epigenetic modification mechanism by which histone modifications regulate the expression of ACAN, COL2A1, and SOX9, thereby affecting matrix synthesis. Nham et al. found that HDAC10 and sirtuin6 controlled the activity of E1 and E2 enhancers to regulate the expression of ACAN and COL2A1 [186]. UKITA et al. discovered that the levels of H3K9 methylation were decreased in OA cartilage, which decreased SOX9 and COL2A1 expression and negatively regulated cartilage homeostasis [187]. Lian et al. reported a non-classical mechanism for the regulation of anabolic genes through synergistic interactions between histone modifications [188]. It was discovered that OA chondrocytes had an abnormality of elevated levels of both H3K27me2/3 demethylase UTX expression and H3K27me3 modification. Furthermore, they observed that H3K27me3 enrichment at the Sox9 promoter was instead facilitated by enforced UTX expression [188]. Further exploration of the mechanism revealed that UTX targets H3K27me3 at the Eed and Suz12 promoters, the core components of histone methyltransferase PRC2, increasing their expression. The increased expression of PRC2 results in heightened H3K27me3 levels in the Igf2 promoter region, reduced Igf2 expression, leading to diminished ECM synthesis, and disrupted cartilage homeostasis, thereby promoting OA progression [188].

The dual regulatory impacts of histone methylation on anabolism and catabolism were described by Shao et al. and Jin et al. [189]. Shao et al. found that histone methyltransferase NSD1 expression was down-regulated in OA. Furthermore, in NSD1 knockout mice, the expression of SOX9 and COL2A1 was reduced, while the expression of MMP3, MMP13, and ADAMTS5 was elevated [189]. Mechanistically, down-regulation of NSD1 leads to decreased Osr2 H3K36 methylation levels and decreased Osr2 expression, which inhibits anabolism and enhances catabolism, disrupting cartilage homeostasis and facilitating OA progression [189]. Jin et al. observed an upregulation of histone demethylase JMJD3 in chondrocytes following the application of abnormal stress [190]. JMJD3 promotes NR4A1 expression by reducing H3K27me3 levels at its promoter. This results in the downregulation of COLII and SOX9, an increase in MMP13 and IL-1β expression, and subsequently promotes OA [190].

3.2.2. Histone modifications in other OA pathological processes

Beyond disrupted cartilage metabolism, OA involves chondrocyte death, synovial inflammation and subchondral bone formation [191]. While most studies on histone modifications mainly focus on cartilage homeostasis, current evidence for their roles in these additional processes is limited but gradually emerging.

3.2.2.1. Chondrocyte death

Chondrocyte death, including apoptosis, pyroptosis, and ferroptosis, is a hallmark of OA and accelerates cartilage degradation [192]. These processes amplify matrix breakdown and inflammatory responses, fueling disease progression [193]. Recent studies indicate that histone modifications regulate chondrocyte death pathways by controlling gene transcription, adding a novel epigenetic dimension to OA pathogenesis.

Apoptosis, the earliest recognized form of programmed cell death (PCD), is a tightly regulated, non-inflammatory process that eliminates damaged or dysfunctional cells [194]. Chondrocyte apoptosis, widely observed in OA patients and animal models, is closely associated with disease severity and contributes to extracellular matrix degradation and cartilage calcification [193]. Emerging evidence suggests that histone modifications influence key apoptotic signaling pathways. Studies have shown that HDACs play an important role in regulating chondrocyte apoptosis. Wang et al. found that HDAC2 lowers H3K27ac at the miR-503-5p promoter, thereby reducing its expression and increasing Caspase-3 and Bax, which promotes chondrocyte apoptosis [195]. Li et al. reported that HDAC4 is downregulated in human and rat OA cartilage. HDAC4 suppresses ATF4 and CHOP expression, reducing oxidative stress and inhibiting chondrocyte apoptosis [196]. Using the Acan-Cre^ERT2; HDAC4^fl/fl genetic mouse model, their team further confirmed this finding, highlighting the potential of HDAC4 as a therapeutic target for OA [197]. The role of JMJD3 in regulating chondrocyte apoptosis has also been revealed. Jin et al. found that JMJD3 promotes NR4A1 expression by H3K27me3 demethylation at its promoter, thereby influencing chondrocyte apoptosis and cartilage degeneration [190]. Similarly, Huang et al. reported that JMJD3 activates the NOTCH1 expression, which upregulates Bax and Caspase-3 while suppressing Bcl-2 expression, promoting IL-1β-induced chondrocyte apoptosis [198].

Pyroptosis is an inflammation-related PCD, triggered by the assembly of inflammasomes and activation of specific caspases, including caspase-1 and caspase-4/5/11 [199]. Research indicates that chondrocyte pyroptosis elevates the release of proteolytic enzymes and inflammatory cytokines, which in turn exacerbate inflammation and cartilage degradation in OA [200]. However, current research on the regulation of chondrocyte pyroptosis by histone modifications remains very limited. Chen et al. were the first to uncover the mechanism by which EZH2 regulates chondrocyte pyroptosis [201]. Their study showed that EZH2 expression positively correlates with the severity of pyroptosis and the progression of OA. EZH2 mediates epigenetic silencing of miR-142-3p through H3K27me3, resulting in increased HMGB1 expression, which aggravates endoplasmic reticulum stress-induced chondrocyte pyroptosis and accelerates OA development.

Ferroptosis, a newly identified PCD characterized by iron-dependent lipid peroxidation, is driven by elevated intracellular free iron, glutathione depletion, and GPX4 inactivation, resulting in the membrane lipid peroxide accumulation and loss of cellular integrity [202]. Ferroptosis is increasingly recognized as an important contributor to OA pathogenesis. During disease progression, iron accumulates in cartilage while antioxidant defenses such as GPX4 and GSH decline, resulting in iron overload and redox imbalance [203]. This disruption induces chondrocyte ferroptosis, which enhances MMP13 expression, suppresses COL2, and ultimately accelerates cartilage degradation and OA progression [204]. Currently, increasing attention has been given to the role of histone modifications in regulating chondrocyte ferroptosis. Wang et al. reported that NSD1 promotes H3K36me2 modification and upregulates SOX9 expression. This suppresses ACSL4 transcription, inhibits chondrocyte ferroptosis, and contributes to cartilage integrity [205]. Zhang et al. reported that lactate dehydrogenase B (LDHB) and H3K18 lactylation increase during OA. LDHB elevates H3K18la and upregulates ACSL4 expression, leading to chondrocyte ferroptosis and accelerated OA progression [206].

