Excessive generation of reactive oxygen species (ROS) can lead to oxidative-antioxidative imbalance in the organism, resulting in oxidative stress. Hematopoietic stem/progenitor cells (HSPCs) exhibit high sensitivity to changes in ROS levels, and high levels of ROS can impair self-renewal capacity of HSPCs, leading to oxidative damage and even death. Wnt/β-catenin signaling pathway regulates hematopoiesis and plays an important role in determining the fate of stem cells, such as self-renewal, proliferation and differentiation of HSPCs. Studies have shown that Wnt/β-catenin signaling pathway is also closely related to oxidative stress. This article summarizes the relevant literature, and reviews the role of Wnt/β-catenin signaling pathway in oxidative stress, its impact on hematopoietic system, and the current research status of related mechanisms.

To prepare clinical-grade human umbilical cord-derived mesenchymal stem cells (hUC-MSC) with high expression of hematopoietic supporting factors and evaluate their stem cell characteristics.

To report the clinical characteristics, diagnosis, treatment and prognosis of one patient with primary cutaneous CD8+ aggressive epidermotropic cytotoxic T-cell lymphoma (CD8+ PCAECTL), and to strengthen the understanding of this extremely rare type of lymphoma.

The healing of the tendon-bone interface is a complex dynamic process involving the interaction of multiple cellular and molecular signaling pathways. Bone mesenchymal stem cells (BMSCs) have the potential to differentiate into various types of cells, including osteoblasts, chondrocytes and adipocytes, etc., and have the potential to regenerate damaged tissues. They are potential seed cells for promoting tendon-bone healing. How to precisely regulate the proliferation and differentiation of BMSCs to accelerate the process of tendon-bone healing is a current research hotspot. Monomers of traditional Chinese medicine can promote tendon-bone healing by regulating signaling pathways such as Wnt/β-catenin and BMP/Smad to induce osteogenic and chondrogenic differentiation of BMSCs. This article reviews from several aspects such as the regulatory role of related signaling pathways on tendine-bone healing, traditional Chinese medicine monomers and their mechanism of regulating BMSCs to promote tendine-bone healing in order to providing new ideas for promoting tendine-bone healing.

Calvarial stem cells are essential for maintaining the health and function of the craniofacial complex and the central nervous system. Under physiological conditions, these stem cells primarily reside within specialized microenvironments known as stem cell niches, located in the bone marrow, periosteum, and sutures of cranial bones. The heterogeneous cellular populations within the microenvironment dynamically regulate the quantity and function of stem cells. Due to their distinct spatial distribution, these stem cells exhibit unique functional characteristics and play crucial roles in the development and progression of various diseases, as well as in relevant therapeutic applications. Herein, we summarize the latest research advances concerning various types of calvarial stem cells, elaborating on their respective functions, microenvironmental regulation, and therapeutic potential, thereby providing new perspectives for both basic research and clinical applications in this field.

To explore the mechanism by which Tougu Xiaotong Capsule (TXC) promotes chondrogenic differentiation and cartilage repair in mice with osteoarthritis (OA).

Steroid-induced osteonecrosis of femoral head (SONFH) is a disease characterized by femoral head collapse and local pain caused by excessive use of glucocorticoids. Peroxisome proliferator-activated receptor-γ (PPARγ) is mainly expressed in adipose tissue. Wnt/β-catenin, AMPK and other related signaling pathways play an important role in regulating adipocyte differentiation, fatty acid uptake and storage. Bone marrow mesenchymal cells (BMSCs) have the ability to differentiate into adipocytes or osteoblasts, and the use of hormones upregulates PPARγ expression, resulting in BMSCs biased towards adipogenic differentiation. The increase of adipocytes affects the blood supply and metabolism of the femoral head, and the decrease of osteoblasts leads to the loss of trabecular bone, which eventually leads to partial or total ischemic necrosis and collapse of the femoral head. MicroRNAs (miRNAs) are a class of short non-coding RNAs that regulate gene expression by inhibiting the transcription or translation of target genes, thereby affecting cell function and disease progression. Studies have shown that miRNAs affect the progression of SONFH by regulating PPARγ lipid metabolism-related signaling pathways. Therefore, it may be an accurate and feasible SONFH treatment strategy to regulate adipogenic-osteoblast differentiation in BMSCs by targeted intervention of miRNA differential expression to improve lipid metabolism. In this paper, the miRNA-mediated PPARγ-related signaling pathways were classified and summarized to clarify their effects on lipid metabolism in SONFH, providing a theoretical reference for miRNA targeted therapy of SONFH, and then providing scientific evidence for SONFH precision medicine.

