TSP50 promotes hepatocyte proliferation and tumour formation by activating glucose‐6‐phosphate dehydrogenase (G6PD)

Background & AimsHepatocellular carcinoma (HCC) is a common malignant tumour with high morbidity and mortality. Metabolic regulation by oncogenes is necessary for tumour growth. Testes‐specific protease 50 (TSP50) has been found to promote cell proliferation in multiple tumour types. However, the mechanism that TSP50 promotes HCC progression are not known.MethodsHepatocyte proliferation was analysed by MTT and BrdU incorporation after TSP50 transfection. Furthermore, LC‐MS/MS, co‐immunoprecipitation and GST pull‐down assays were performed to analyse protein(s) binding to TSP50. Moreover, the site‐specific mutation of G6PD was used to reveal the key site critical for G6PD acetylation mediated by TSP50. Finally, the role of G6PD K171 acetylation regulated by TSP50 in cell proliferation and tumour formation was investigated.ResultsOur data suggest that the overexpression of TSP50 accelerates hepatocyte proliferation. In addition, G6PD is an important protein that binds to TSP50 in the cytoplasm. TSP50 activates G6PD activity by inhibiting the acetylation of G6PD at the K171 site. In addition, TSP50 promotes the binding of G6PD to SIRT2. Furthermore, the K171ac of G6PD regulated by TSP50 is required for TSP50‐induced cell proliferation in vitro and tumour formation in vivo. Additionally, according to The Cancer Genome Atlas (TCGA) programme, TSP50 and G6PD are negatively correlated with the survival of HCC patients.ConclusionsCollectively, our findings demonstrate that TSP50‐induced cell proliferation and tumour formation are mediated by G6PD K171 acetylation.     

Optogenetic control of YAP cellular localisation and function

YAP, an effector of the Hippo signalling pathway, promotes organ growth and regeneration. Prolonged YAP activation results in uncontrolled proliferation and cancer. Therefore, exogenous regulation of YAP activity has potential translational applications. We present a versatile optogenetic construct (optoYAP) for manipulating YAP localisation, and consequently its activity and function. We attach a LOV2 domain that photocages a nuclear localisation signal (NLS) to the N‐terminus of YAP. In 488 nm light, the LOV2 domain unfolds, exposing the NLS, which shuttles optoYAP into the nucleus. Nuclear import of optoYAP is reversible and tuneable by light intensity. In cell culture, activated optoYAP promotes YAP target gene expression and cell proliferation. Similarly, optofYap can be used in zebrafish embryos to modulate target genes. We demonstrate that optoYAP can override a cell''s response to substrate stiffness to generate anchorage‐independent growth. OptoYAP is functional in both cell culture and in vivo, providing a powerful tool to address basic research questions and therapeutic applications in regeneration and disease.     

Triptolide promotes autophagy to inhibit mesangial cell proliferation in IgA nephropathy via the CARD9/p38 MAPK pathway

BackgroundMesangial cell proliferation is the most basic pathological feature of immunoglobulin A nephropathy (IgAN); however, the specific underlying mechanism and an appropriate therapeutic strategy are yet to be unearthed. This study aimed to investigate the therapeutic effect of triptolide (TP) on IgAN and the mechanism by which TP regulates autophagy and proliferation of mesangial cells through the CARD9/p38 MAPK pathway.MethodsWe established a TP‐treated IgAN mouse model and produced IgA1‐induced human mesangial cells (HMC) and divided them into control, TP, IgAN, and IgAN+TP groups. The levels of mesangial cell proliferation (PCNA, cyclin D1, cell viability, and cell cycle) and autophagy (P62, LC3 II, and autophagy flux rate) were measured, with the autophagy inhibitor 3‐Methyladenine used to explore the relationship between autophagy and proliferation. We observed CARD9 expression in renal biopsies from patients and analyzed its clinical significance. CARD9 siRNA and overexpression plasmids were constructed to investigate the changes in mesangial cell proliferation and autophagy as well as the expression of CARD9 and p‐p38 MAPK/p38 MAPK following TP treatment.ResultsAdministering TP was safe and effectively alleviated mesangial cell proliferation in IgAN mice. Moreover, TP inhibited IgA1‐induced HMC proliferation by promoting autophagy. The high expression of CARD9 in IgAN patients was positively correlated with the severity of HMC proliferation. CARD9/p38 MAPK was involved in the regulation of HMC autophagy and proliferation, and TP promoted autophagy to inhibit HMC proliferation by downregulating the CARD9/p38 MAPK pathway in IgAN.ConclusionTP promotes autophagy to inhibit mesangial cell proliferation in IgAN via the CARD9/p38 MAPK pathway.

