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. 2026 May 8;50(4):101061. doi: 10.1016/j.jgr.2026.101061

Li-ginseng powder alleviates cancer cachexia in mice by regulating the ubiquitin-proteasome pathway and reducing inflammation

Xinghui Jin a,b,1, Wenyuan Liu a,1, Gang-Ao Li a,b, Ya-Ni Wang a, Kwang-Il To a, Yang Li a,b,⁎, Ying-Hua Jin a,b,⁎
PMCID: PMC13323883  PMID: 42395026

Abstract

Background

As a debilitating syndrome, cancer cachexia (CC) manifests as ongoing weight reduction and skeletal muscle atrophy, which severely compromise patients’ well-being and life expectancy, with no approved treatment available to date. Rare ginsenosides such as Rh2, Rg5, Rk1, and Rh4 have been reported to modulate Nuclear factor kappa-B (NF-κB) and Signal Transducer and Activator of Transcription 3 (STAT3) activity and attenuate inflammatory signaling pathways implicated in CC progression. Li-Ginseng powder (LGP), a specially processed Panax ginseng enriched in rare ginsenosides, including Rk1, Rk3, Rh4, Rg3, and Rg5 represents a potential therapeutic candidate for CC.

Methods

The anti-cachexia effects of LGP were evaluated in a BALB/c mouse model of CC and in a cellular CC model using mouse myoblast C2C12 cells. Body weight, skeletal muscle atrophy, and histopathological analyses were performed to assess in vivo efficacy. Network pharmacology was applied to predict key regulatory pathways, and mechanistic validation was conducted using Western blotting, immunohistochemistry, and Enzyme-linked immunosorbent assay.

Results

LGP treatment significantly attenuated body weight loss and skeletal muscle atrophy in CC mice. Mechanistically, LGP suppressed activation of the ubiquitin-proteasome pathway in the gastrocnemius muscle and reduced systemic and local inflammatory responses. Network pharmacology analysis identified NF-κB and STAT3 signaling as major targets of LGP, which was further confirmed in both muscle tissues and C2C12 cells. Consistently, LGP alleviated myotube atrophy and inhibited UPP, NF-κB, and STAT3 activation in vitro.

Conclusion

These findings demonstrate that LGP exerts protective effects against CC by modulating muscle proteolysis and inflammation-related signaling pathways, highlighting its potential as a ginseng-based therapeutic strategy for CC.

Keywords: Cancer cachexia, Li-ginseng powder, Gastrocnemius muscle, UPP, STAT3, NF-κB

Graphical abstract

graphic file with name ga1.jpg

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Abbreviations

CC

Cancer cachexia

Fbxo32

F-box protein 32

IMP

Imperatorin

LGG

Li-Ginseng ginsenoside fraction

LGP

Li-Ginseng powder

NF-κB

Nuclear factor kappa-B

STAT3

Signal Transducer and Activator of Transcription 3

TCM

Tumor-conditioned medium

Trim63

Tripartite motif containing 63

UPP

Ubiquitin-proteasome pathway

1. Introduction

As a syndrome marked by intricate, multifactorial pathogenesis, cancer cachexia (CC) is strongly predictive of poor prognosis. [1]. More than half of all cancer patients are impacted by this syndrome, which is further responsible for approximately 20% of cancer-attributed deaths [2]. The hallmark features of cachexia include systemic inflammation and significant weight loss [3]. Persistent elevation of inflammatory cytokines, such as TNF-α, IL-1β, and IL-6, characterizes this condition [4]. These pro-inflammatory factors activate key transcription factors, Nuclear factor kappa-B (NF-κB) and Signal Transducer and Activator of Transcription 3 (STAT3), driving the progression of cachexia [5,6]. The ubiquitin-proteasome pathway (UPP)-mediated protein degradation is central to skeletal muscle atrophy and subsequent weight loss during cachexia progression [3]. E3 ubiquitin-ligases, particularly Tripartite Motif Containing 63 (Trim63) and F-Box Protein 32 (Fbxo32), play a critical role in muscle atrophy by promoting the degradation of MyoD, a key myogenic differentiation protein, via the UPP [7,8]. Although this syndrome profoundly compromises both quality of life and prognosis, no effective intervention for CC is currently available [9]. Megestrol acetate, a commonly used medication for managing cachexia, primarily increases body fat rather than enhancing skeletal muscle mass, limiting its therapeutic efficacy [10].Panax ginseng—a medicinal plant with deep historical roots in East Asia—has been established as possessing wide-ranging pharmacologic effects [11]. Previous studies, primarily focusing on ginsenosides, suggest that ginseng and its derivatives exhibit potential anti-CC effects by mitigating CC-related symptoms and skeletal muscle atrophy [12]. Ginsenosides are regarded as the principal bioactive constituents of ginseng. Notably, rare ginsenosides—which are present in minimal amounts in raw ginseng (less than 0.1% of total ginsenosides) but become abundant in processed forms such as Li-ginseng (constituting about 60% of total ginsenosides) — demonstrate superior bioactivity and bioavailability compared with common ginsenosides [11]. We have previously reported that rare ginsenosides—including Rh2, Rg5, Rk1, and Rh4—suppress cancer cell proliferation and elicit anti-inflammatory responses via modulation of the NF-κB and STAT3 signaling cascades. [[13], [14], [15]]. Li-Ginseng powder (LGP) is a specially processed form of Panax ginseng, enriched with a variety of rare ginsenosides, including Rk1, Rk3, Rh4, Rg3, and Rg5 [11]. Evidence obtained in our previous study indicates that the active constituents counteract alcoholic liver disease by promoting the metabolic removal of acetaldehyde and sustaining cellular stability [11].