Mitochondrial metabolism is crucial for providing energy and heme precursors during erythroid development. Oxoglutarate dehydrogenase complex (OGDC) is a key enzyme in the mitochondrial tricarboxylic acid (TCA) cycle, and its level gradually increases during erythroid development, indicating its significant role in erythroid development. The aim of the present study was to explore the role and mechanism of OGDC in erythroid development. In this study, we treated erythroid progenitor cells with CPI-613, a novel lipoic acid analog that competitively inhibits OGDC. The results showed that CPI-613 inhibited erythropoietin (EPO)-induced differentiation and enucleation of human CD34+ hematopoietic stem cells into erythroid cells, suppressed cell proliferation, and induced apoptosis. The results of in vivo experiments showed that CPI-613 also hindered the recovery of mice from acute hemolytic anemia. Further mechanism research results showed that CPI-613 increased reactive oxygen species (ROS) in erythroid progenitor cells, inhibited mitochondrial respiration, caused mitochondrial damage, and suppressed heme synthesis, thereby inhibiting erythroid differentiation. Clinical research results showed that oxoglutarate dehydrogenase (OGDH) protein expression levels were up-regulated in bone marrow cells of polycythemia vera (PV) patients. Treatment with CPI-613 significantly inhibited the excessive proliferation and differentiation of erythroid progenitor cells of the PV patients. These findings demonstrates the critical role of OGDC in normal erythroid development, suggesting that inhibiting its activity could be a novel therapeutic strategy for treating PV.

Objective:To investigate the impact of Waardenburg syndrome（WS） -associated Sox10 p.S100Rfs*9 mutation on inner ear function and its mechanism in noise-induced hearing impairment. Methods:A mice model carrying the Sox10 p.S100Rfs*9 mutation was established using CRISPR-Cas9 gene editing technology. Auditory phenotypes were assessed under baseline conditions and after noise exposure（96 dB SPL, 2 hours）. Auditory brainstem response（ABR） tests were performed to evaluate hearing function, combined with immunofluorescence staining of cochlear basilar membrane whole-mounts to observe hair cells and ribbon synapses. Transcriptome sequencing was conducted to analyze molecular mechanisms. Results:Sox10 p.S100Rfs*9 heterozygous mice exhibited normal hearing thresholds with characteristic ventral pigmentation abnormalities under baseline conditions. Following noise exposure, mutant mice showed significantly higher ABR thresholds at 24 000 Hz compared to wild-type controls（[60.00±6.12]vs[48.13±4.28]dB SPL, P<0.000 1）, and a significant reduction in ribbon synapses（CtBP2-positive puncta） in the basal turn（[55.0±2.3]vs[64.8±3.3]per inner hair cell, P=0.006 6）, while hair cell morphology and number remained intact. Transcriptome analysis revealed altered expression of genes involved in immune regulation, membrane structures, ion channels, and neuroactive ligand-receptor interactions. Conclusion:The Sox10 p.S100Rfs*9 mutation does not alter baseline hearing function but significantly increases inner ear susceptibility to noise damage, primarily manifested as enhanced ribbon synapse vulnerability, especially in high-frequency regions. This gene-environment interaction reveals that Sox10 haploinsufficiency may compromise noise tolerance by affecting synaptic stability and inner ear protective mechanisms. These findings provide new perspectives on the phenotypic heterogeneity in WS patients and theoretical basis for individualized noise protection strategies for patients carrying SOX10 mutations.