3.2.2.2. Synovial inflammation and subchondral bone remodeling

Synovial inflammation is a key pathological feature in the development of OA. Although OA has traditionally been regarded as a cartilage degenerative disease, increasing evidence indicates that synovial inflammation contributes to OA onset and progression [207]. It is characterized by synovial hyperplasia, angiogenesis, and infiltration of immune cells such as macrophages, T cells, and B cells [208]. These cells release pro-inflammatory cytokines and chemokines, which amplify inflammation, induce MMPs and ADAMTS, and accelerate cartilage matrix degradation [209]. Evidence remains limited, but existing studies underscore the role and therapeutic potential of histone modifications in regulating synovial inflammation in OA. The histone deacetylase inhibitor trichostatin A was shown to reduce synovial inflammation and prevent cartilage damage in an arthritis model [210]. Lin et al. demonstrated that synovial macrophages in OA mice shift toward M1 polarization and promote cartilage degradation via exosome secretion. These exosomes transfer miR-350-3p to chondrocytes, suppressing NSD1 expression and reducing H3K36 methylation, which consequently elevates the expression of MMP13, P16, and P21 [211].

Subchondral bone remodeling imbalance is a key pathological feature of OA. During disease progression, the subchondral bone undergoes increased bone density, trabecular thickening, and osteophyte formation [212]. These alterations not only disrupt joint biomechanics but also accelerate cartilage degeneration through bone–cartilage crosstalk [213,214]. Recent studies suggest that histone modifications may play a role in subchondral bone remodeling, offering potential new targets for OA therapy. Animal studies have demonstrated that histone methylation significantly influences subchondral bone remodeling in OA. Intra-articular overexpression of KDM6A in mice led to pronounced osteophyte formation and increased osteophyte volume around the injured knee, whereas KDM6A knockout alleviated OA features, including cartilage loss, osteophyte development, and subchondral trabecular bone loss [188,215]. Likewise, curcumenol treatment upregulated KDM6B and reduced H3K27me3 levels, thereby improving cartilage integrity and preventing subchondral bone loss in knee OA models [216]. Lian et al. demonstrated that subchondral mesenchymal stem cells (SMSCs) from OA knees possess higher osteogenic and chondrogenic potential than bone marrow–derived MSCs, contributing to subchondral bone over-mineralization and osteophyte formation. This mechanism is mediated by microRNA-29a-induced downregulation of HDAC4, underscoring the role of histone acetylation in subchondral bone remodeling during OA.

In summary, histone modifications play a central role in OA progression by enhancing catabolic activity, suppressing anabolic pathways, and disturbing cartilage homeostasis. Beyond these well-established mechanisms, emerging evidence suggests their involvement in chondrocyte death, synovial inflammation, and subchondral bone remodeling. However, research in these areas remains scarce, leaving their precise roles largely undefined. Future studies should aim to elucidate these mechanisms and explore the therapeutic potential of targeting histone modifications to develop epigenetic-based strategies for OA management.

3.3. Histone modifications and intervertebral disc degeneration

Intervertebral disc degeneration, a spinal degenerative disease characterized by progressive deterioration of the structure and function of the nucleus pulposus (NP), annulus fibrosus (AF), and cartilage endplate (CEP), is the leading cause of lower back pain (LBP), which leads to a serious disease burden worldwide [9,[217], [218], [219]]. Relevant research has demonstrated that histone modifications are crucial to the occurrence and development of IDD [220,221] (Fig. 4).

Fig. 4.

Histone modifications contribute to nucleus pulposus and cartilage endplate degeneration in intervertebral disc disease. They accelerate ECM degradation, induce nucleus pulposus cell senescence and death, and promote cartilage endplate degeneration.

3.3.1. Histone modifications in extracellular matrix homeostasis

The extracellular matrix (ECM) refers to the non-cellular components within tissues, which offer structural support to cells and establish a microenvironment favorable for cell survival while regulating a range of cellular physiological functions [222]. The main components of the intervertebral disc's ECM include proteoglycans, collagen, and non-collagenous proteins, playing crucial roles in regulating cell phenotype, absorbing water, facilitating nutrient diffusion, and maintaining mechanical microenvironments [223]. Enhanced catabolism and suppressed anabolism lead to ECM degradation, which results in reduced hydration and elasticity of the nucleus pulposus, further leading to dehydration and fibrosis, and significantly disrupting the flexibility and load-bearing capacity of the intervertebral disc [224,225]. Moreover, alterations in ECM components induce changes in the mechanical microenvironment and disrupt cell–cell interactions, leading to phenotypic changes in nucleus pulposus cells (NPCs) and impairing their normal functions [225]. Consequently, ECM degradation is closely associated with IDD. Furthermore, histone modification participates in ECM degradation and IDD processes by influencing gene expression connected to ECM synthesis and decomposition.

ADAMTS is one of the major ECM-degrading enzymes that recognize ECM components through its thrombospondin motifs [226]. It is a typical marker of catabolism, and its upregulation in expression and activity is often associated with ECM metabolism dysregulation [227]. Wang et al. discovered that histone methylation modifications directly regulate the expression of ADAMTS-5, mediating ECM degradation. They found a reduction in RREB1 expression in degenerated NPCs, resulting in increased levels of transcriptionally activating modifications H3K4me2 and H3K4me3 on the ADAMTS-5 promoter region, and decreased levels of inhibitory modifications H3K9me3 and H3K27me3, leading to increased expression of ADAMTS-5, ECM degradation, and promotion of IDD [228].

By increasing the synthesis of associated matrix-degrading enzymes, the NF-κB signaling pathway facilitates ECM degradation [229]. Related research has indicated that SIRT6 participates in ECM metabolism by regulating the NF-κB signaling pathway. It has been demonstrated that the histone deacetylase SIRT6 controls the expression of NF-κB related genes by mediating H3K9 deacetylation, which regulates the signal transmission of the NF-κB pathway at the gene level [230]. Kang et al. observed decreased SIRT6 expression in degenerated NP tissues and IL-1β-induced NPCs, and demonstrated that silencing SIRT6 induces ECM degradation in human NP cells [231]. Mechanistically, the downregulation of SIRT6 expression weakens its inhibitory effect on the NF-κB pathway, leading to increased production of MMP3 and MMP13, and decreased production of Collagen II and Aggrecan, thereby promoting ECM degradation and IDD [231].