In this study, we demonstrate that LGP effectively alleviates CC symptoms in mice by inhibiting gastrocnemius muscle degradation and myotube atrophy. These achieved primarily through significant downregulation of inflammation, the UPP, and the activation of NF-κB and STAT3.

2. Materials and methods

2.1. Reagents and antibodies

LGP was obtained from Yanbian ADKH Biotechnology Co., Ltd. Li-ginseng ginsenoside fraction (LGG) extracted using the published laboratory procedure [11]. Imperatorin (IMP) (EFEBIO, China, E052319), DMEM high glucose medium (Gibco, USA, 11965092), RPMI 1640 medium (Gibco, USA, 12633012), fetal bovine serum (Gibco, USA, A5670402), horse serum (Solarbio, China, 12633012) were used for cell culture.

The following antibodies were used: Fbxo32 antibody (Proteintech, USA, 67172-1-IG), Trim63 antibody (Proteintech, US, 55456-1-AP), MyoD antibody (Proteintech, USA, 18943-1-AP), STAT3 antibody (Santa cruz, USA, sc-8019), phosphorylated-STAT3 antibody (Santa cruz, USA, sc-8059), p50 antibody (Santa cruz, USA, sc-8414), phosphorylated-p50 antibody (Santa cruz, USA, sc-271908), p65 antibody (Santa cruz, USA, sc-8008), phosphorylated-p65 antibody (Santa cruz, USA, sc-136548), GAPDH antibody (Cell Signaling Technology, USA, 97166T), Pierce Goat Anti-Mouse IgG (Thermo Scientific, USA, 31168) Pierce Goat Anti-Rabbit IgG (Thermo Scientific, USA, 31216)

2.2. Tumor-conditioned medium preparation

CT26 mouse colon cancer cells were obtained from Shanghai Enzyme Research Science and Technology Bioengineering Co., Ltd. Cells were cultured in RPMI 1640 medium supplemented with 10% (v/v) fetal bovine serum, 100 U/mL penicillin, and 100 μg/mL streptomycin, and maintained under conventional incubation conditions (5% CO2, 37 °C, saturated humidity). To prepare tumor-conditioned medium (TCM), CT26 cells were replaced in DMEM medium containing 2% horse serum after they reached the logarithmic growth phase [16]. The culture supernatant was obtained after 48 h, subjected to centrifugation at 1000 rpm for 10 min to remove particulate matter, and the resulting fraction was applied to construct an in vitro CC system.