Objective To investigate the effects and underlying mechanisms of human placental chorionic plate-derived mesenchymal stem cells (hpcMSCs) on cognitive dysfunction, neuroinflammation, neuronal damage and synaptic plasticity in a mouse model of stroke. Methods A mouse model of middle cerebral artery occlusion (MCAO) was adopted. The mice were randomly divided into three groups: sham operation group, MCAO group and hpcMSCs treatment group, with seven mice in each group. The hpcMSCs treatment group received hpcMSCs transplantation on the 1st, 3rd and 10th day after MCAO. One month after MCAO, the cognitive ability of the mice was evaluated by Morris water maze and Y maze behavioral tests; the morphological changes and synaptic functions of hippocampal neurons were analyzed by HE staining, Nissl staining, Golgi staining and immunofluorescence staining techniques; the density and activation status of microglia was analyzed by Fluorescent labeling method; the levels of tumor necrosis factor α (TNF-α), interleukin 1β (IL-1β) and IL-6 in brain tissue were analyzed by ELISA; the expressions of phosphorylated-mitogen-activated protein kinase kinase 1 (p-MEK1), phosphorylated-extracellular regulated protein kinase (p-ERK) and phosphorylated-cAMP-response element binding protein (p-CREB) and other proteins related to neuroprotection in the signal pathways were detected by Western blotting; and electrophysiological detection was performed using hippocampal slices in vitro. Results Compared with the MCAO group, mice in the hpcMSCs treatment group showed significant improvements, including improved cognitive ability, alleviated neuroinflammation (demonstrated by reduced microglial activation and decreased levels of inflammatory factors TNF-α, IL-1β and IL-6), and increased neuronal density with normalized morphology of neurons in the hippocampal CA1 region. The treatment group also demonstrated a significantly increased number of Nissl-positive cells and density of dendritic spines of hippocampal neurons, along with restored frequency of miniature excitatory postsynaptic potential (mEPSP). Moreover, hpcMSCs treatment significantly increased the expression levels of p-MEK1, p-ERK and p-CREB in the hippocampus. Conclusion Transplantation of hpcMSCs ameliorates cognitive dysfunction and hippocampal neuronal injury in stroke mice through the reduction of neuroinflammation, restoration of hippocampal neuronal function, promotion of synaptic plasticity and activation of the MEK/ERK/CREB signaling pathway. These findings suggest a new potential therapeutic approach for post-stroke neural repair.

Airway basal cells (BCs) are adult stem cells in the airways, playing a crucial role in airway renewal and post-injury regeneration and repair. Chronic obstructive pulmonary disease (COPD) is characterized by persistent airway inflammation and remodeling leading to incompletely reversible airflow limitation, significantly impairing patients' quality of life. COPD-related pathogenic factors can influence the genetic expression, proliferation, differentiation, and inflammatory regulation of BCs by altering their gene expression and molecular phenotypes, ultimately contributing to COPD-associated pathological changes. This article systematically reviews the changes in BCs during COPD progression and explores the potential and prospects of targeting BCs as a therapeutic approach for COPD from the perspective of regenerative medicine.