The proliferation of glomerular mesangial cells is the most basic pathological feature for IgA nephropathy (IgAN). Galactose‐deficient IgA1 (Gd‐IgA1) immune complex deposition activates mesangial cells to mediate specific intracellular signal transduction, promotes mesangial cell proliferation, and initiates kidney damage. Autophagy is involved in mesangial cell proliferation. CARD9 is a risk gene for IgAN. The triptolide (TP), purified from Tripterygium wilfordii hook F., promotes autophagy to inhibit mesangial cell proliferation in IgAN via the CARD9/p38 MAPK pathway.     

Fibroblast growth factor 10 protects against UVB‐induced skin injury by activating the ERK/YAP signalling pathway

ObjectivesUltraviolet light B (UVB) irradiation can induce skin injury and result in keratinocytes proliferation inhibition. However, the molecular understanding of the repair during UVB‐induced cell proliferation inhibition remains poorly understood. The purpose of this study was to explore the role and potential mechanism of FGF10 in promoting keratinocytes cell cycle and proliferation after UVB injury.Materials and MethodsExpression of FGF10 protein was analysed in skin treated with UVB radiation by immunohistochemistry. The proliferation potential was examined by Immunofluorescence, Western Blot and RT‐PCR under UVB radiation, treated with FGF10 protein or overexpression of FGF10 using adeno‐associated virus. CCK8 kit was used to further detect cell proliferation ability.ResultsWe found that FGF10 is highly expressed in skin treated with UVB. Overexpression of FGF10 has a protective effect against UVB‐induced skin damage by balancing epidermal thickness and enhancing epidermal keratinocytes proliferation. Importantly, FGF10 is found to alleviate UVB‐induced downregulation of YAP activity, then promoting keratinocytes proliferation. Disruption of YAP function, either with the small molecule YAP inhibitor Verteporfin (VP) or YAP small‐interfering RNA (siRNA), largely abolishes the protective activity of FGF10 on epidermal keratinocytes proliferation. Meanwhile, disruption of ERK kinase (MEK) activity with U0126 or ERK siRNA hinder the positive influence of FGF10 on UVB‐induced skin injury.ConclusionFGF10 promotes epidermal keratinocytes proliferation during UVB‐induced skin injury in an ERK/YAP‐dependent manner.

Schematic illustration of the protective effects of FGF10 on keratinocytes under UVB radiation. UVB radiation would cause skin injury and proliferation inhibition. In this study, we revealed that FGF10 promotes keratinocytes cell cycle and proliferation via ERK/YAP signalling pathway.     

Ageing causes an aortic contractile dysfunction phenotype by targeting the expression of members of the extracellular signal‐regulated kinase pathway

The extracellular signal‐regulated kinase (ERK) pathway is a well‐known regulator of vascular smooth muscle cell proliferation, but it also serves as a regulator of caldesmon, which negatively regulates vascular contractility. This study examined whether aortic contractile function requires ERK activation and if this activation is regulated by ageing. Biomechanical experiments revealed that contractile responses to the alpha1‐adrenergic agonist phenylephrine are attenuated specifically in aged mice, which is associated with downregulation of ERK phosphorylation. ERK inhibition attenuates phenylephrine‐induced contractility, indicating that the contractile tone is at least partially ERK‐dependent. To explore the mechanisms of this age‐related downregulation of ERK phosphorylation, we transfected microRNAs, miR‐34a and miR‐137 we have previously shown to increase with ageing and demonstrated that in A7r5 cells, both miRs downregulate the expression of Src and paxillin, known regulators of ERK signalling, as well as ERK phosphorylation. Further studies in aortic tissues transfected with miRs show that miR‐34a but not miR‐137 has a negative effect on mRNA levels of Src and paxillin. Furthermore, ERK phosphorylation is decreased in aortic tissue treated with the Src inhibitor PP2. Increases in miR‐34a and miR‐137 with ageing downregulate the expression of Src and paxillin, leading to impaired ERK signalling and aortic contractile dysfunction.     