2.3. CC in vivo model establishment and treatment

Fifty six-week-old male BALB/c mice (20 ± 2g) were purchased from Beijing Vital river Technology Co., Ltd. (SCXK (Jing) 2021-2006). All mice were accommodated in a temperature and humidity-controlled facility, received food and water without restriction, and were entrained to a 12-h photoperiod. CT26 colon cancer cells, suspended at 1.0 × 106 cells/mL, were administered subcutaneously in a 100 μL volume into the right axillary fossa of each mouse. Tumors were evident by approximately one week, and euthanasia was carried out on day 14. Tumor masses were harvested, mechanically dissociated, filtered through a 200-mesh sieve, and processed into a tumor tissue suspension. For the second-generation tumor model, mice were randomly assigned into five groups (n = 8). To establish the CC model, 100 μL of tumor tissue suspension was administered subcutaneously into the right axillary fossa of each mouse across four groups. These groups were designated as follows: Model group (Received daily oral gavage of water), LGP(L) group (Received 85 mg/kg LGP dissolved in water via oral gavage), LGP(H) group (Received a double dose of LGP(L) (170 mg/kg)), IMP group (Received 50 mg/kg IMP dissolved in water via oral gavage) and Control group (non-tumor-bearing) received the same treatment as the Model group. Throughout the study, body weight and food ingestion were documented daily (Fig. 1A). All mice were euthanized on day 18, after which serum and various tissue samples—namely heart, liver, spleen, lungs, kidneys, and gastrocnemius muscle—were excised for subsequent assays.

Fig. 1.

Fig. 1

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LGP significantly improved the condition of CC mice. (A) Establishment and treatment experimental design of CC mice (n = 8); (B) Tumor volume change curve in each group (n = 8); (C) Body weight change curve in each group (n = 8); (D) Food intake change curve in each group (n = 8); (E) Gastrocnemius muscle weight in each group (n = 8); (F) Organs weights (Heart, Kidney and Liver) in each group (n = 8); (G) Images of mouse leg muscles and HE staining of gastrocnemius muscle sections (Scale bar: 200 μm).

2.4. Hematoxylin-eosin staining

The mouse gastrocnemius muscle was harvested, fixed in 4% paraformaldehyde, and embedded in paraffin. Tissue blocks were sectioned at a thickness of 4 μm, mounted onto slides, and baked at 60 °C for 30 min. Following dewaxing and rehydration, the sections were stained with hematoxylin, differentiated, counterstained with eosin, then dehydrated, cleared, and mounted. The stained sections were ultimately examined under a light microscope.

2.5. Immunohistochemistry staining

The paraffin-embedded gastrocnemius muscle sections were dewaxed and subjected to antigen retrieval, followed by blocking with a non-specific staining blocker at room temperature for 20 min. The sections were then incubated overnight at 4 °C with the primary antibody diluted in antibody diluent in a humidified chamber. Biotin-labeled goat anti-rabbit IgG polymer was applied, followed by a 20-min incubation. Subsequently, streptavidin-biotin-peroxidase was added and incubated for another 20 min. The reaction was visualized using diaminobenzidine chromogenic solution, and the process was stopped by rinsing with phosphate-buffered saline. The sections were then counterstained with hematoxylin, dehydrated, cleared, and mounted for microscopic analysis.

2.6. Western blotting

Processed C2C12 cells and gastrocnemius muscle were centrifuged at 12000 rpm for 15 min at 4 °C. The protein concentration was quantified and normalized, followed by denaturation in 5 × loading buffer and boiling for 5 min. Proteins were separated by SDS-PAGE and PVDF membranes. The membranes were blocked with 5% skim milk for 1 h, then incubated overnight at 4 °C with the primary antibodies. The next day, membranes were incubated with the secondary antibody for 1 h. Protein bands were visualized using an ECL detection kit (Thermo Fisher, Waltham, MA, USA). The chemiluminescent signal was detected and quantified using ImageJ 1.53T.

2.7. Network pharmacological analysis

BATMAN-TCM (http://bionet.ncpsb.org.cn/batman-tcm) and PubChem (https://pubchem.ncbi.nlm.nih.gov) databases were used to identify potential targets of Rk1, Rk3, Rh4, Rg3, and Rg5. Targets obtained from both databases were merged to generate a comprehensive target list. A drug-target interaction network was then constructed using Cytoscape. To identify CC-associated targets, the DisGeNET, Gene Cards, and OMIM databases were queried using the keyword "cancer cachexia". Targets from these databases were aggregated and filtered to create a unified disease target set. The intersection of ginsenoside targets and CC-related gene sets was analyzed using Venny 2.1.0, thereby pinpointing candidate targets through which rare ginsenosides may exert anti-CC effects. The STRING database was then employed to generate a protein-protein interaction network, which was further analyzed using Cytoscape to assess the degree of correlation among key targets within the "ginsenoside-cancer cachexia" network.

2.8. Gene ontology and pathway enrichment analysis

Metascape was utilized to perform GO annotation and KEGG pathway enrichment analysis on the identified targets of rare ginsenosides in CC. These investigations were designed to uncover the biological functions and pathway-level mechanisms by which ginsenosides mediate their anti-CC activity.