Oxidative stress refers to cellular oxidative damage caused by the excessive production and insufficient clearance of reactive oxygen species (ROS) [232]. Studies have demonstrated that excess ROS induces the upregulation of catabolism markers such as MMPs and ADAMTS through the initiation of related signaling pathways, such as MAPK and NF-κB pathway, while the expression of anabolism markers such as Coll Ⅱ and Aggrecan is downregulated, leading to extracellular matrix (ECM) degradation and promoting IDD [233]. Relevant research indicates the involvement of histone modifications in these molecular mechanisms. Xu et al. found that ROS induced by H₂O₂ activates the p38/MAPK pathway, promoting the phosphorylation of histone methyltransferase KMT2D and inhibiting its ubiquitination degradation, thereby increasing its expression [234]. Upregulated KMT2D subsequently increases the expression of degradation markers MMPs by enhancing the H3K4me1 levels in the enhancer regions of Mmp3, Mmp9, and Mmp13, thereby leading to ECM degradation and promoting IDD [234].

3.3.2. Histone modifications in nucleus pulposus cell senescence

Intervertebral disc degeneration, an age-related affliction, is intricately intertwined with cellular senescence. Senescent nucleus pulposus cells unleash SASPs, unleashing a cascade of inflammatory cytokines and degradation factors such as MMPs and ADAMTS, thereby fueling extracellular matrix (ECM) degradation and accelerating IDD progression [235]. Presently, related research underscores the close interplay between NPC senescence and alterations in histone modifications, revealing that these modifications serve as pivotal epigenetic regulators linking inflammation, oxidative stress, autophagy, and RNA methylation to NPC senescence.

During IDD, inflammation and NPC senescence of are highly intertwined. Chronic inflammatory cytokines, such as TNF-α and IL-1β, activate inflammatory signaling pathways like NF-κB, inducing cell cycle arrest and the expression of SASPs, thereby promoting inflammatory senescence [236]. Inflammatory signaling pathways, particularly MAPK and NF-κB, exert pivotal roles in orchestrating NPC senescence within the inflammatory microenvironment [[237], [238], [239]]. Zhou et al. observed EZH2 promotes NPC senescence by enhancing MAPK-mediated inflammatory responses. Further, mechanistic insights reveal that EZH2 suppresses miR-129-5p expression by meditating H3K27me3, thereby alleviating its inhibitory effect on MAPK1 expression [240]. As discussed earlier, SIRT6 mitigates NF-κB target gene expression through histone deacetylation, effectively halting its signaling cascade [230]. Xie et al. documented a decline in SIRT6 protein expression and an increase in NF-κB p65 protein expression, accompanied by heightened H3K9ac levels in TNF-α-induced NPCs. SIRT6 downregulation activates the NF-κB pathway, culminating in NPC inflammatory senescence [241]. The cGAS/STING pathway mediates the inflammatory microenvironment by activating NF-κB signaling, driving local inflammation and SASPs secretion [242]. Zheng et al. reported that EZH2 regulates the cGAS/STING pathway via H3K27me3 and suppresses inflammatory senescence. In contrast, the downregulation of EZH2 in degenerated NP tissues may lift the transcriptional repression of STING, resulting in activation of the cGAS/STING signaling pathway and the induction of inflammatory responses [243].

NADPH oxidase 4 (NOX4)-associated oxidative stress emerges as one of the driving factors of NPC senescence [244]. Liu et al. noted diminished EZH2 expression and heightened NOX4 expression in degenerated rat NP [245]. EZH2 downregulation precipitates NPC senescence, as reduced H3K27me3 levels in the NOX4 promoter region contribute to elevated NOX4 expression. Moreover, NOX4 overexpression suppresses EZH2 expression via Wnt/β-catenin, forming a positive feedback [245].

Emerging evidence underscores the role of a synergistic interplay between histone modifications and m6A methylation in driving NPC senescence and accelerating IDD. Li et al. and Wu et al. reported the synergistic interaction between m6A methyltransferases WTAP and METTL3 and histone methylation. Li et al. discovered that in TNF-α-induced senescent NP cells, the expression of lysine demethylase KDM5a markedly decreases, leading to heightened histone H3K4me3 epigenetic modification in the promoter region of m6A methyltransferase WTAP, thereby promoting its transcription [246]. The heightened expression of WTAP significantly boosts the m6A modification of lncRNA NORAD, amplifying its degradation. Furthermore, via the NORAD/PUMILIO/E2F3 axis, it reduces the expression of E2F3, thereby fostering NPC senescence and IDD [246]. Wu et al. demonstrated that KMT2A-mediated H3K4me3 upregulates METTL3, resulting in m6A methylation of ATG4a mRNA and decreased transcript stability. This impairs autophagy and triggers the GATA4-driven NPC senescence, accelerating IVDD progression [247]. Li et al. revealed an upregulation in m6A demethylase ALKBH5 expression in senescent NP cells induced by TNF-α [248]. Subsequent investigations unveiled that this alteration stemmed from heightened lysine demethylase KDM4a expression, resulting in diminished H3K9me3 levels in the ALKBH5 promoter region, thereby facilitating its expression. ALKBH5 boosts DNMT3B expression through its m6A demethylation function, subsequently leading to DNA methylation-mediated inhibition of E4F1 transcription, ultimately fostering NPC senescence and IDD [248].

3.3.3. Histone modifications in nucleus pulposus cell death

NPC death, including apoptosis, pyroptosis, and ferroptosis, plays a pivotal role in the initiation and progression of IDD [199]. Dysregulation of these pathways contributes to the depletion of disc cells, degradation of extracellular matrix components, and amplification of local inflammatory responses [249]. Notably, growing evidence suggests that histone modifications promote IDD by disturbing the homeostasis of programmed cell death in NPCs.

Apoptosis, a classic form of cell death, contributes to progressive cell loss, cytokine release, and ECM degradation in IDD, thereby accelerating structural deterioration and functional impairment of the intervertebral disc [249]. Recent studies have highlighted the critical role of histone modifications in modulating NPC apoptosis. Apoptosis signal-regulating kinase 1 (ASK1) is a key apoptosis-related kinase that triggers apoptosis via activation of mitochondria-dependent caspases [250]. Recent studies indicate that ASK1 promotes nucleus pulposus cell apoptosis and contributes to IDD [251,252]. Lin et al. found that HDAC4 promotes the expression of ASK1 through KLF5, thereby exacerbating IDD [252]. In addition, Zheng et al. found that HDAC/H3K27ac-mediated transcriptional activation of NDUFA3 inhibits nucleus pulposus cell apoptosis, improves mitochondrial function, and protects against high glucose-induced damage [253]. While these studies highlight the significant roles of histone acetylation, Liu et al. found that KMT2D promotes the expression of miR-133a-5p through H3K4me2. This facilitates miR-133a-5p binding to PFKFB2, resulting in transcriptional repression of PFKFB2 and subsequently enhancing apoptosis and inhibiting proliferation of NPCs [254].

