A review of the relationship between gut microbiota and osteoporosis in high altitude environments

Abstract

Osteoporosis is the most common bone metabolic disease, but the altitude environment increases the incidence of osteoporosis. Gut microbiota is a key potential target for osteoporosis. However, it is not clear how plateau environment (hypoxia/hypothermia) interferes with the development of osteoporosis by affecting gut microbiota. Therefore, the aim of this paper is to explain that hypoxia and hypothermia environment is involved in bone metabolism regulation by affecting gut microbiota, which may be one of the pathways for the early development of osteoporosis. This paper reviews a large number of clinical and basic studies to systematically evaluate the pathway by which gut microbiota is involved in regulating bone metabolism, and further discuss the potential effects of hypoxia/hypothermia on gut microbiota in regulating bone metabolism. This review summarizes that gut microbiota was mainly involved in the regulation of bone metabolism through immune, hormone and metabolite levels, while hypothermia/hypoxia affected bone metabolism mainly through the effects of microbial immune response and short-chain fatty acids (SCFAs) secretion. In addition, our interpretation of Tibetan dietary patterns reveals a new potential complementary therapy for osteoporosis intervention. Although the initial results are exciting, more trials are needed to understand the interactions between diet, gut microbiota, and bone metabolism.

Introduction

Osteoporosis is a systemic bone disease characterized by low bone mass, damage to the microstructure of bone tissue, increased bone fragility, and easy fracture [1]. At present, osteoporosis has been defined as a worldwide public health problem. According to the statistics of the World Health Organization (WHO), more than 200 million people in the world suffer from osteoporosis, and with the intensification of global population aging, this number is still rising [2, 3]. Age and gender are the most important risk factors for osteoporosis. With the increase of age, the bone reconstruction process of the human body is unbalanced, and bone absorption is greater than bone formation, resulting in gradual loss of bone mass. In women after menopause, due to the sharp decline in estrogen levels, the inhibitory effect of estrogen on osteoclasts is weakened, bone absorption is significantly accelerated, and osteoporosis is more likely to occur. Among people over 50 years of age, the prevalence of osteoporosis is about one third in women and one fifth in men [2]. However, due to the rise of"internal volume culture"[4], long-term sedentary working mode, staying up late and bad eating habits [5, 6], osteoporosis is no longer only the"patent"of the elderly, and more and more young people are also affected by osteoporosis in recent years [7–9]. In addition, at present, the medical and health care costs of osteoporosis treatment are expensive, which brings huge economic pressure to individuals and governments [10], so how to prevent osteoporosis in an early stage has become the focus of attention. However, we found that the unique environment of the plateau aggravated the incidence of osteoporosis [11–14], but the incidence of osteoporosis in Tibetan people in the plateau was lower than that in Han people. Interestingly, this phenomenon may not only be affected by genes, but also be related to the Tibetan diet [15–17]. The absorption of dietary nutrients is mainly through the intestine. Therefore, we will discuss the causes of osteoporosis in the plateau environment from the level of gut microbiota changes, and look for possible early intervention methods for osteoporosis from the Tibetan diet, so as to provide some new ideas for preventing osteoporosis.

Key role of gut microbiota in osteoporosis.

Bone is a dynamic organ, which requires osteoblasts and osteoclasts to maintain a relative dynamic balance in order to maintain its normal function; otherwise, it will lead to bone diseases, such as osteoporosis [18]. Bone homeostasis is mainly regulated by estrogen, parathyroid hormone and immune cells [19]. Recent studies have found that osteoporosis is closely related to gut microbiota, and gut microbes may become a key regulator of bone physiology [20]. Therefore, in the following discussion, we mainly discuss the role of gut microbiota in bone metabolism regulation and its influence on ameliorating osteoporosis from the perspective of immune system, hormone regulation, gut metabolism and gut-bone axis (Fig. 1).

Fig. 1.

Gut microbiota regulates bone metabolism through three pathways. Note: In immune regulation, Th17 cells are induced and differentiated by specific components of the intestinal microbiota, such as SFB, E. lenta, and Bifidobacterium adolescens, thereby promoting osteoclastogenesis via receptor activator of nuclear factor-κB ligand (RANKL) signaling. Meanwhile, Treg cells, under the influence of intestinal Clostridium species, secrete anti-inflammatory cytokines including IL-4, IL-10, and TGF-β to suppress osteoclast activity and promote osteogenesis. In terms of hormonal regulation, estrogen metabolism is modulated by GUS enzymes associated with Lactobacillus and other gut microbiota. Parathyroid hormone (PTH) facilitates bone loss mediated by Th17 cells through the action of intestinal SFB, while requiring intestinal butyrate to enhance osteogenesis via the Treg/CD8 + T cell-WNT10b pathway. Dysbiosis of the intestinal microbiota may also impact bone metabolism through 5-HT synthesis pathways. Among the intestinal metabolites, acetic acid inhibits osteocalcin secretion, whereas propionic acid and butyric acid exert protective effects against ovariectomy-induced bone loss. Collectively, the intestinal microbiota and its metabolites regulate the equilibrium between osteogenesis and osteoclastogenesis via immune, hormonal, and other physiological pathways