This study primarily investigates the effect of Liuwei Dihuang Pills on the activation and proliferation of female germline stem cells(FGSCs) in the ovaries and cortex of mice with premature ovarian failure(POF), and how it improves ovarian function. ICR mice were randomly divided into the control group, model group, Liuwei Dihuang Pills group, Liuwei Dihuang Pills double-dose group, and estradiol valerate group. A mouse model of POF was established by intraperitoneal injection of cyclophosphamide. After successful modeling, the mice were treated with Liuwei Dihuang Pills or estradiol valerate for 28 days. Vaginal smears were prepared to observe the estrous cycle and body weight. After the last administration, mice were sacrificed and sampled. Serum levels of estradiol(E_2), follicle-stimulating hormone(FSH), luteinizing hormone(LH), and anti-Müllerian hormone(AMH) were measured by enzyme-linked immunosorbent assay(ELISA). Hematoxylin-eosin(HE) staining was used to observe ovarian morphology and to count follicles at all stages to evaluate ovarian function. Immunohistochemistry was used to detect the expression of mouse vasa homolog(MVH), a marker of ovarian FGSCs. Immunofluorescence staining, using co-labeling of MVH and proliferating cell nuclear antigen(PCNA), was used to detect the expression and localization of specific markers of FGSCs. Western blot was employed to assess the protein expression of MVH, octamer-binding transcription factor 4(Oct4), and PCNA in the ovaries. The results showed that compared with the control group, the model group exhibited disordered estrous cycles, decreased ovarian index, increased atretic follicles, and a reduced number of follicles at all stages. FSH and LH levels were significantly elevated, while AMH and E_2 levels were significantly reduced, indicating the success of the model. After treatment with Liuwei Dihuang Pills or estradiol valerate, hormone levels improved, the number of atretic follicles decreased, and the number of follicles at all stages increased. MVH marker protein and PCNA proliferative protein expression in ovarian tissue also increased. These results suggest that Liuwei Dihuang Pills regulate estrous cycles and hormone disorders in POF mice, promote the proliferation of FGSCs, improve follicular development in POF mice, and enhance ovarian function.

Dental-derived mesenchymal stem cells (DMSC), owing to their self-renewal capacity and multi-lineage differentiation potential, have demonstrated significant promise in the field of oral tissue regeneration. Mitochondria, as essential organelles within cells, can not only be responsible for energy metabolism but also influence the maintenance of stem cell stemness and the direction of differentiation by regulating cellular metabolic processes, apoptosis, and signal transduction pathways. In recent years, the mechanisms by which mitochondria participating in the osteogenic and odontogenic differentiation of DMSC have gradually become a research focus. This paper reviews the regulatory mechanisms of mitochondria in the osteogenic and odontogenic differentiation of DMSC, with emphasis on how mitochondrial energy metabolism (oxidative phosphorylation and glycolysis) and quality control processes (namely biogenesis, dynamics as fission and fusion, and autophagy) modulate the differentiation of DMSC. The aim is to provide a theoretical basis and reference for the research and application of dental pulp regeneration, periodontal tissue regeneration, and other aspects of oral tissue engineering.

Objective: To investigate the effects and underlying mechanisms of Stat3 knockout in donor T cells on acute gastrointestinal graft-versus-host disease (GI-aGVHD) . Methods: BALB/c mice were exposed to lethal irradiation and transplanted with bone marrow and spleen cells from BALB/c mice (syngeneic control group), C57BL/6 mice (wild-type T cell group, WT group), or C57BL/6J-Stat3(em1cyagen) mice (Stat3 gene knockout T cell group, Stat3-KO group) via tail vein injection to establish the aGVHD model. The survival rate, body weight changes, and clinical scores of mice were monitored. Cytometric bead array (CBA) was used to detect the concentrations of serum cytokines. Lymphocytes were isolated from tissues for flow cytometric analysis. H&E staining was performed to observe intestinal pathological changes. FITC-dextran assay was conducted to assess intestinal permeability. Immunohistochemistry was used to evaluate the expression of Ki67 and Muc2. Real-Time Quantitative Reverse Transcription PCR (qRT-PCR) was employed to analyze the gene expression levels of Olfm4, Lysozyme, and Muc2 in the small intestine. Metabolomics was conducted to detect metabolites in serum and intestinal tissues. An in vitro GI-aGVHD organoid model was established by coculturing intestinal organoids with allogeneic T cells, where the number and area of small intestinal organoids were recorded. The GVL effect was assessed using luciferase-transfected ALL cells (ALL/Luc) and bioluminescent imaging. Results: Compared with the WT group, Stat3 knockout T cells alleviated body weight loss, reduced symptoms-such as hunchback and diarrhea-in mice, improved survival rate (P<0.05), and reduced serum interleukin (IL) -2, IL-6, interferon-γ, tumor necrosis factor-α, IL-17A, and IL-10 levels (all P<0.05), intestinal inflammatory cell infiltration (P<0.05), and intestinal mucosal permeability. Further, Muc2 and Ki67 expression levels in the small intestine of the Stat3 knockout group were markedly increased, and Olfm4, Lysozyme, and Muc2 gene expression levels were significantly increased (all P<0.05). In vitro, the Stat3 knockout group demonstrated better organoid development than the WT group. Metabolomic analyses indicated that Stat3 knockout in T cells may affect the pathways associated with bile acid secretion and unsaturated fatty acids. ALL/Luc cells in the GVL mouse model proliferated rapidly in the TCD-BM group; however, 80% of the mice in the Stat3-KO group survived tumor-free for >100 days (P<0.05) . Conclusion: Knocking out Stat3 in graft T cells reduces T cell damage to intestinal stem cells, thereby ultimately alleviating GI-aGVHD while maintaining a stable GVL effect.