Pyroptosis, an inflammatory mode of cell death, is increasingly recognized as a key driver of IDD progression [255]. Pyroptosis of NPCs results not only in cell loss but also in the release of pro-inflammatory cytokines, which promote ECM degradation and disrupt tissue homeostasis [255]. Yao et al. demonstrated that EZH2 mediates transcriptional repression of DKK1 via H3K27me3, which triggers NPCs pyroptosis through activation of NLRP3 and NAIP/NLRC4 inflammasomes, contributing to the progression of IDD [256].

Ferroptosis, an iron-dependent form of cell death driven by lipid peroxidation, has recently attracted attention for its potential role in IDD. Iron overload and ferroptosis upregulate MMP3 and MMP13 while downregulating type II collagen, thereby accelerating ECM degradation and structural deterioration of the intervertebral disc [257,258]. Recent studies have shown that histone modifications within the altered metabolic microenvironment play a crucial role in regulating ferroptosis in NPCs during IDD. Wu et al. found that GLS1-mediated glutamine metabolism enhances acetyl-CoA production and increases H3K9ac and H3K27ac at the NFS1 promoter, leading to NFS1 upregulation. Elevated NFS1 levels preserve Fe²⁺ homeostasis and prevent oxidative stress–induced matrix degradation, ferroptosis, and senescence in NPCs [259]. Sun et al. reported that enhanced glycolysis and increased lactate production in degenerated discs induce ACSL4 expression by elevating H3K18 lactylation and promote ACSL4 protein lactylation. These metabolic and epigenetic alterations activate NPCs ferroptosis and accelerate IDD [260]. These findings highlight a mechanistic axis in which metabolic reprogramming shapes the epigenetic landscape to regulate iron-dependent cell death, revealing a critical link between altered energy metabolism, histone modification, and ferroptosis in the pathogenesis of IDD.

3.3.4. Histone modifications in cartilage endplate and annulus fibrosus degeneration

The cartilage endplate (CEP) is essential for intervertebral disc health, facilitating nutrient exchange and providing biomechanical support [261]. CEP degeneration disrupts nutrient supply, making the disc susceptible to stress and injury, and is often linked to the breakdown of ECM components [[262], [263], [264]]. Jiang et al. observed elevated expression of histone methyltransferase EZH2 in degenerated rat endplate chondrocytes (EPCs) and patients' CEP. Notably, EZH2 overexpression promotes ADTAMTS5 and MMP13 expression, and inhibits Coll Ⅱ and Aggrecan expression, consequently driving CEP and IDD [265]. Mechanistically, EZH2 suppresses Sox-9 transcription by inducing H3K27me3 modification at its promoter region, resulting in the downregulation of Sox-9 [265].

The annulus fibrosus (AF) is a critical component of the intervertebral disc, composed of concentric lamellae of collagen fibers that provide tensile strength and maintain disc integrity [217]. During IDD, AF undergoes structural and biochemical alterations, including disorganization of collagen fibers, decreased proteoglycan content, and increased cell apoptosis, which collectively compromise the mechanical properties and contribute to disc herniation and chronic low back pain [266]. Recent studies have revealed the crucial role of epigenetic regulation in AF degeneration. For instance, DNA methylation alterations have been identified in AF during IDD, while noncoding RNAs such as lncRNA GAS5 and miR-221-3p influence AF cell osteogenic differentiation and apoptosis [267,268]. In contrast, research on histone modifications in AF degeneration remains extremely scarce, and the underlying mechanisms are largely unexplored. Recently, Ramteke et al. reported that SIRT6 conditional knockout mice develop accelerated AF degeneration, characterized by the loss of the NP-AF boundary, formation of AF fissures, and degradation of organized collagen matrix, highlighting the potential role of histone modifications in AF pathology [269].

In summary, histone modifications are emerging as key regulators in IDD, influencing ECM degradation, NPC senescence, multiple forms of regulated cell death, and potentially cartilage endplate and annulus fibrosus degeneration. Nevertheless, current research on histone modifications remains largely focused on nucleus pulposus, with limited understanding of their roles in other disc components. Future studies should aim to comprehensively elucidate these mechanisms and explore histone-modifying enzymes as promising therapeutic targets for IDD.

4. Histone-modifying enzymes provide the potential therapeutic targets for degenerative skeletal diseases

In light of the crucial roles of histone modifications and their modifying enzymes in degenerative skeletal diseases, the exploration of small-molecule modulators including inhibitors and activators targeting these histone-modifying enzymes holds promise for treatment. Currently, small-molecule inhibitors targeting histone-modifying enzymes have made significant advances, especially in oncology, where they have shown promising therapeutic effects. However, their clinical application in degenerative skeletal diseases remains largely unexplored, with no clinical trials reported so far. Numerous preclinical studies have demonstrated their potential to slow disease progression, highlighting the urgent need for further translational research and early-phase clinical trials (Table 4).

Table 4.

Small-molecule modulators of histone modification and their effects on degenerative skeletal diseases.