Gut microbiota regulates bone metabolism through the immune system

Most studies have proved that the gut microbiota is closely related to the host immune system [21]. However, few clinical studies have been conducted to explain the changes of bone metabolism through immune regulation of gut microbiota. Sjogren et al. first discovered in 2012 that gut microbiota has the potential to regulate bone mass through the immune system [22]. Compared with traditional mice, germ-free mice showed decreased expression of inflammatory cytokines (tumor necrosis factor-α (TNF-α), receptor activator of nuclear factor-κB ligand (RANKL), Interleukin (IL) −6) in bone and bone marrow, and decreased number of CD4 T cells in bone. This immune dysregulation leads to reduced osteoclast production, which leads to reduced bone resorption. After re-colonizing the gut microbiota, the bone mass of the mice normalized [22]. Recent data suggest that osteoporosis and gut inflammatory diseases may share a common immune component, with increased levels of many pro-inflammatory osteoclast activators in patients with inflammatory bowel disease, including TNF-α, IL-1α and 1β, IL-6, IL-11 (Interleukin-11), and IL-17 [23]. At present, a new pathway of bone metabolism research is called the receptor activator of nuclear factor-κB (RANK)-RANKL-osteoprotegerin (OPG) pathway. RANKL is expressed on the surface of osteoblasts, synovial stromal cells, and activated T cells. RANKL can bind to osteoclast precursors expressing the RANKL receptor, RANK, or the soluble decoy receptor OPG produced by osteoblasts. If RANKL and RANK interact, osteoclasts differentiate and mature, resulting in increased bone loss [24]. Recent studies suggest that changes in the RANKL to OPG ratio may be responsible for bone loss in patients with inflammatory bowel disease. Plasma OPG and RANKL levels are associated with bone mineral density and current treatments for inflammatory bowel disease [25, 26]. Moschen et al. [26] found that plasma levels of OPG were 2.4 times higher in Crohn's disease and 1.9 times higher in ulcerative colitis. These results suggest that elevated OPG levels may attempt to counteract RANKL or TNF-α to drive osteoclast production and maintain normal bone mass. In addition, Peek et al. [27] found significant bone loss in chemical, T-cell-driven, and inflammatory infection models of gut inflammation. Bone loss is associated with an increase in osteoclastogenic cytokines and an expansion of the specific Cd11b⁻/Ly6C^hi osteoclast precursor (OCP) population. Gut inflammation leads to altered OCP expression of surface receptors involved in osteoclast differentiation and function, including the osteoclastogenic coreceptor myeloid DAP12-associating lectin-1 (MDL-1).

As described above, RANKL is an important signaling molecule involved in the connection between immunity and bone metabolism. In previous studies, activated T cells were found to be an important source of RANKL [28]. However, not all T cells have the ability to stimulate osteoclast differentiation after activation. At present, only Th17 cells (T helper 17 cells) selectively express TNF-α of RANKL [28]. According to some studies, segmented filamentous bacteria (SFB) in the intestine [29, 30], E. lenta strains [31], Bifidobacterium adolescentis [32], can increase Th17 cell differentiation. The balance between Th-17 cells and Treg (Regulatory T) cells is crucial for the regulation of inflammatory response [33]. In contrast to the influence of Th17 cells, Treg cells are CD4 T cells with immunosuppressive function and have a beneficial effect on bone remodeling [34]. Treg cells can secrete some anti-inflammatory cytokines IL-4 (Interleukin-4), IL-10 (Interleukin-10) and TGF-β (transforming growth factor-β) to promote the increase of osteoblasts and inhibit the secretion of osteoclasts [35–37]. IL-10 secreted by Treg cells helps to down-regulate the expression of RANKL and enhance the secretion of OPG, thus inhibiting osteoclast differentiation [38]. Existing studies have found that gut microbiota can regulate Treg cell differentiation. Stefka et al. [39] found that when Clostridia was implanted in the intestine of germ-free mice, ^Foxp3+Tregs were significantly increased in the colon gut lamina propria. Another study also demonstrated that 17 strains belonging to the Clostridia IV, XIVa, and XVIII clusters provide bacterial antigens and a TGF-β-rich environment as a community, promoting the expansion and differentiation of Treg cells [40]. In summary, these studies reflect the close relationship between gut microbiota and the immune system, and regulate bone metabolism and maintain bone balance through immune response (Fig. 1).

Gut microbiota regulates bone metabolism through hormones

Bone homeostasis is regulated by estrogen, parathyroid hormone, and serotonin (5-HT); however, the gut microbiota has the ability to regulate concentrations of these hormones. Due to many studies on estrogen in bone metabolism, we first describe the regulation of estrogen by gut microbiota.

1. Estrogen.

It is well known that estrogen deficiency is the main cause of postmenopausal osteoporosis in women [41]. Estrogen regulates the balance between osteoblasts and osteoclasts to maintain bone mass [42]. Studies have shown that estrogen enhances OPG production in osteoblasts by inhibiting RANKL expression in CD 3 T cells and CD 20 B cells [43]. Estrogen deficiency leads to T cell-mediated increase of TNF-α [44] and activation of the Nuclear factor kappa-B (NF-κB) pathway [45], which enhances osteoclast differentiation and leads to bone loss. Now more and more studies have proved that gut microbiota β-glucuronidase (GUS) is the core of regulating the body's estrogen metabolism [46]. Masamune [47] first identified GUS genes in E. coli and other enterobacteriaceae in 1934, and preliminarily identified GUS's involvement in estrogen metabolism in 1944 [48]. Approximately 279 GUS genes have been obtained in the gastrointestinal (GI) database of the Human Microbiome Project. Among them, 93.5% are classified into Bacteroidetes (52%), Firmicutes (43%), Verrucomicrobia (1.5%) and Proteobacteria (0.5%) [49]. And GUS and its related enzymes have been found in Bacteroidaceae, Bifidobacteriaceae, Clostridiaceae, Enterobacteriaceae, Lactobacillaceae, Ruminococcaceae and Verrucomicrobiaceae [50]. Ma et al. [51] reported that after ovariectomy in rats, changes occurred in the gut microbiota. They speculated that Ruminococcus flavefaciens and Prevotella might be the possible pathological causes of osteoporosis induced by steroid hormone deficiency. Another study also demonstrated that there was a correlation between GUS activity and the relative abundances of Lactobacillaceae, Ruminococcaceae and Streptococcaceae in the gut contents [50]. In addition, Li [52] also found that gut microbiota dysbiosis was related to bone loss associated with steroid hormone deficiency. These results suggest that GUS may be an important mediator in the regulation of bone metabolism through the interaction between gut microbiota and estrogen. GUS may possess the potential to be a biomarker for bone metabolism. However, there is currently a lacking research on the regulatory role of GUS in bone metabolism (Fig. 1).