2.9. Measurement of inflammatory cytokines in serum

Enzyme-linked immunosorbent assay kits for mouse TNF-α, IL-6, and IL-1β (Jianglai Biotechnology Co., Ltd., China) were used to quantify cytokine levels.

2.10. CC in vitro model establishment and treatment

C2C12 mouse myoblasts were obtained from Shanghai EK-BIO Science Co., Ltd. and cultured in DMEM supplemented with 10% (v/v) fetal bovine serum, 100 U/mL penicillin, and 100 μg/mL streptomycin under standard conditions (5% CO2, 37 °C, and saturated humidity). Cell differentiation was induced by replacing the growth medium with DMEM medium containing 2% horse serum once the cells reached the logarithmic growth phase. The medium was changed daily until multinucleated myotubes were fully formed. After differentiation was complete, the culture medium was discarded, and TCM was mixed with normal culture medium at a 1:1 ratio before being added to the culture dish to induce myotube atrophy. Cells were treated with LGP and IMP for 48 h, after which they were collected for further analysis. For the control group, C2C12 cell supernatants were similarly mixed with normal culture medium at a 1:1 ratio.

2.11. Statistical analysis

Experimental data were analyzed using GraphPad Prism 8.0. One-way ANOVA was performed for statistical comparisons, with a p-value<0.05 considered statistically significant. Statistical significance in all figures is indicated as follows: ∗p< 0.05, ∗∗p< 0.01, ∗∗∗p< 0.001, ∗∗∗∗p < 0.0001; #p < 0.05, ##p < 0.01, ###p < 0.001, ####p< 0.0001. Comparisons are specified in each figure legend (e.g., ∗ vs. control, # vs. model group).

3. Result

3.1. LGP treatment significantly improved the condition of CC mice

Treatment commenced on the third day, once the tumor mass became palpable. IMP, was employed as a positive control in vivo study which belongs to naturally sourced has been proved directly inhibiting STAT3 to alleviate CC [16]. Following 2 weeks of LGP administration, CC mice exhibited a significant decrease (p < 0.001, Fig. 1B) in tumor volume and increase (p < 0.0001, Fig. 1C) in body weight compared with model mice. Notably, LGP(H) group exhibited a more pronounced increase (p < 0.0001, Fig. 1C), with body weight approaching that of the control group. Loss of appetite is a contributor to CC-induced weight loss [17]. LGP treatment significantly improved food intake (p < 0.01, Fig. 1D), with a greater effect in LGP(H) group (p < 0.001, Fig. 1D). A hallmark of CC is skeletal muscle atrophy, characterized by progressive muscle degradation and loss of muscle mass [1]. LGP treatment significantly attenuated muscle atrophy, as evidenced by reduced leg muscle wasting (Fig. 1G) and an increase in gastrocnemius muscle weight (p < 0.0001, Fig. 1E). In the model group, muscle fiber atrophy and widened inter-fiber gaps were observed, whereas LGP treatment markedly improved muscle fiber integrity (Fig. 1G). LGP also reversed organ weight abnormalities in CC mice. Heart (p < 0.05, Fig. 1F) and kidney weights (p < 0.05, Fig. 1F) were effectively restored following LGP treatment. Conversely, liver weight, which was elevated in CC mice, was significantly reduced after LGP treatment (p < 0.01, Fig. 1F).

3.2. LGP regulated the UPP related proteins in CC mice

In CC, the UPP is the primary mechanism driving skeletal muscle degradation [18], and the specific E3 ubiquitin ligases, Fbxo32 and Trim63 are upregulated in CC, leading to skeletal muscle atrophy [19]. Immunohistochemical staining was performed to detect Fbxo32 and Trim63 expression in the gastrocnemius muscle. In the model group, both Fbxo32 and Trim63 were significantly upregulated, whereas LGP treatment markedly downregulated their expression (Fig. 2A). These findings were further confirmed by western blotting, which demonstrated a significant reduction in Fbxo32 (p < 0.0001) and Trim63 (p < 0.05) levels following LGP treatment (Fig. 2B–D). Additionally, LGP treatment significantly reduced MyoD degradation in the gastrocnemius muscle (Fig. 2B–E, p < 0.05), with a stronger effect observed in the LGP(H) group (p < 0.001). No significant difference was detected in the IMP group (Fig. 2B–E).