Classification	Small molecules	Targets	Diseases	Experimental model/subjects	Pharmacological functions	References
HMT inhibitors	Tazemetostat	EZH2	osteoarthritis	IL-1β-induced chondrocytes; meniscectomy mice	reduces inflammation and catabolism; reduces cartilage degradation and joint pain	[274]
	DZNep	EZH2	osteoarthritis	IL-1β-induced chondrocytes	reduces MMPs expression	[275]
HDM inhibitors	GSK2879552	LSD1	osteoporosis	RANKL-treated BMMs; OVX mice	inhibits RANKL-induced osteoclast differentiation; prevents cortical bone loss	[278]
	KDM5A-IN-1	KDM5C	osteoporosis	RANKL-treated BMMs	inhibits RANKL-induced osteoclast differentiation	[279]
	GSK-J4	KDM6A	osteoarthritis	DMM mice	reduces articular cartilage loss and synovial hyperplasia	[215]
	GSK-J4	JMJD3	osteoarthritis	IL-1β-induced chondrocytes; DMM mice	inhibits inflammation and catabolism; prevents cartilage degradation	[280]
	Daminozide	KDM2/7	osteoarthritis	C28/I2 cells; DMM mice	increases anabolism; reduces cartilage degradation and osteophyte formation	[281]
HAT inhibitors	A-485	p300/CBP	osteoporosis	RANKL-treated BMMs; OVX mice	inhibits RANKL-induced osteoclast differentiation; alleviates bone loss	[285]
	Salidroside	PCAF	osteoarthritis	TNF-α-induced chondrocytes; DMM mice	inhibits ECM degradation, inflammation, and apoptosis; alleviates cartilage destruction and synovitis	[286]
HDAC inhibitors	Vorinostat	class I and II HDAC	osteoarthritis	IL-1β-induced chondrocytes	inhibits the expression of catabolic markers	[[290], [291], [292]]
	Vorinostat	class I and II HDAC	osteoporosis	RANKL-treated BMMs; OVX mice	inhibits RANKL-induced osteoclast differentiation; ameliorates bone loss	[293]
	Panobinostat	HDAC	osteoarthritis	ACLT rats	attenuates subchondral bone remodeling, cartilage degradation, and articular cartilage hypertrophy; alleviates pain	[295]
	Panobinostat	HDAC	osteoarthritis	IL-1β-induced chondrocytes; DMM mice	inhibits catabolism; attenuates cartilage degradation, subchondral bone remodeling, and synovitis	[296]
	Chidamide	benzamide-type HDAC	osteoporosis	adipogenic medium-induced BM-MSCs	suppresses adipogenic differentiation of BM-MSCs	[298]
	Chidamide	benzamide-type HDAC	osteoarthritis	ACLT rats	reduces cartilage degradation, osteophyte formation, and pain	[314]
	Tubastatin A	HDAC6	osteoarthritis	DMM mice	inhibits cartilage degradation and synovial tissue thickening; alleviates hyperalgesia	[299]
	Tubastatin A	HDAC6	osteoarthritis	TBHP-induced chondrocytes; DMM mice	reduces apoptosis, oxidative stress, and ECM degradation, and enhances autophagy in chondrocytes; inhibits cartilage degradation and aberrance	[300]
	ACY-1215	HDAC6	osteoarthritis	IL-1β-induced chondrocytes	decreases the expression of catabolic markers and promotes the expression of COL2A1	[301]
SIRT1 activator	Resveratrol	SIRT1	osteoporosis	H2O2-treated osteoblasts; OVX mice	enhances proliferation, differentiation, and suppresses apoptosis of H2O2-treated osteoblasts; ameliorates bone loss and promotes osteogenesis	[327]
	Resveratrol	SIRT1	osteoporosis	postmenopausal women	slows bone loss in the lumbar spine and femoral neck	[328]
	Resveratrol	SIRT1	osteoarthritis	DMM mice	inhibits cartilage degradation, subchondral bone remodeling	[330]
	Resveratrol	SIRT1	osteoarthritis	postmenopausal women	reduce chronic pain in age-related osteoarthritis and improve menopause-related quality of life in postmenopausal women	[331]
	Resveratrol	SIRT1	intervertebral disc degeneration	NPCs	inhibited NPCs apoptosis and promoted ECM synthesis	[333]

Note: BMMs: bone marrow-derived macrophages; OVX: ovariectomy; DMM: destabilized medial meniscus; ACLT: anterior cruciate ligament transection.

4.1. HMT inhibitors

Numerous inhibitors targeting histone methyltransferases (HMTs) have been identified and developed, with EZH2 inhibitors demonstrating significant advancements in drug development. In particular, in January 2020, the FDA approved Tazemetostat, the first EZH2 inhibitor in the world that was co-developed by Epizyme and Eisai, for marketing to treat epithelioid sarcoma [270]. In addition, Valemetostat Tosilate, a dual EZH1/2 inhibitor, developed by Daiichi Sankyo Company in Japan, received marketing approval from the MHLW in September 2022 for addressing relapsed/refractory adult T-cell leukemia [271].

Multiple investigations have revealed elevated EZH2 expression levels in degenerative skeletal diseases (OP, OA, IDD), strongly associated with disease progression [265,272,273]. Utilizing siRNA or shRNA to knock down EZH2 has shown promising results in ameliorating tissue cell degeneration, impeding disease advancement, and exerting a protective effect [265,272,273]. This indicates that treating degenerative skeletal diseases with EZH2 inhibitors is feasible. Allas et al. conducted a preclinical study utilizing Tazemetostat to target EZH2 in OA treatment [274]. They observed that Tazemetostat application reduced the expression of catabolic genes in IL-1β-induced chondrocytes, decreased ECM degradation, attenuated cartilage degradation, and enhanced motor function in OA mice [274]. Aury-Landas et al. also reported that DZNep, an EZH2 inhibitor, exhibits chondroprotective effects by downregulating the expression of IL-1β-induced cartilage matrix degradation genes [275]. Extensive clinical studies are necessary to determine the safety and effectiveness of EZH2 inhibitors in the treatment of OA, despite the promising therapeutic utility of these in vivo and in vitro research.

4.2. HDM inhibitors

Histone demethylase (HDM) inhibitors can be categorized into two main groups: those aimed at LSD1/KDM1A and those targeting the JMJC subfamily. The former group of inhibitors has advanced more rapidly than the latter group, with numerous drugs currently progressing through clinical phases II and III, primarily for the treatment of hematological, lymphatic disorders, and neoplasms [276,277]. While clinical trials evaluating the efficacy of HDM inhibitors in treating degenerative skeletal diseases are yet to be conducted, a significant number of preclinical investigations have highlighted their promising therapeutic potential in this context.

GSK2879552, an LSD1 inhibitor, has demonstrated the efficacy of reducing RANKL-induced osteoclast differentiation, while also exhibiting the capacity to partially ameliorate cortical bone loss induced by OVX in murine models of OP [278]. Liu et al. investigated the utilization of the pan-KDM5 inhibitor (KDM5A-IN-1), which acts by targeting KDM 5C, thus impairing energy metabolism and curtailing osteoclastogenesis in both murine and human subjects [279]. Additionally, GSK-J4, acting as an inhibitor of the KDM6 family, functions dually to inhibit UTX/KDM6A and JMJD3/KDM6B. Relevant studies have highlighted its prospective application in mitigating OA. Lian et al. administered intra-articular injections of GSK-J4 to inhibit KDM6A, observing significant attenuation of DMM-induced OA alterations, such as synovial hyperplasia and articular cartilage loss, in murine models [215]. Jun et al. noted upregulated JMJD3 expression in OA cartilage, and further demonstrated that GSK-J4 administration led to reduced production of catabolic enzymes, inhibited extracellular matrix degradation, and mitigated cartilage damage in murine models of OA [280]. Furthermore, Daminozide, a KDM2/7 subfamily inhibitor, has been reported to elevate H3K79 methylation and glycosaminoglycans levels in chondrocytes following intra-articular injection, thus conferring protective effects against cartilage damage in DMM mice [281]. Collectively, all of these investigations show that HDM inhibitors have the potential to be innovative treatment options for degenerative skeletal diseases.