• (2)Parathyroid hormone (PTH).

The PTH produced by the parathyroid gland is essential for bone development and bone homeostasis. However, persistent hyperparathyroidism promotes bone resorption, resulting in decreased net bone mass. This is also an important factor in osteoporosis and fracture [53]. Intermittent use of PTH analogues is the only bone formation promoting therapy approved by the U.S. Food and Drug Administration (FDA) for the treatment of osteoporosis [54]. However, there are only a few studies on the relationship between PTH and gut microbiota. Yu et al. [55] found that PTH only resulted in bone loss in mice treated with antibiotics or germ-free mice whose microbiota was enriched by the SFB induced by Th17 cells. It is speculated that SFB microbiota enables PTH to expand gut TNF-T and Th17 cells, increase sphingosine-1-phosphate receptor 1 (S1PR1), and mediate outflow and recruitment from the gut to bone marrow (BM), thereby leading to bone loss. In addition, C-X-C motif chemokine receptor 3 (CXCR3) -mediated TNF-T cell homing to BM upregates the Th17 chemoattractant C–C motif chemokine ligand 20 (CCL20) and recruits Th17 cells to BM. One study has also shown that PTH is required by gut microbiota to stimulate bone formation and increase bone mass. When the gut microbiota is depleted, the levels of PTH and the gutmetabolite butyrate are reduced, and bone metabolism is disturbed. However, after the restoration of gut microbiota, butyrate and PTH levels recovered and bone metabolism maintained balance. Therefore, the researchers speculated that butyrate may be responsible for connecting the communication between the bones of the intestine, and PTH needs butyrate to increase the number of Treg cells in the BM. Treg cells stimulate the production of osteogenic wingless-related integration site (Wnt) ligand Wnt10b through BM CD⁸⁺ T cells, thereby activating WNT-dependent bone formation [56] (Fig. 1). These studies reveal the mechanisms of gut microbiota-mediated gut-bone communication in mouse models, which may facilitate early intervention and treatment. Targeting the gut microbiota and its metabolites may be a therapeutic approach to treating diseases associated with abnormal osteogenic metabolic disorders of PTH.

• (3)5-HT

Existing studies have found that 5-HT plays an important regulatory role in bone development and maintenance [57, 58], and about 95% of 5-HT exists in enterochromaffin cells [59]. Through literature investigation, we found that osteoblasts and osteoclasts contain 5-HT receptors, and the increase of 5-HT in serum may lead to the decrease of bone mass in mice [22, 60–66]. Lavoie et al. also reported that Dextran Sulfate Sodium Salt (DSS) induced colitis in mice, which promoted the increase of 5-HT in the blood, resulting in bone loss in mice. However, the subsequent administration of 5-HT synthesis inhibitor and 5-HT1B receptor antagonist in DSS mice could improve the bone loss in colitis mice [62]. In addition, it has been found that Tph1 (rate-limiting enzyme tryptophan hydroxylase 1) is the initial enzyme in gut5-HT biosynthesis, and inhibition of enteric-derived 5-HT synthesis using LP533401, a small molecule inhibitor of Tph1, can prevent bone loss induced by OVX (oophorectomy) [63]. In addition, Sjogren et al. [22]. also demonstrated that the serum 5-HT content of germ-free mice was lower than that of normal mice, and the bone mass was higher. These animal studies suggest that gut microbiota plays an important role in regulating 5-HT in maintaining bone metabolism. This was thus confirmed in clinical trials, one of which, osteoporotic Fractures in Men Study (MrOS) Sweden, analyzed serum 5-HT levels and bone mineral density in 950 men (aged 69 to 81 years), and found that 5-HT was linearly correlated with hip fracture, with patients with high 5-HT levels having three times the risk of fracture [64]. In addition, Modder et al. [65] also found that high 5-HT levels were negatively correlated with Bone Mineral Density (BMD) of the female spine, femur, and trabecular bone. These results suggest that 5-HT levels can predict the probability of osteoporosis. However, the relationship between 5-HT levels and BMD remains controversial, with most studies suggesting that high levels of 5-HT are associated with an increased risk of osteoporosis. However, a study on the relationship between serum 5-HT and BMD in postmenopausal women in China found that lower serum 5-HT levels were associated with lower BMD in lumbar, femur, and neck bones in postmenopausal women [66]. More research is needed to explain the association between 5-HT and BMD.

In summary, dysregulation of the gut microbiota leads to bone loss by regulating 5-HT synthesis. The synthesis of 5-HT regulated by gut microbiota was negatively correlated with BMD (Fig. 1). BMD is known to be an important factor in the development of osteoporosis, so the regulation of 5-HT synthesis by gut microbiota may be a means to intervene in the development of osteoporosis [67].

Gut microbiota regulates bone metabolism through metabolites

The gut microbiota is also able to regulate distal organs by producing secondary metabolites, which are often referred to as postbiotics [68]. Epigenome mainly includes SCFAs, enzymes, peptides, vitamins and phosphatides. Here we mainly discuss the regulatory effects of SCFAs on bone metabolism. SCFA (including acetate, propionate, and butyrate) is produced by the gut microbiota by ferment of indigestible dietary fiber, acetate can be produced by many types of bacteria, but propionate and butyrate are only produced by specific bacteria [69–71]. For example, Akkermansia muciniphila produces propionic acid from the digestion of the intestinal mucus layer [70]. However, butyrate is generated by Faecalibacterium prausnitzii, Eubacterium rectale, Eubacterium hallii, Ruminococcus bromii and other small gut microbiota [72]. Interestingly, acetate, propionate, and butyrate regulate bone mass gain through different pathways. Lucas et al. [73] found that acetate inhibited osteocalcin secretion in a T-cell and B-cell dependent manner, while propionate and butyrate effectively prevented OVX-induced bone loss by reducing osteocalcin and serum CTX-I, (C-terminal telopeptide of type I collagen) levels (Fig. 1). At present, the relationship between SCFAs and bone metabolism is mainly focused on butyrate. It has been found that butyrate can inhibit histone deacetylation in cells [74], and inhibition of histone deacetylation can reduce the osteoclast-specific mRNA expression of cathepsin K and calcitonin receptors [75], and inhibit RANKL-induced osteoclast formation by inhibiting the induction of the transcription factor c-Fos produced by osteoclasts [76]. These results suggest that butyrate as a histone deacetylase inhibitor may be a novel treatment for typical bone diseases.