Objective: To explore the role and mechanism of Prussian blue nanoparticle (PBNP) in the apoptosis of mouse adipose-derived stem cells (ADSCs) treated with hydrogen peroxide, providing a reference for chronic wound treatment. Methods: This research was an experimental research. PBNP with a cubic micromorphology was synthesized via the hydrothermal method. ADSCs were isolated from 6 male 6-8 weeks old Institute of Cancer Research mice using enzymatic digestion. ADSCs were divided into control group with normal culture, hydrogen peroxide group treated with hydrogen peroxide at final molarity of 200 μmol/L, and low PBNP group and high PBNP group pretreated with PBNP at final mass concentration of 10 and 20 μg/mL respectively and then treated as that in hydrogen peroxide group. After 24 h of culture, the reactive oxygen species (ROS) level was detected by fluorescence probe method, the superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), catalase (CAT), malondialdehyde (MDA) levels, and lactate dehydrogenase (LDH) release rate were measured by colorimetric method, the cell survival rate was assessed by cell counting kit-8, and the protein expression levels of B-cell lymphoma-2 (Bcl-2), Bcl-2-associated X protein (Bax), cytochrome C (Cyt-C), cleaved cysteinyl aspartate specific protease-3 (caspase-3), cleaved caspase-9, phosphatidylinositide 3-kinase (PI3K), phospho-PI3K (p-PI3K), protein kinase B (Akt), and phospho-Akt (p-Akt) were detected by Western blotting, with ratios of p-PI3K/PI3K and p-Akt/Akt being calculated. Another batch of ADSCs were divided into control group, hydrogen peroxide group, high PBNP group, which were treated as before, and N-acetyl-L-cysteine (NAC) group, high PBNP+LY294002 group, and high PBNP+MK-2206 group pretreated with NAC at final molarity of 5 mmol/L, PBNP at final mass concentration of 20 μg/mL and LY294002 at final molarity of 10 μmol/L, and PBNP at final mass concentration of 20 μg/mL and MK-2206 at final molarity of 100 μmol/L, respectively, and then treated as that in hydrogen peroxide group. After 24 h of culture, the p-PI3K/PI3K and p-Akt/Akt ratios were detected and calculated, and protein expression levels of Bcl-2, Bax, Cyt-C, cleaved caspase-3, and cleaved caspase-9 were measured as before. There were 3 samples in all experiments. Results: After 24 h of culture, the ROS level in cells in hydrogen peroxide group was 29.0±1.1, which was significantly higher than 2.6±1.1 in control group, 16.5±0.9 in low PBNP group, and 5.3±0.9 in high PBNP group (with P values all <0.05). Compared with those in hydrogen peroxide group, the levels of SOD, CAT, and GSH-Px, the cell survival rate, the Bcl-2 protein expression level, and the ratios of p-PI3K/PI3K and p-Akt/Akt were markedly increased in cells in control group, low PBNP group, and high PBNP group (P<0.05), the MDA level and LDH release rate in cells in control group and high PBNP group and the LDH release rate in cells in low PBNP group were significantly decreased (P<0.05), and the protein expression levels of Bax, Cyt-C, cleaved caspase-9, and cleaved caspase-3 in cells in control group, low PBNP group, and high PBNP group were significantly decreased (P<0.05). After 24 h of culture, compared with those in hydrogen peroxide group, the ratios of p-PI3K/PI3K and p-Akt/Akt, as well as Bcl-2 protein expression level were significantly increased in cells in control group, NAC group, and high PBNP group (P<0.05), while the protein expression levels of Bax, Cyt-C, cleaved caspase-9, and cleaved caspase-3 were significantly decreased (P<0.05). Compared with those in high PBNP group, the ratios of p-PI3K/PI3K and p-Akt/Akt, as well as Bcl-2 protein expression level were significantly decreased in cells in high PBNP+LY-294002 group and high PBNP+MK-2206 group (P<0.05), while the protein expression levels of Bax, Cyt-C, cleaved caspase-9, and cleaved caspase-3 were significantly increased (P<0.05). Conclusions: PBNP can inhibit apoptosis of mouse ADSCs caused by oxidative stress through activating the PI3K/Akt signaling pathway and reducing the expression level of apoptosis-related proteins in cells.