4.3. HAT inhibitors

HAT inhibitors function by targeting the bromodomain and HAT domains of histone acetyltransferases (HATs) [282]. Their drug development is currently in the preclinical and clinical phase I/II stages [283]. CBP/p300 BRD inhibitors Inobrodib (CCS1477) and NEO2734 (EP31670) have progressed into Phase 1/2 clinical trials targeting tumor diseases, such as multiple myeloma and non-Hodgkin lymphoma [284]. Additionally, the KAT6 catalytic inhibitor PF-07248144 has progressed into Phase 1 clinical trials aimed at treating metastatic ER+/HER2– breast cancer and other cancers [284]. Relevant preclinical studies have indicated the potential of HAT inhibitors in the treatment of degenerative skeletal diseases.

CBP/p300 inhibitor A-485 has shown promise in protecting against OP. According to Huo et al., A-485 reduced OVX-induced bone loss in the mouse model in vivo and suppressed osteoclast-specific gene expression and osteoclast differentiation in vitro [285]. Salidroside has been demonstrated to inhibit histone acetylation against OA. Chen et al. discovered that Salidroside inhibited ECM degradation, inflammation, and apoptosis in OA chondrocytes and alleviated OA symptoms in the mouse DMM model by pharmacologically blocking PCAF [286]. These studies highlight the application prospects of HAT inhibitors in degenerative skeletal diseases.

4.4. HDAC inhibitors

Since the first HDAC inhibitor, trichostatin A, was discovered more than three decades ago, a great deal of progress has been achieved in the drug development of HDAC inhibitors [287]. Five representative HDAC inhibitors, Vorinostat, belinostat, panobinostat, romidepsin, and chidamide, have been permitted for therapies for hematologic malignancies, with several others undergoing clinical trials in phases II and III [288]. An increasing amount of preclinical research emphasizes the potential therapeutic utility of HDAC inhibitors in degenerative skeletal diseases.

The effectiveness of vorinostat, panobinostat, and chidamide in treating degenerative skeletal diseases has been substantiated by extensive preclinical research. Vorinostat, FDA-approved in 2006 for cutaneous T-cell lymphoma treatment [289], has exhibited promising results in inhibiting degradation markers like MMP-13 in OA chondrocytes, while also reversing the downregulation of COL2A1 and ACAN expression [[290], [291], [292]]. Peng et al. evaluated Vorinostat's therapeutic effects on bone loss in OVX mice via intraperitoneal injection, finding reduced osteoclast count, alleviated estrogen deficiency-induced bone loss, and trabecular structural damage without notable toxicity [293]. Panobinostat, FDA-approved in February 2015 for relapsed/refractory multiple myeloma [294], significantly downregulated RUNX2 and MMP13 expression in chondrocytes, mitigated chondrocyte hypertrophy, and alleviated joint inflammation and cartilage degeneration in ACLT-induced osteoarthritic rats, thus delaying OA progression [295]. Ohzono et al. reduced cartilage, synovium, and subchondral bone degeneration in osteoarthritic mice through intraperitoneal administration, improving pain behavior [296]. Independently developed by Chipscreen Biosciences, chidamide obtained CFDA approval in December 2014 for relapsed/refractory peripheral T-cell lymphoma treatment [297]. Zhang et al. highlighted chidamide's inhibitory effect on BMSC adipogenic differentiation, suggesting therapeutic potential [298]. Additionally, HDAC6, upregulated in OA, promotes ECM degradation, contributing to disease progression [299]. Its inhibitors, Tubastatin A and ACY-1215, reduce inflammatory factors and MMP expression, inhibit ECM degradation, improve cartilage degeneration, and halt OA progression [[299], [300], [301]].

These studies indicate that HDAC inhibitors exhibit tissue-protective effects, making them promising candidates for the treatment of degenerative skeletal diseases. Nevertheless, further clinical trials are necessary to confirm their efficacy and safety.

4.5. Translational potential and challenges of histone-targeted therapeutics

Epigenetic modulation through histone modification has emerged as a promising therapeutic approach for degenerative skeletal diseases. As outlined above, small-molecule inhibitors targeting histone-modifying enzymes have demonstrated compelling preclinical efficacy. These small-molecule inhibitors have been demonstrated to regulate critical pathological processes underlying disease progression, including inflammation, extracellular matrix degradation, and apoptosis. Importantly, they have not only reversed disease phenotypes in vitro but also improved tissue integrity and function in relevant animal models, highlighting their therapeutic value. In addition, their extensive exploration and application in the oncology field have provided valuable insights for their potential translation to orthopedic disorders. Despite these encouraging preclinical findings, substantial challenges still hinder the clinical translation of histone-modifying enzyme inhibitors for degenerative skeletal diseases.

4.5.1. Challenges in clinical translation

Suboptimal pharmacokinetic characteristics remain a major obstacle to the clinical development of histone-modifying enzyme inhibitors. A substantial proportion of histone-modifying agents exhibit poor aqueous solubility, limited drug permeability, low oral bioavailability, rapid systemic clearance, and short plasma half-lives [302,303]. These limitations not only impair the attainment of therapeutic concentrations and reduce pharmacological efficacy, but also elevate the risk of dose-limiting toxicities, thereby representing a significant barrier to broader clinical application [302]. For instance, vorinostat, one of the most widely studied HDAC inhibitors, is classified as a Biopharmaceutical Classification System (BCS) class IV drug owing to its poor aqueous solubility (0.2 mg/mL), low membrane permeability (log P = 1.9) and limited intestinal absorption (2 × 10⁻⁶ cm/s) [304]. Moreover, its rapid metabolic clearance further compromises systemic exposure and restricts tissue distribution [304]. Similarly, panobinostat exhibits low oral bioavailability (approximately 21.4 %) due to poor solubility and extensive first-pass metabolism [305,306]. It also shows considerable interindividual variability, with clearance varying by up to 74 % [305,306].

Moreover, side effects, toxicity, and resistance also limit the clinical application of histone-modifying enzyme inhibitors. Notably, HDAC inhibitors have been shown to induce a variety of adverse effects, including gastrointestinal reactions, myelosuppression, and cardiac toxicity [307,308]. These toxicities are primarily attributed to the lack of selectivity of most HDAC inhibitors, especially pan-HDAC inhibitors, which target multiple HDAC enzymes and modulate a broad spectrum of biological pathways [307]. Furthermore, the emergence of resistance is likely driven by the activation of alternative signaling pathways, such as the AKT and CDK pathways, which effectively negate the therapeutic efficacy [309].

To tackle the challenges outlined above, substantial efforts have been directed towards several critical areas.