In addition, the researchers also found that supplementation with Lactobacillus rhamnosus GG in mice can increase the level of butyrate, stimulate the secretion of Wnt ligand Wnt10b by CD⁸⁺ T cells by increasing the number of Treg cells in the intestine and BM, and activate the Wnt signaling pathway in osteoblasts to stimulate bone formation improve osteoporosis [77]. Ge et al. [78] also found that long-term Pb leakage in mice destroys bone metabolism, resulting in osteoporosis, and causes imbalance of butyrate producing gut microbiota (such as Butyrivibrio crossotus and Clostridium sp.JN9), resulting in butyrate reduction. The supplementation of butyrate restored the gut microbiota and structural damage of Pb burst mice, increased the abundance of Treg cells, and alleviated osteopenia.

In summary, SCFA has the potential role of regulating bone metabolism and ameliorating osteoporosis. One study reviewed the effects of dietary habits on bone health over the past decade and found that the Mediterranean diet has a protective effect on bones [79], increasing SCFA levels by supplementing dietary fiber or being rich in SCFA-producing bacteria [73, 80]. This could be a way to intervene early in osteoporosis.

Influence of gut microbiota changes on bone metabolism in plateau environment

In the above, we have described the important regulatory role of gut microbiota on bone metabolism. However, in addition to the pathological status of the body itself, environmental factors are also a key factor affecting the abundance of microbiota. In high altitude environment, the diversity of human gut microbiota may decrease. Due to the combination of hypoxia, low pressure, hypothermia and other special environmental conditions on the plateau, as well as changes in diet structure and other factors, some types of bacteria that are more sensitive to environmental changes will decrease. Li et al. [81] found in a comparative analysis of gut microbiota of people at low or high elevations in 2015 that Firmicutes increased in the guttract of people at high elevations while Bacteroidetes decreased compared with those at low elevations. This microbiota change was mainly due to a significant reduction in Prevotella. Another study also found that 51 bacterial microbiota species increased and 57 bacterial microbiota species decreased in the human guttract at high altitude, and most of these bacterial microbiota species belonged to Prevotella and Bacteroides, which had nothing to do with the human population [82].

Therefore, these results indicate that there is a direct relationship between gut microbiota and plateau environment, and the dominant microbiota is mainly Firmicutes, Prevotella and Bacteroides. Studies have shown that Firmicutes, Prevotella and Bacteroides are closely related to bone metabolism [83–85]. Due to its unique geographical conditions, including hypoxia, hypothermia and high radiation, the incidence of osteoporosis among residents in the plateau region is significantly different from that in the plain region, which has attracted the attention of many scholars. Most studies believe that people in plateau environment are more prone to osteoporosis [11–14]. Therefore, high-altitude environments influence bone health through both direct (hypoxia/hypothermia) and indirect (gut microbiota-mediated) mechanisms. Below, we synthesize these pathways, highlighting the crosstalk between environmental stressors and the gut-bone axis.

Hypoxia-microbiota-bone metabolism

Hypoxia is a primary characteristic of high-altitude environments and exerts direct effects on bone metabolism by activating the hypoxia inducible factor (HIF) signaling pathway and inducing inflammatory responses (Fig. 2). For instance, hypoxia inhibits osteoblast mineralization and promotes osteoclast differentiation via HIF-1α and HIF-2α, while also upregulating pro-inflammatory cytokines (TNF-α, IL-6) through the NF-κB pathway [86–95]. These direct effects disrupt bone remodeling balance. Concurrently, hypoxia influences bone metabolism indirectly via gut microbiota dysregulation. Beyond direct effects, hypoxia-induced gut microbiota dysregulation exacerbates bone loss. Studies show that prolonged hypoxia reduces the abundance of SCFA-producing bacteria (e.g., Blautia, Bifidobacterium) and increases pro-inflammatory Bacteroides [84, 96–101]. This dysregulation impairs gut mucosal barrier integrity, allowing Lipopolysaccharide (LPS) translocation and NF-κB-mediated inflammation [102–105]. Additionally, reduced SCFA synthesis (e.g., butyrate) disrupts osteoblast-osteoclast balance, as SCFAs inhibit osteoclastogenesis via histone deacetylase inhibition [73–76]. However, due to the lacking detailed studies, there is no specific explanation for the relationship between hypoxia and bone metabolism in gut microbiota.

Fig. 2.

Hypoxic environment directly and indirectly regulates bone metabolism via A HIF signaling and B gut microbiota. Note: (A) Under hypoxic conditions, HIF-1α mediates the regulation of the RANKL/OPG pathway via osteoblasts, while HIF-2α directly inducesTNF receptor associated factor 6 (TRAF6) expression, thereby promoting osteoclastogenesis. Additionally, the inactivation of PHD enzymes activates the NF-κB signaling pathway, leading to enhanced expression of inflammatory genes. Notably, NF-κB and HIF-1α exhibit mutual upregulation through a positive feedback loop. (B) Hypoxic conditions can result in a decreased abundance of intestinal microbiota associated with the synthesis of SCFAs, such as Ruminococcaceae and Trichospiraceae. The reduction in acetic acid may enhance osteocalcin secretion via an immune-mediated mechanism. Propionic acid and butyric acid increase osteocalcin levels and serum CTX-I, thereby contributing to bone loss. Additionally, hypoxia-induced dysbiosis of the gut microbiota compromises the integrity of the intestinal mucosal barrier, allowing bacterial products to activate the NF-κB pathway through Toll-like receptor 4 (TLR4). This leads to the release of pro-inflammatory cytokines, including IL-1β, IL-6, and TNF-α, which inhibit osteoblast activity and ultimately result in bone loss