Fig. 2.

Fig. 2

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LGP inhibits gastrocnemius muscle atrophy in CC mice by regulating the UPP related proteins. (A) Immunohistochemical staining of Fbxo32 and Trim63 in gastrocnemius muscle (Scale bar: 200 μm); (B) Western blotting analysis of Fbxo32, Trim63, and MyoD in gastrocnemius muscle; (C-E) Quantification of western blotting for Fbxo32, Trim63 and MyoD (n = 4).

3.3. LGP has numerous potential targets for CC

HPLC analysis of LGG revealed that the rare ginsenosides Rk1, Rk3, Rh4, Rg3, and Rg5 were present at 16.12%, 5.65%, 12.03%, 15.21%, and 22.57% respectively [11]. To identify potential molecular targets of LGP, a database search for Rk1, Rk3, Rh4, Rg3, and Rg5 identified 57, 46, 42, 98, and 88 target proteins, respectively. After removing duplicates, a total of 199 unique ginsenoside-targeting proteins were identified (Fig. 3C). To investigate the involvement of these targets in CC, disease-related proteins were retrieved from DisGeNET (110 targets), GeneCards (2562 targets), and OMIM (496 targets), resulting in a combined dataset of 2891 CC-associated proteins (Fig. 3A). The potential therapeutic targets of rare ginsenosides in CC were determined by identifying the intersection between ginsenoside targets and CC disease targets using the Venny 2.1 tool, yielding 121 overlapping target proteins (Fig. 3B). A protein-protein interaction network was constructed using the STRING database, comprising 121 nodes and 2581 edges. Network analysis performed using Cytoscape identified AKT1, TP53, CASP3, and STAT3 as central nodes, with NF-κB also playing a key role in the network (Fig. 3D).

Fig. 3.

Fig. 3

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Potential targets proteins of LGP in CC (A) Venn diagram of CC-associated targeting proteins (B) Venn diagram of intersection of CC-associated targeting proteins and rare ginsenoside targeting proteins; (C) Network diagram of 199 rare ginsenosides-targeting proteins; (D) Protein-protein interaction network between rare ginsenosides and CC-associated overlapping targets proteins.

3.4. LGP regulated the STAT3, NF-κB and inflammation in CC mice

The 121 overlapping target proteins were analyzed using Metascape for GO analysis and KEGG enrichment analysis. GO analysis revealed that these targets were primarily involved in kinase binding, protein kinase activity, transcription factor binding, and cytokine receptor binding in terms of molecular function (Fig. 4A); Response to hormone, response to inorganic substance, positive regulation of phosphorylation, response to lipopolysaccharide, regulation of neuron death, positive regulation of cell death and positive regulation of cell motility in biological process (Fig. 4B); Membrane raft, side of membrane, transcription regulator complex, transferase complex, and receptor complex in cellular component (Fig. 4C). KEGG pathway enrichment analysis indicated that rare ginsenosides targets in CC were predominantly associated with cancer pathways, lipid and atherosclerosis, the FoxO signaling pathway, proteoglycans in cancer, MAPK signaling pathway, and the NF-κB signaling pathway (Fig. 4D).

Fig. 4.

Fig. 4

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Network pharmacology analysis and following proved experiments (A) Molecular function analysis of GO; (B) Biological process analysis of GO; (C) Cellular component analysis of GO; (D) KEGG enrichment analysis; (E) Serum levels of TNF-α, IL-6 and IL-1β in each group (n = 8); (F) Western blotting analysis of p65, p-p65, p50, p-p50, STAT3 and p-STAT3 (n = 4).

CC is characterized by systemic inflammation, leading to elevated circulating levels of pro-inflammatory factors [20]. LGP treatment significantly reduced serum TNF-α (p < 0.0001), IL-6 (p < 0.0001) and IL-1β (p < 0.0001) (Fig. 4E). Based on network pharmacological analysis, we focused on STAT3 and NF-κB, as these transcription factors play critical roles in CC-associated inflammation. Notably, the NF-κB and STAT3 pathways also regulate the expression of Fbxo32 and Trim63 in the UPP [21,22]. To further assess the regulatory effects of LGP, the expression and activation of NF-κB subunits (p50 and p65) and STAT3 in the gastrocnemius muscle were evaluated. The results showed that the expression levels of p65 (p < 0.001), p-p65 (p < 0.0001), p50 (p < 0.05), p-p50 (p < 0.001), STAT3 (p < 0.05) and p-STAT3 (p < 0.01) were significantly downregulated after the treatment of LGP (Fig. 4F).