Objective To investigate the effects of cucurbitacin B (CucB) on alleviating skin lesions and inflammation in psoriasis mice via the cGAS-STING signaling pathway. Methods The expression of genes associated with the cGAS-STING signaling pathway in psoriatic lesions and non-lesional skin was analyzed, and hallmark gene set enrichment analysis was performed. The cytotoxicity of CucB on BMDMs was evaluated using the CCK-8 assay. The expression levels of genes and proteins related to the cGAS-STING signaling pathway, along with the secretion of inflammatory cytokines, were measured at different concentrations of CucB using quantitative PCR, Western blotting, and ELISA. Imiquimod-induced psoriasis BALB/c mice were divided into four groups: normal group, model group, low-dose CucB group [0.1 mg/ (kg.d)], and high-dose CucB group [0.4 mg/ (kg.d)], with five mice per group. PASI scoring was performed to assess the severity of psoriasis after 6 days of treatment, and HE staining was conducted to observe pathological damage. Meanwhile, the mRNA levels of inflammatory cytokines and their secretion were detected by qPCR and ELISA. Results Most cGAS-STING signaling-related genes were upregulated in lesional skin of psoriasis patients, and the hallmark gene set enrichment analysis revealed that the most significantly upregulated genes were primarily associated with immune response signaling pathways. CucB inhibited dsDNA-induced phosphorylation of interferon regulatory factor 3 (IRF3) and STING proteins in both bone-marrow derived macrophages(BMDMs) and THP-1 cells. CucB also suppressed dsDNA-induced mRNA expression of IFNB1, TNF, IFIT1, CXCL10, ISG15, and reduced the secretion of cytokines such as IFN-β, IL-1β, and TNF-α in THP-1 cells. In the imiquimod-induced psoriasis mouse model, CucB treatment reduced psoriatic symptoms, alleviated skin lesions, and attenuated inflammation. ELISA and qPCR results showed that CucB significantly reduced serum secretion levels of IL-6, TNF-α, and IL-1β, as well as the mRNA levels of IL23A, IL1B, IL6, TNF, and IFNB1. Conclusion CucB inhibits cytoplasmic DNA-induced activationc of the GAS-STING pathway. CucB significantly attenuates skin lesions and inflammation in IMQ-induced psoriatic mice, and the potential molecular mechanism may be related to the down-regulation of the cGAS-STING pathway.

To review recent advances in the application of hair transplantation in wound healing and scar repair in special areas.