4.5.2. Innovations in drug delivery systems

The development of innovative delivery approaches, including prodrugs, nanoparticle-based delivery systems, and proteolysis targeting chimeras (PROTACs), has garnered significant attention [302]. Prodrugs are inactive, bioreversible derivatives of active compounds that undergo enzymatic or chemical transformation to release the parent drug, including aqueous soluble, lipophilic, carrier-mediated, and dual function prodrug [310,311]. They are commonly employed to address the physicochemical limitations of drugs and improve their pharmacokinetic properties, including solubility, lipophilicity, bioavailability, half-life, release profiles, and targeted tissue delivery [311]. Du et al. designed and synthesized aqueous soluble prodrugs of SAHA and belinostat, which exhibited significantly enhanced water solubility (up to 600-fold higher than the parent compounds), while maintaining similar antiproliferative potency [312]. Additionally, the ester prodrug of butyric acid HDAC inhibitor (BA), AN-113, has been reported to show considerable improvements in physicochemical properties, pharmacokinetic parameters, efficacy, and safety [313].

Epigenetic drug delivery systems utilizing nanoparticles have emerged as a prominent area of research. A diverse array of nanocarriers, including lipid-based, polymeric-based, protein-based, and inorganic-based nanocarriers, have been explored for their potential in drug development [303]. Ye et al. utilized PLGA microcapsules to encapsulate chidamide, facilitating gradual and uninterrupted release for uniform intra-articular dispersion and sustained therapeutic effects [314]. Intra-articular injection of HDACi Microspheres exhibited extended retention time and alleviated pain in ACLT rats, safeguarding articular cartilage and showing excellent biocompatibility [314]. Additionally, Liu et al. designed mesoporous silica nanoparticles (MSN) loaded with 4-octyl itaconate (OI), which regulates epigenetic modifications such as histone acetylation and DNA demethylation [315]. This system effectively restored mitochondrial function and modulated the osteoimmune microenvironment, showing promising results in alleviating osteoporotic bone loss.

PROTACs are emerging as an innovative and promising strategy in drug development, particularly in regulating epigenetic processes. These bifunctional molecules induce selective degradation of target proteins by recruiting an E3 ubiquitin ligase and initiating ubiquitin proteasome system [316]. Compared to traditional inhibitors, PROTACs offer higher selectivity, minimize toxic side effects, and significantly reduce drug resistance by continuously degrading the target protein rather than merely inhibiting its activity [317]. For example, Suzuki et al. designed and synthesized a series of PROTAC compounds based on the HDAC8 inhibitor NCC-149-A, with pomalidomide as the E3 ligand, demonstrating excellent targeted degradation activity and pharmacological efficacy [318].

4.5.3. Development of isoform-selective small-molecule inhibitors

Given the toxic side effects associated with non-selective histone modification inhibitors, the development of isoform-selective inhibitors is critical for enhancing drug safety and facilitating their broader clinical application. Among these, the development of isoform-selective HDAC inhibitors has seen the most significant advancements. HDAC inhibitors are generally classified according to their selectivity for specific HDAC isoforms, such as HDAC1, HDAC2, and HDAC3 [319,320]. These selective inhibitors offer the potential to precisely modulate distinct epigenetic pathways, thereby enhancing therapeutic efficacy while minimizing off-target effects and associated adverse reactions [321]. For instance, Bresciani et al. reported selective class I HDAC inhibitors utilizing an ethylketone as the zinc-binding group (ZBG) [322]. They developed a novel series of HDAC3-selective inhibitors, among which compound 32 demonstrated potent and highly selective inhibition of HDAC3, exhibiting a 50-fold greater selectivity compared to the commonly used HDAC3 tool compound, RGFP9669. Furthermore, Sellmer et al. introduced Marbostat-100, a selective HDAC6 inhibitor derived from Tubastatin A, featuring a hydroxamic acid group linked to tetrahydro-β-carboline derivatives [323]. Their research demonstrated that Marbostat-100 is a potent HDAC6 inhibitor with strong selectivity and exhibits anti-inflammatory effects in in in vivo models.

4.5.4. Advancements in dual-/multi-target small-molecule inhibitors

Due to the complexity of diseases, single-target therapies often exhibit limited efficacy, along with challenges such as drug resistance and toxicity [320]. Consequently, the development of dual- and multi-target agents, capable of synergistically modulating disease-related networks, has emerged as a promising avenue in drug development. Among the multi-target strategies, multi-target HDAC inhibitors have garnered significant attention. Recent studies have shown that these compounds enhance therapeutic efficacy, reduce adverse effects, and decrease drug resistance, positioning them as a promising class of drugs in the field of epigenetic-based therapy [309].

Currently, a diverse array of multi-target HDAC inhibitors has been designed. Dual or multi-target drugs that simultaneously target HDAC and transcription factors have shown enhanced therapeutic effects. Tilekar et al. designed and synthesized a series of novel TZD-based naphthylidene derivatives aimed at simultaneously targeting HDAC and PPARγ for cancer therapy [324]. Among these, compound 7i emerged as the most potent dual-targeting agent, showing selective inhibition of HDAC4 (IC50 = 1.1 μM) and partial activation of PPARγ (EC50 = 0.245 μM). This compound demonstrated significant antiproliferative effects, induced apoptosis, and exhibited in vivo tumor regression, thus proving the efficacy of multi-target HDAC inhibitors in combating cancer. In the development of multi-target HDAC inhibitors, protein kinases have emerged as critical therapeutic targets, enabling precise modulation of cellular signaling pathways and contributing to improved therapeutic outcomes. Cheng et al. successfully designed and synthesized a series of novel dual HDAC/CDK inhibitors. Representative compounds 7c and 14a exhibited potent antiproliferative activity across multiple human cancer cell lines, demonstrated nanomolar inhibitory potency against both HDACs and CDKs. In vivo, compound 7c showed favorable pharmacokinetic properties and significant antitumor efficacy in the HCT116 xenograft model (TGI = 51.0 %) [325].

4.5.5. Identification and development of natural epigenetic modulators

Natural compounds, particularly those derived from traditional Chinese medicine, have attracted increasing interest as potential epigenetic modulators for the treatment of skeletal disorders. Among them, resveratrol, a natural activator of SIRT1, has been identified to exert an osteoprotective role in various studies [326]. Jiang et al. found that resveratrol promoted osteogenesis by activating SIRT1/FoxO1 pathway and prevented OVX-induced bone loss in the mouse model [327]. In a clinical trial, postmenopausal women who regularly took 75 mg of resveratrol twice daily showed improvements in bone density in the lumbar spine and femoral neck as well as a decrease in the bone resorption marker CTX [328]. Furthermore, both preclinical and clinical trials have confirmed the protective effect of resveratrol on OA [329]. Zhou et al. observed that the application of resveratrol inhibited chondrocyte apoptosis, increased trabecular bone number of the subchondral bone, and elevated bone density by activating SIRT1 [330]. A clinical trial confirmed that persistent resveratrol administration reduced chronic pain in age-related OA of postmenopausal women [331]. In addition, resveratrol has shown promise in ameliorating the progression of IDD by preventing further degeneration of intervertebral disc cells [332]. Wu et al. found that the treatment of resveratrol inhibited NPCs apoptosis and promoted ECM synthesis, by suppressing IL‐6/JAK/STAT3 pathway [333].