3.5. LGG alleviated myotube atrophy and regulated UPP, NF-κB and STAT3 signaling pathways in C2C12 cells

LGG is the ginsenosides fraction of LGP which is the main active components of LGP and convenient to use in cell-base assay because of easily soluble. Differentiation of C2C12 mouse myoblasts resulted in cell fusion on the fifth day, forming myotubes (Fig. 5A). To evaluate whether LGG exerts a protective effect on myotubes under CC conditions, C2C12 cells were induced with TCM and treated with 15 μg/mL or 20 μg/mL LGG for 48h respectively. The IMP concentration (20 μg/mL) was selected based on a previous report [16]. HE staining results revealed that the TCM-induced myotubes exhibited sparse distribution and reduced diameter, indicating atrophy. However, LGG treatment significantly improved the atrophy of myotubes (Fig. 5B). After LGG treatment, the expression levels of Fbxo32 (p < 0.01) and Trim63 (p < 0.05) were significantly downregulated while MyoD (p < 0.01) were significantly upregulated in the TCM-induced myotubes (Fig. 5C and E); Additionally, p50 (p < 0.05), p-p50 (p < 0.001), STAT3 (p < 0.05) and p-STAT3 (p < 0.001, Fig. 5C and D). These effects were consistent with findings in gastrocnemius muscle.

Fig. 5.

Fig. 5

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LGG inhibits the atrophy and regulated the CC related pathway in C2C12 cells (A) Differentiation of C2C12 myoblasts into myotubes induced by horse serum (Scale bar: 100 μm); (B) H&E staining of myotubes (Scale bar: 100 μm); (C) Western blotting analysis of p50, p-p50, STAT3, p-STAT3, Fbxo32, Trim63 and MyoD (D) Quantification of western blotting results for p50, p-p50, STAT3 and p-STAT3 (E) Quantification of western blotting results for Fbxo32, Trim63 and MyoD (n = 4).

4. Discussion

As a complex and multifactorial disorder, CC is defined by progressive muscle wasting. The syndrome afflicts over 50% of individuals with malignancy and severely compromises their physical function and longevity. [23,24]. Although the precise pathogenesis of cachexia remains unclear, research suggests that tumor secretions, tumor-induced immune responses, and metabolic alterations are closely associated with its development.

Ginsenosides are considered the primary bioactive components in ginseng [25,26]. Rare ginsenosides constitute only a minor fraction of total saponins in unprocessed raw ginseng, yet they exhibit markedly superior bioactivity compared to their common counterparts. [27]. LGP is a specially processed ginseng product rich in rare ginsenosides, with Rg3, Rg5, Rk1, Rk3, and Rh4 making up 60% of the total ginsenoside content. Our previous studies have shown that Rg5 and Rk1 inhibit the NF-κB pathway by targeting Annexin A2 [28], while Rh4 exerts anti-inflammatory effects by suppressing both the NF-κB and STAT3 signaling pathways, along with downregulating proinflammatory cytokines including IL-6, IL-1β, and TNF-α [15]. The median lethal dose (LD50) of LGP was not reached in an acute toxicity study, with no adverse effects observed in mice following administration at up to approximately 134 times the standard therapeutic dose (22.9 g/kg body weight). These results, demonstrating a favorable safety profile, suggest that LGP, a product rich in rare ginsenosides, holds promise as an effective natural therapeutic agent for the treatment of CC.

This study investigated the effects of LGP on CC and its related signaling pathways, both in vivo and in vitro. A second-generation CT26 colon cancer cell-induced cachexia mouse model was established for this investigation. LGP significantly alleviated CC-related symptoms in mice, including weight loss, reduced food intake, and organ condition (Fig. 1). Muscle atrophy, a prominent feature of CC, is closely associated with the UPP [29]. LGP treatment significantly mitigated weight loss and gastrocnemius muscle atrophy in CC mice (Fig. 1B–D). Notably, heart and kidney weights were significantly reduced (Fig. 1E), while the liver weight was markedly increased in CC mice. These results support that mice were on late stage of CC which as a systemic disorder, characterized not only by muscle atrophy but also by mass alterations and functional impairment in multiple organs such as the heart, kidney, and spleen especially in late stage [30,31]. LGP treatment substantially restored the organ weights, underscoring its multifunctional therapeutic potential.