Fanconi anemia (FA) is a hereditary bone marrow failure syndrome that is characterized by genomic instability and heightened sensitivity to DNA cross-linking agents. In recent years, the CRISPR-Cas9 technology has exhibited groundbreaking progress in the field of gene therapy for FA. The traditional CRISPR-Cas9 technology has been successfully applied in FA gene editing. Further, single-base editing technology, based on the CRISPR/Cas9 system, performs precise and efficient gene repair for prevalent gene mutations in patients with FA. The prime editing technology provides new possibilities for gene editing; however, its application in FA has not been initiated. Despite significant advancements in FA gene editing technology, several challenges remain, including the collection of sufficient hematopoietic stem cells, the risk of increased tumorigenesis postgene editing, chromosomal instability, and off-target effects. Future research is recommended to focus on optimizing sgRNA and Cas9 nucleases, designing stricter PAM sequences to reduce off-target effects, and devising personalized gene editing strategies. Further, ethical and regulatory issues as well as long-term follow-ups are crucial priorities for future gene editing work. With continuous technological advancements and in-depth clinical trials, we expect more breakthroughs in FA treatment using the CRISPR-Cas9 technology in the future. This article reviews the latest research progress of CRISPR technology in FA treatment and analyzes the advantages and disadvantages of this technology in FA gene therapy.

A 20-year-old male patient with T-lymphoblastic lymphoma/leukemia received 9/10 human leukocyte antigen-compatible unrelated peripheral blood stem cell transplantation. He was transplanted with 5.91×10(8) mononuclear cells/kg and 2.88×10(6) CD34(+) cells/kg, and neutrophil engraftment was obtained at +11 days and platelet engraftment at +9 days. After transplantation, he presented with repeatedly increased serum creatinine levels, BK virus (BKV) -associated hemorrhagic cystitis, and BKV viremia. BK virus nephropathy was diagnosed based on renal biopsy and metagenomic next-generation sequencing. After adjusting the immunosuppressant, intravenous immunoglobulin, and donor lymphocyte infusion treatment, the patient's renal function deteriorated progressively, and he eventually died of multiple organ failure at +289 days.

This study aims to elucidate the role and mechanism of clematichinenoside AR(CAR) in protecting bone marrow mesenchymal stem cells(BMSCs) from hypoxia-induced apoptosis. BMSCs were isolated by the bone fragment method and identified by flow cytometry. Cells were cultured under normal conditions(37℃, 5% CO_2) and hypoxic conditions(37℃, 90% N_2, 5% CO_2) and treated with CAR. The BMSCs were classified into eight groups: control(normal conditions), CAR(normal conditions + CAR), hypoxia 24 h, hypoxia 24 h + CAR, hypoxia 48 h, hypoxia 48 h + CAR, hypoxia 72 h, and hypoxia 72 h + CAR. The cell counting kit-8(CCK-8) assay and terminal-deoxynucleoitidyl transferase mediated nick end labeling(TUNEL) were employed to measure cell proliferation and apoptosis, respectively. The number of mitochondria and mitochondrial membrane potential were measured by MitoTracker®Red CM-H2XRo staining and JC-1 staining, respectively. The level of reactive oxygen species(ROS) was measured with the DCFH-DA fluorescence probe. The protein levels of B-cell lymphoma-2 associated X protein(BAX), caspase-3, and optic atrophy 1(OPA1) were determined by Western blot. The results demonstrated that CAR significantly increased cell proliferation. Compared with the control group, the hypoxia groups showed increased apoptosis rates, reduced mitochondria, elevated ROS levels, decreased mitochondrial membrane potential, upregulated expression of BAX and caspase-3, and downregulated expression of OPA1. In comparison to the corresponding hypoxia groups, CAR intervention significantly decreased the apoptosis rate, increased mitochondria, reduced ROS levels, elevated mitochondrial membrane potential, downregulated the expression of BAX and caspase-3, and upregulated the expression of OPA1. Therefore, it can be concluded that CAR may exert an anti-apoptotic effect on BMSCs under hypoxic conditions by regulating OPA1 to maintain mitochondrial homeostasis.