Although preclinical studies have demonstrated the therapeutic potential of histone-modifying modulators in alleviating degenerative skeletal diseases, their clinical translation remains hindered by challenges such as suboptimal pharmacokinetics, systemic toxicity, adverse effects, and drug resistance. Future research should focus on improving drug delivery strategies, advancing the design of isoform-specific inhibitors, developing dual- or multi-target agents, and initiating early-phase clinical trials. Moreover, a deeper understanding of their molecular mechanisms and epigenetic context is essential for bridging the gap between experimental findings and clinical application.

5. Conclusion and prospects

In summary, histone modification contributes significantly to the pathogenesis of degenerative skeletal diseases. Abnormalities in histone modification induced by various pathogenic factors affect the normal physiological functions of cells by influencing crucial genes expression, promoting tissue degeneration, and leading to the onset and progression of diseases. In this paper, we reviewed the role of histone modification on major degenerative bone diseases (OP, OA, IDD), discussed the research progress of their related regulatory mechanisms, and provided some elaboration and prospects for the therapeutic role and application of histone modifying enzyme modulators in degenerative skeletal diseases.

Compared to existing reviews, such as Sun P et al. (2022) on OP and Ma Z et al. (2023) on EZH2 in musculoskeletal diseases, this review takes a more comprehensive approach. While Sun et al. focus on OP, our review also includes other degenerative skeletal diseases. Additionally, while Ma et al. focus on EZH2, we cover a variety of histone-modifying enzymes and their diverse roles in disease pathogenesis, offering new insights and therapeutic avenues. Another strength of our review is the in-depth discussion of histone-modifying enzyme inhibitors, highlighting their therapeutic potential and clinical translation challenges, such as pharmacokinetic issues, toxicity, and side effects. We also emphasize the gaps in clinical trial data for degenerative skeletal diseases, underscoring the need for further research to move from preclinical findings to clinical applications.

However, current research on histone modifications in degenerative skeletal diseases still faces several limitations. First, there is insufficient exploration of the cell-specific effects of histone modifications. While many studies focus on histone modifications in BMSCs, osteoblasts, and chondrocytes, research on other cell types, such as synovial cells, subchondral bone cells, cartilage endplate cells, and annulus fibrosus cells, remains relatively underexplored. This results in an incomplete understanding of histone modifications in different cell types, thereby hindering a comprehensive elucidation of their multifaceted regulatory roles in degenerative skeletal diseases. Second, there is an imbalance in the types of histone modifications studied, with a predominant emphasis on methylation and acetylation, while other modifications, such as phosphorylation, ubiquitination, and lactylation, have been less extensively investigated in the context of degenerative skeletal diseases. Third, research on the interactions between histone modifications and other disease pathways is limited. Most studies are confined to isolated signaling pathways or specific epigenetic modifications, neglecting the intricate interplay within broader disease networks. Fourth, there is a gap in the translation of basic research into clinical applications. Current studies on histone modifications in skeletal diseases are predominantly confined to cell and animal models, with limited clinical validation. Fifth, the development of pertinent therapeutic agents and their clinical applications remain fraught with challenges. Although numerous therapeutic agents have been developed, issues related to pharmacokinetics, toxicity, adverse effects, and drug resistance continue to curtail their clinical efficacy.

Consequently, future research should focus on several key areas. One important direction is comprehensive mechanistic exploration, which involves further investigation into the role of histone modifications across a broader spectrum of cell types and modification types. Emphasis should be placed on elucidating the intricate interactions within complex mechanisms to gain a more holistic understanding of their pathogenic role in degenerative skeletal diseases. Another critical area is translational research, including the initiation of further clinical trials to substantiate the clinical relevance of histone modifications and their potential therapeutic applications. Finally, innovative drug development should be prioritized, advancing the development of cutting-edge drug delivery systems, enhancing the selectivity of histone-modifying agents, and designing multi-target therapeutic strategies to address the complexity and heterogeneity of degenerative skeletal diseases.

In addition, the integration of emerging technologies such as single-cell sequencing and gene editing offers powerful tools to dissect histone modification patterns at single-cell resolution and functionally validate their roles in specific cellular subpopulations. These approaches are expected to provide deeper insights into the epigenetic landscape within the pathological microenvironment and facilitate the identification of novel, more precise therapeutic targets. Currently, single-cell RNA-seq studies have revealed distinct subpopulations of chondrocytes with unique transcriptional profiles in osteoarthritic cartilage, highlighting cell-specific pathogenic mechanisms [334,335]. Moreover, Bittner et al. constructed a comprehensive 3D chromatin conformation map of primary osteoarthritic chondrocytes. This work identified novel candidate effector genes regulated by chromatin interactions, offering valuable insights into epigenetic regulation in OA [336].

In conclusion, histone modification plays an important role in the regulation of degenerative skeletal diseases, and further exploration of the related pathogenesis and the development of related targeted drugs are of great significance for the treatment of diseases.

Data availability

Data sharing not applicable to this article as no datasets were generated or analysed during the current study.

Author contributions

Yao Zhang, Jiale Wang and Di Hua drafted the manuscript, designed, and developed the figures and tables; Dechun Geng, Xiexing Wu, and Haiqing Mao conceptualized and structured the article, polished the language of this manuscript, and supervised and revised the work critically for important intellectual content; Chunyang Fan selected the title, conducted a literature review, and proofread the first draft; Wei He, Yongkang Deng, Maoting Tang collected related papers and supervised this work. All authors have read and approved the final manuscript and agree to its publication in the journal.

Funding

This work was supported by the National Natural Science Foundation of China (81972104, 82272547, 82572793), the Postdoctoral Science Foundation Project of China (2024M752336), the Natural Science Foundation of Jiangsu Province (BK20220252, BK20241795), Suzhou Key Clinical Disease Diagnosis and Treatment Technology Special Project (LCZX202203), Clinical Research Project of the First Affiliated Hospital of Soochow University (BXLC017).

Declaration of competing interest

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.