Hypothermia-microbiota-bone metabolism

Hypothermia is another key factor that affects bone metabolism through neuroendocrine regulation (hypothalamic-pituitary-thyroid axis (HPT axis)/hypothalamo–pituitary–adrenal axis (HPA axis)), reduced bone tissue perfusion, and inhibition of osteoblast activity [20, 106–114]. For instance, activated thyroid hormones can increase bone renewal, while prolonged exposure to cold can reduce the alkaline phosphatase activity of osteoblasts and damage bone matrix synthesis [115–117] (Fig. 3A). These direct effects are further regulated by the gut microbiota.Hypothermia-induced gut microbiota shifts (e.g., reduced Coprococcus, Lachnospiraceae; increased Clostridium) disrupt SCFA production, particularly butyrate [118–125]. Butyrate, critical for bone formation via Treg cell-mediated Wnt10b signaling [56, 77], is reduced under hypothermia, leading to decreased osteoblast activity. Supplementation with butyrate restores microbiota balance and bone mass [122], highlighting the pivotal role of gut metabolites in cold-induced bone loss (Fig. 3B). We mentioned in Sect. 2.4 that SCFA, a major metabolite in the gut, is also involved in regulating bone metabolism, and hypothermia may also affect bone loss by affecting SCFA synthesis. However, as far as we know, there are no studies on the participation of gut microbiota in bone metabolism under hypothermia environment. So this part of the story lacks a clear enough explanation.

Fig. 3.

Hypothermia regulates bone metabolism through A neuroendocrine pathways and B gut microbiota-derived SCFAs. Note: (A) hypothermia can promote the secretion of thyroid hormones by activating the HPT axis, thereby enhancing osteoblast activity. Simultaneously, it can activate the HPA axis, leading to high concentrations of adrenal cortical hormones that inhibit osteoblast function. Additionally, hypothermia reduces blood perfusion and nutrient absorption in bone tissue through peripheral vasoconstriction, which in turn decreases the metabolic activity of bone cells and the alkaline phosphatase activity of osteoblasts, ultimately reducing bone matrix synthesis. Prolonged exposure to hypothermias may result in bone loss. (B) Chronic cold exposure has been shown to reduce the abundance of butyrate-producing microbiota (Fecococcus, Trichospiraceae, Ruminococcaceae) in the intestinal tract while increasing the prevalence of Clostridium, Riomycetidae, and Bacteroides. It is hypothesized that this alteration disrupts bone formation by decreasing butyric acid synthesis, which reduces the number of Treg cells in the bone marrow and inhibits the production of osteogenic Wnt10b ligands. Additionally, the Mucoricaceae family exhibits a positive correlation with bone mineral density, whereas the Bacteroides genus demonstrates a negative correlation with bone mineral density and is associated with an increased risk of osteoporosis. Nevertheless, the precise mechanisms by which low temperatures influence bone metabolism via the gut microbiota remain underexplored and warrant further detailed investigation

Diet in high altitude environment interferes with osteoporosis

At present, there are two main treatments for osteoporosis: bone resorption inhibitors and bone formation promoters. Bone resorption inhibitors are mainly bisphosphonates (alendronate, risedronate), estrogen and hormone replacement therapy. This treatment mainly promotes osteoclast apoptosis, reduces bone resorption, and thus enhances bone strength [126]. According to Association for the Advancement of Computing in Education (AACE/ACE) guidelines, first-line treatment for most Postmenopausal Osteoporosis (PMO) patients at high risk of fracture includes alendronate, risedronate, zoledronic acid, and dinomumab [127]. Bone formation promoters mainly increase the activity of osteoblasts, thereby promoting bone formation. The most commonly used bone formation enhancer is parathyroid hormone [126]. Other treatments include some active vitamin D and its analogues, vitamin K2, strontium salts, Chinese medicines or compounds [18, 128]. However, the existing treatment has many side effects and high drug cost, such as the gastrogutadverse reactions of bisphosphonates and the high price of parathyroid hormone analogues [129]. Therefore, researchers have been looking for better therapies to achieve early intervention or adjuvant treatment of osteoporosis. Because gut microbiota plays an important role in bone metabolism, gut microbiota may be a better target for the prevention and treatment of osteoporosis. Over the past decade, dietary patterns have been found to be involved in bone health [79].

Especially in plateau areas, we found that the bone density of plateau people was higher than that of plain people [130], which may be related to plateau diet. In the plateau areas of different regions of the world, the unique geographical environment has nurtured the unique food culture. In the Tibetan Plateau in Asia, the Alps in Europe, the Andes Plateau in the Americas and the Ethiopian Plateau in Africa, the active ingredients in the diet have a profound impact on gut microbiota, and there is a complex correlation with bone mineral density (Table 1). The study of these relationships is helpful to understand the healthy adaptation mechanism of plateau population, and can also provide reference for the healthy diet of people in other areas. Therefore, here are some simple descriptions of how dietary characteristics in high altitude areas interfere with bone density through gut microbiota.

1. European Plateau:

Table 1.