MyoD is one of the most important factors which downregulated in CC synchronize with muscle atrophy [32]. In addition, NF-κB signaling inhibits MyoD transcription and induces skeletal muscle atrophy. [33]. E3 ligases, Fbxo32 and Trim63 accelerate the muscle atrophy and even induce the ubiquitination and degradation of MyoD. Interestingly, the expressions of Fbxo32 and Trim63 were markedly downregulated and the degradation of MyoD was alleviated in the gastrocnemius muscle in LGP treatment CC mice (Fig. 2B–E).

To investigate the underlying mechanism of LGP's effect in CC, network pharmacology was employed to identify the common targets of the major ginsenosides in LGP (Rk1, Rk3, Rh4, Rg3, and Rg5) and their association with CC. TNF, STAT3, and NF-κB were identified as potential key proteins involved in the LGP-mediated regulation of inflammatory processes in CC (Fig. 3). Based on network pharmacology predictions, we focused on inflammation and related signaling pathways. LGP treatment reduced the levels of major inflammatory factors, including TNF-α, IL-6, and IL-1β, in the serum of CC mice (Fig. 4E), suggesting that LGP modulates systemic inflammation.

Furthermore, LGP treatment significantly suppressed the expression and activation of STAT3, as well as the p50 and p65 subunits of NF-κB, in the gastrocnemius muscle (Fig. 4F). An in vitro CC model was also established to explore the protective effects of LGG on muscle in CC. LGG treatment inhibited the expression and activation of STAT3, as well as the p50 and p65 subunits of NF-κB in CC-induced myotube cells (Fig. 5C and D). Additionally, LGP treatment downregulated the expression of Fbxo32 and Trim63 in the myotube cells, further supporting its potential as a therapeutic agent for CC (Fig. 5C and D).

In this study, we observed that LGP improved the condition of CC mice in several ways, with a particular focus on the primary symptom of CC—muscle atrophy. LGP primarily regulated the STAT3 and NF-κB signaling pathways, the UPP, and inflammation to protect against muscle atrophy in CC. Interestingly, the liver weight in model mice was increased compared to the control mice. This change in liver weight may reflect systemic inflammation, which warrants further exploration. In C2C12 cells, MyoD expression in TCM-induced myotube cells was not downregulated. However, treatment with 20 μg/mL LGG upregulated MyoD expression compared to the model cells. These findings suggest that in vitro and in vivo cachexia models do not fully recapitulate identical signaling events. In C2C12 myotubes, the expression of MyoD and its UPP-mediated degradation may reach a dynamic equilibrium, thereby maintaining relatively stable MyoD levels. Notably, LGG treatment inhibited UPP activity, which may contribute to the upregulation of MyoD observed in LGG-treated cells. This hypothesis requires further investigation to confirm its underlying mechanisms. In this study, we focused on the main rare ginsenosides in LGP as the active components involved in the treatment of CC. However, it is important to recognize that LGP also contains other bioactive compounds, such as ginseng polysaccharides and ginseng peptides, which may contribute to its pharmacological effects [27,28]. The exceptional safety and potent anti-CC efficacy of LGP are likely due to the combined actions of these active substances. The exact mechanisms by which these components contribute to LGP's effectiveness in CC progression remain to be further explored and fully elucidated.

In summary, this study explored the beneficial effects of LGP in the CC model, with a particular focus on its mechanism of protecting against muscle atrophy during the CC progression. Our findings provide a foundation for the development of anti-CC therapies based on Panax ginseng and highlight the need for further investigations to fully understand its therapeutic potential in this context.

Ethics statement

This study was approved by the Institutional Animal Ethics Committee of Jilin University (Approval Number YNPZSY2022011).

Conflicts of interest

The authors declare no conflicts of interest.

Acknowledgment

This work was supported by Specific Funding of Development and Reform Commission of Jilin Province (2021FGWCXNLJSSZ01) and The Leading Team of the Changbai Mountain Talent Engineering Project (000009).

Footnotes

Appendix A

Supplementary data to this article can be found online at https://doi.org/10.1016/j.jgr.2026.101061.

Contributor Information

Yang Li, Email: liyang915@jlu.edu.cn.

Ying-Hua Jin, Email: yhjin@jlu.edu.cn.

Appendix A. Supplementary data

The following are the supplementary data to this article:

Multimedia component 1
mmc1.docx (17.5KB, docx)

figs1.

figs1

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