Association of high altitude diet with gut microbiota and bone metabolism

	Name	Altitude	Special diet and active ingredients	Gut Microbiota	Relationship with bone mineral density
European plateau	Central Siberian plateau	Between 500–1000 m above sea level	Protein in reindeer meat; Trace elements (zinc, copper and magnesium) and polyunsaturated fatty acids in fish products; omega-3 fatty acids in the white fish family [160]	Promote the growth and metabolism of beneficial bacteria such as Lactobacillus and Bifidobacterium [134]; Regulation of gut metabolites such as SCFAs [161, 162]	1. Lactic acid produced by Lactobacillus fermentation can combine with calcium to form calcium lactate, improve the solubility of calcium and promote the absorption of calcium. After calcium is absorbed into the blood, it can be used to mineralize bones and increase bone density [135, 136]
	Alpine mountains	The average elevation is about 3,000 m	Dairy products: probiotics in cheese, yogurt; Polyphenols in olive oil; Protein in air-dried beef; Dietary fiber in rye bread [133]		2. Olive oil polyphenols and whole grain dietary fiber can play a role in regulating the metabolites of gut microbiota, reducing the production of gut inflammation-related metabolites, and also helping to reduce the burden of bone inflammation and protect bone density [137]
American plateau	Andean plateau	Around 3000—4000 m	Quinoa: Rich in protein, dietary fiber and a variety of minerals; Meat products: llama and alpaca meat; Dairy products: alpaca milk and milk and cheese; Vegetables: potatoes, sweet potatoes and other root vegetables; Fruits: Blueberries, cactus fruits are rich in anthocyanins and vitamin C [139, 140]	Quinoa polysaccharides can be considered prebiotics because of their ability to increase Bifidobacterium and Collinsella, and to regulate short-chain fatty acid acid [141, 142]	1. Dietary fiber in quinoa can promote SCFAs produced by beneficial bacteria, improve the absorption efficiency of calcium in the intestine, and participate in bone mineralization after calcium is absorbed [141–143]. 2. Blueberry anthocyanins indirectly promote the absorption of nutrients by regulating gut microbiota and also contribute to the maintenance of bone mineral density [144, 145]
African plateau	Ethiopian plateau	The average elevation is between 2,500–3,000 m	Teff: Rich in plant fiber; Baobab fruit: rich in nutrients such as vitamin C, fiber and polyphenols [146–148]	Plant fibers in teff and baobab fruits can be decomposed and utilized by various gut microbiota in the gut, promoting the growth of beneficial bacteria such as BIfidobacteria and Prevotella [149, 150]	The plant fiber in teff and baobab fruit promotes SCFAs produced by beneficial bacteria, which can improve the gut absorption efficiency of calcium and other minerals and improve bone mineral density [141–143, 149]
Asian plateau	Tibetan plateau	The average elevation is over 4,000 m	Qingke: Rich in beta-glucan. Yak beef and yak milk: conjugated linoleic acid, immunoglobulin and other components; Wolfberry: Wolfberry polysaccharide; Sea buckthorn: vitamin C, flavonoids, FLA: rich in omega-3 fatty acids [151–154]	1. Qingke β-glucan has prebiotic potential, which can promote the production of short chain fatty acids such as acetic acid, propionic acid and butyric acid [155]. 2.FLA can increase Bifidobacterium and decrease Firmicutes [135]. 3. Polysaccharides in Wolfberry and sea buckthorn can increase Akkermansia and Lactobacillus [159, 163]	1. SCFAs produced during the growth and reproduction of active ingredients such as Qingke β-glucan can promote the absorption of minerals such as calcium and phosphorus in the intestine [78, 141, 155]. 2. CLA in yak milk also contributes to gut absorption of nutrients, provides sufficient raw materials for bones, and promotes the increase of bone density [156–158]. 3.FLA can reduce Firmicutes and improve bone density by increasing Bifidobacterium [85, 102, 135]. 4. Lycium barbarum polysaccharides may reduce inflammatory levels and improve bone metabolism by increasing probiotics [140, 141]

In the highland regions of Europe, such as the Alps, dairy products such as cheese and milk are an important part of the diet, rich in protein, calcium, phosphorus and other nutrients. In addition, olive oil also plays an important role in the diet of the Mediterranean coast plateau, which is rich in polyphenols, such as hydroxytyrosol, oleuropein and so on. There are also whole grain foods, which are rich in dietary fiber, vitamins and minerals [131]. Among them, the protein in dairy products is digested and decomposed into amino acids and small peptides in the intestine, which can provide nutrients for gut microbiota and promote the growth and metabolism of beneficial bacteria such as Lactobacillus and Bifidobacterium [132]. Lactobacillus produced by lactic acid bacteria fermentation can combine with calcium to form calcium lactate, improve the solubility of calcium and promote the absorption of calcium. After calcium is absorbed into the blood, it can be used to mineralize bones and increase bone density [133, 134]. In addition, olive oil polyphenols and whole grain dietary fiber can play a role in regulating the metabolites of gut microbiota, reducing the production of gutinflammatory metabolites, and also helping to reduce the burden of bone inflammation and protect bone mineral density [135, 136].

• (2)American Plateau:

In the American highlands such as the Andean highlands, quinoa is a representative food, rich in protein, dietary fiber, minerals (such as calcium, magnesium, iron, etc.) and many vitamins [137, 138]. In addition, some local characteristic fruits such as blueberries are also more common, and blueberries are rich in antioxidant components such as anthocyanins. Among them, minerals such as calcium, magnesium and protein in quinoa are of great significance for maintaining and improving bone density. Moreover, dietary fiber in quinoa can promote SCFAs produced by beneficial bacteria (Bifidobacterium, Lactobacillus, etc.), improve the absorption efficiency of calcium in the intestine, and participate in bone mineralization after calcium is absorbed [139–141]. In addition, blueberry anthocyanins indirectly promote the absorption of nutrients by regulating gut microbiota, and also contribute to the maintenance of bone mineral density [142, 143].

• (3)African Plateau:

In African highlands such as the Ethiopian Highlands, the traditional staple food is a food called Injera. It is made from teff, a local grain. Teff is rich in plant fiber. Baobab fruit is also a common food, rich in nutrients such as vitamin C, fiber and polyphenols [144–146]. Plant fibers in teff and baobab fruits can be decomposed and utilized by various gut microbiota in the gut, promoting the growth of beneficial bacteria such as Bifidobacteria and Prevotella [147, 148]. The SCFAs produced by these beneficial bacteria in the process of utilizing plant fiber can improve the absorption efficiency of calcium and other minerals in the intestine and improve bone density [139–141, 147].

• (4)Asian Plateau:

Asian plateau areas, such as the Tibetan Plateau, as the world's first plateau has a rich variety of special foods. Qingke is one of the main food crops in the region and is rich in β-glucan. Yak meat and yak milk are also important food sources, yak milk contains conjugated linoleic acid (CLA), immunoglobulin and other components, and yak meat is a high-quality protein provider. In addition, plant foods such as wolfberry, sea buckthorn and flaxseed (FLA) are also relatively common, among which wolfberry contains wolfberry polysaccharide, sea buckthorn is rich in vitamin C and flavonoids, and FLA is rich in omega-3 fatty acids [149–152]. The SCFAs produced by beneficial bacteria in the growth and reproduction process using the active ingredients such as Qingke β-glucan can promote the absorption of calcium, phosphorus and other minerals in the intestine [77, 139, 153]. For example, butyric acid can up-regulate the expression of calcium-binding protein in gutepithelial cells, increase calcium absorption channels, and thus improve calcium absorption rate. CLA in yak milk also contributes to gut absorption of nutrients, provides sufficient raw materials for bones, and promotes the increase of bone density [154–156]. By regulating gut microbiota, Lycium barbarum polysaccharide indirectly affects the immune state of the whole body, and also helps to maintain the immune balance of bones, promote the activity of osteoblasts and inhibit the function of osteoclasts, thus contributing to the maintenance and improvement of bone mineral density [157, 158]. In addition, the omega-3 fatty acids in FLA can also improve gut microbiota and increase bone mineral density [84, 100, 159].

Tibetan diet and osteoporosis

As the Qinghai-Tibet Plateau is the first plateau in the world and has a long cultural history, we focus on the relationship between diet and osteoporosis in the Qinghai-Tibet Plateau population. Some Chinese studies have found that in the Qinghai-Tibet Plateau, the bone of the Tibetan population is significantly stronger than that of the Han population, which may not only be due to ethnic genes, but also may be related to dietary patterns [15–17]. This may indicate that the Tibetan diet has some early preventive and protective effects on osteoporosis in the plateau environment. Therefore, here we mainly discuss the effects of dietary patterns on bone health of people living on the Tibetan Plateau for a long time. In the Qinghai-Tibet Plateau in Asia, most areas are not suitable for food production, and the diet structure is relatively simple. At the same time, in the Hypothermia environment, more calories and nutrients may be needed to maintain the basic metabolism of human body on the plateau [162]. Research on dietary patterns in Tibetan populations is very limited, with only a few studies discussing Tibetan dietary intake.

1. CLA intake

A 2023 study, the first to report a comprehensive analysis of the dietary patterns of Tibetan adults by sex, region, and age group, found that meat intake (100 ± 26 g/day) was higher than the recommended value (40 to 75 g/day), but the meat intake structure was monotonous, mainly yak meat and mutton, and Fish intake was minimal (2 ± 0.1 g/d) [149]. This may be related to religious belief [150]. However, as the main part of daily food intake in plateau areas, yak butter, beef and mutton contain a large proportion of CLA. Some studies have shown that CLA, as one of the gutmetabolites [154], can inhibit gutinflammation in mice [155], and dietary CLA supplementation can enhance the anti-inflammatory effect of mouse bone marrow cells [156]. This may be one of the ways in which dietary meat intake improves bone mass by raising levels of the gutmetabolite CLA early on, enhancing the body's ability to fight hypoxia-induced inflammation.

• (2)Qingke β-glucan intake

Due to the harsh climate of the plateau compared with the plain, the intakes of vegetables (90 ± 19 g/d), fruits (97 ± 25 g/d) and grains (117 ± 27 g/d) were insufficient, which were 70%, 51.5% and 53.2% lower than the recommended values, respectively [149]. However, some studies have also found that the Tibetan population has a high frequency of grain intake and the fact that grain foods provide major nutrients [150]. Qingke is one of the main crops in the Qinghai-Tibet Plateau, which has the highest β-glucan among wheat crops in the world [163], and is also the main grain consumed by Tibetan people. Some studies have proved that Qingke β-glucan has probiotic potential, which can promote the production of SCFAs such as acetic acid, propionic acid and butyric acid [153], and Qingke β-glucan can also reduce the increase of pro-inflammatory gut microbiota abundance induced by capsaicin [164]. It has been mentioned in Part 3.3 that SCFAs regulate bone metabolism [78], so we speculated that the dietary intake of Qingke in the early stage may also have the effect of improving bone mass, which may be related to the enhancement of SCFAs level.

• (3)FLA intake

In the strict sense, FLA is not a typical ingredient in the traditional diet of the plateau. However, in the northwest of China, the planting area of the Qinghai-Tibet Plateau has been expanding [165], and under the influence of modern nutrition concepts, the diet of the Qinghai-Tibet Plateau has been integrated to a certain extent. FLA is also used as a nutritional supplement in the Tibetan diet, such as mixing an appropriate amount of ground flaxseed with highland barley powder, and then adding ghee and tea to stir, and FLA is also added to tea and yak milk. Studies have shown that FLA is one of the best plant sources of omega-3 fatty acids and has the effect of improving the gut microbiota [151]. Yang et al. found that supplementation of FLA to mice on a high-fat diet could prevent the decrease of Bifidobacterium abundance and reduce the abundance of Firmicutes [159]. In Part 4, we have mentioned that the plateau environment can lead to a decline in the abundance of Bifidobacterium and Firmicutes [81, 97], where Bifidobacterium is positively correlated with BMD [84, 100]. In addition, in addition to omega-3 fatty acids, lignans and dietary fiber are also one of the main components of FLA. A study found that lignans in flaxseed can play an anti-osteoporosis role by up-regulating the expression of osteogenic genes [165]. These results suggest that FLA intake is also one of the reasons for enhancing bone mass in Tibetan population.

• (4)Other dietary components

In addition, the Tibetan diet also has a high frequency of intake of some dairy products, such as ghee, yak milk and their products [149]. Tibetan people will make yak milk into yogurt, cheese and other dairy products. Among these dairy products, yak yogurt is unique because the rich Lactobacillus in yogurt helps to improve gut microbiota [166]. Lactobacillus, as one of the main probiotics, is suitable for treating a variety of diseases [167]. Many of these studies have shown that Lactobacillus can improve osteoporosis. McCabe et al. [168]. found that after oral administration of Lactobacillus reuteri ATCC PTA 6475 in male mice, the formation of femur and vertebrae increased, resulting in increased trabecular bone volume in male mice, but there was no significant response to healthy female mice. However, one study showed that treatment with Lactobacillus reuteri prevented bone loss in female ovariectomized mice after menopause [169]. Li et al. [52] further demonstrated that Lactobacillus can treat and prevent estrogen-deficient induced trabecular bone loss in female mice. These results indicate the anti-osteoporosis potential of Lactobacillus, and also indicate the early intervention of Tibetan diet against osteoporosis.

To sum up, we discussed the reasons for the stronger bone mass of Tibetan people than that of Han people from the perspective of Tibetan diet, and provided the relationship between Tibetan diet and osteoporosis for the first time, which will provide some theoretical basis for the early intervention of bone metabolism in people living in plateau areas for a long time.

Conclusions and perspectives

In recent years, more and more studies have been conducted on gut microbiota and osteoporosis. Although some progress has been made, the incidence of osteoporosis in some areas (such as plateau) is different from that in normal environmental populations. Therefore, it is still necessary to discuss the potential interactions and crosstalk between bone and gut microbiota in combination with some environmental factors. Due to this research blind spot, we discussed and analyzed the relationship between bone and gutmicrobiota through a large number of literature studies, combined with hypothermia and hypoxia, two major environmental factors in the plateau, and proposed that the Tibetan diet may achieve early intervention in osteoporosis by regulating gutmicrobiota. Although this diet has some potential to prevent osteoporosis, it needs to be further verified due to the lacking sufficient clinical studies. There are still many problems in this paper that need to be further verified by experiments. The key questions mainly include three points: 1) Whether the Tibetan diet can be accepted by other ethnic groups, and whether long-term use has any adverse effects? 2) How do the active ingredients in Tibetan diet specifically participate in the regulation of bone metabolism through gut microbiota? 3) Is the active ingredient in the Tibetan diet a single component for anti-osteoporosis or a combination of superimposed components to play an anti-osteoporosis role? In conclusion, because the gut is sensitive to the external environment, gut microbes are a key target for studying the occurrence and development of osteoporosis, and the development of targeted active ingredient intervention for the treatment of osteoporosis based on Tibetan diet is a potential alternative therapy. However, because its main mechanism and clinical outcomes are still unknown, more clinical trials are needed to investigate it.

Abbreviations

WHO

World Health Organization

OCR

Oxygen consumption rate

HIF

Hypoxia inducible factor

HIF-1α

Hypoxia Inducible Factor 1 Subunit Alpha

RANKL

Receptor Activator of Nuclear Factor-κ B Ligand

OPG

Osteoprotegerin

HIF-2α

Hypoxia-inducible factor-2 alpha

TRAF6

TNF receptor associated factor 6

IL-1

Interleukin-1

IL-6

Interleukin-6

IL-17

Interleukin-17

NF-κB

Nuclear factor kappa-B

PHD

Prolyl hydroxylase

IKKβ

Inhibitory kappa B kinase beta

IKK

Inhibitor of kappa B kinase

IκB

Inhibitor of NF-κB

HPT axis

Hypothalamic-pituitary-thyroid axis

TNF-α

Tumor necrosis factor-α

HPA axis

Hypothalamo–pituitary–adrenal axis

IL-11

Interleukin-11

RANK

Receptor activator for nuclear factor-κ

OCP

Osteoclast precursor

MDL-1

Myeloid DAP12-associating lectin-1

SFB

Segmented filamentous bacteria

Treg

Regulatory T

IL-4

Interleukin-4

IL-10

Interleukin-10

TGF-β

Transforming growth factor-β

GUS

Gut microbiota β-glucuronidase

GI

Gastrointestinal

FDA

Food and Drug Administration

S1PR1

Sphingosine-1-phosphate receptor 1

BM

Bone marrow

CXCR3

C-X-C motif chemokine receptor 3

CCL20

C-C motif chemokine ligand 20

Wnt

Wingless-related integration site

5-HT

Serotonin

DSS

Dextran Sulfate Sodium Salt

Tph1

Rate-limiting enzyme tryptophan hydroxylase 1

OVX

Oophorectomy

MrOS

Osteoporotic Fractures in Men Study

BMD

Bone Mineral Density

SCFAs

Short-chain fatty acids

CTX-I

(C-terminal telopeptide of type I collagen)

LPS

Lipopolysaccharide

TLR4

Toll-like receptor 4

AACE/ACE

Association for the Advancement of Computing in Education

PMO

Postmenopausal Osteoporosis

CLA

Conjugated linoleic acid

FLA

Flaxseed

Author contributions

All the authors materially participated in the research and article preparation. The roles of all the authors are as follows: Yajun Qiao, Huimin Zheng and Ruiying Cheng: Writing-Original Draft, Drawing. Lin Rong, Juan Guo, Guoqiang Li, Tingting Gao, Lixin Wei: Writing-Review & Editing. Zhongshu Shan, Tingting Gao, Hongtao Bi: Project administration and funding acquisition.

Funding

This work was supported by the Natural Science Foundation of China (Grant No. 82171863), the Innovation Platform Program of Qinghai Province (2020-ZJ-T08), and the Tianfu Emei Project of Sichuan Province.

Data availability

No datasets were generated or analysed during the current study.

Declarations

Ethics approval and consent to participate

Not applicable.

Consent for publication

Not applicable.

Competing interests

The authors declare no competing interests.