ERK/Drp1‐dependent mitochondrial fission contributes to HMGB1‐induced autophagy in pulmonary arterial hypertension

ObjectivesHigh‐mobility group box‐1 (HMGB1) and aberrant mitochondrial fission mediated by excessive activation of GTPase dynamin‐related protein 1 (Drp1) have been found to be elevated in patients with pulmonary arterial hypertension (PAH) and critically implicated in PAH pathogenesis. However, it remains unknown whether Drp1‐mediated mitochondrial fission and which downstream targets of mitochondrial fission mediate HMGB1‐induced pulmonary arterial smooth muscle cells (PASMCs) proliferation and migration leading to vascular remodelling in PAH. This study aims to address these issues.MethodsPrimary cultured PASMCs were obtained from male Sprague‐Dawley (SD) rats. We detected RNA levels by qRT‐PCR, protein levels by Western blotting, cell proliferation by Cell Counting Kit‐8 (CCK‐8) and EdU incorporation assays, migration by wound healing and transwell assays. SD rats were injected with monocrotaline (MCT) to establish PAH. Hemodynamic parameters were measured by closed‐chest right heart catheterization.ResultsHMGB1 increased Drp1 phosphorylation and Drp1‐dependent mitochondrial fragmentation through extracellular signal‐regulated kinases 1/2 (ERK1/2) signalling activation, and subsequently triggered autophagy activation, which further led to bone morphogenetic protein receptor 2 (BMPR2) lysosomal degradation and inhibitor of DNA binding 1 (Id1) downregulation, and eventually promoted PASMCs proliferation/migration. Inhibition of ERK1/2 cascade, knockdown of Drp1 or suppression of autophagy restored HMGB1‐induced reductions of BMPR2 and Id1, and diminished HMGB1‐induced PASMCs proliferation/migration. In addition, pharmacological inhibition of HMGB1 by glycyrrhizin, suppression of mitochondrial fission by Mdivi‐1 or blockage of autophagy by chloroquine prevented PAH development in MCT‐induced rats PAH model.ConclusionsHMGB1 promotes PASMCs proliferation/migration and pulmonary vascular remodelling by activating ERK1/2/Drp1/Autophagy/BMPR2/Id1 axis, suggesting that this cascade might be a potential novel target for management of PAH.     

LRRC8A is essential for volume‐regulated anion channel in smooth muscle cells contributing to cerebrovascular remodeling during hypertension

ObjectivesRecent studies revealed LRRC8A to be an essential component of volume‐regulated anion channel (VRAC), which regulates cellular volume homeostasis. However, evidence for the contribution of LRRC8A‐dependent VRAC activity in vascular smooth muscle cells (VSMCs) is still lacking, and the relevant functional role of LRRC8A in VSMCs remains unknown. The primary goal of this study was to elucidate the role of LRRC8A in VRAC activity in VSMCs and the functional role of LRRC8A in cerebrovascular remodeling during hypertension.Materials and MethodssiRNA‐mediated knockdown and adenovirus‐mediated overexpression of LRRC8A were used to elucidate the electrophysiological properties of LRRC8A in basilar smooth muscle cells (BASMCs). A smooth muscle–specific overexpressing transgenic mouse model was used to investigate the functional role of LRRC8A in cerebrovascular remodeling.ResultsLRRC8A is essential for volume‐regulated chloride current (I _Cl, Vol) in BASMCs. Overexpression of LRRC8A induced a voltage‐dependent Cl⁻ current independently of hypotonic stimulation. LRRC8A regulated BASMCs proliferation through activation of WNK1/PI3K‐p85/AKT axis. Smooth muscle‐specific upregulation of LRRC8A aggravated Angiotensin II‐induced cerebrovascular remodeling in mice.ConclusionsLRRC8A is an essential component of VRAC and is required for cell volume homeostasis during osmotic challenge in BASMCs. Smooth muscle specific overexpression of LRRC8A increases BASMCs proliferation and substantially aggravates basilar artery remodeling, revealing a potential therapeutic target for vascular remodeling in hypertension.

The schematic diagram for LRRC8A role in cerebrovascular remodeling. LRRC8A is an essential component of VRAC in BASMCs. During the challenge of hypertension, the activated LRRC8A channel‐mediated‐Cl⁻ efflux increases WNK1 phosphorylation, which in turn triggers AKT phosphorylation and promotes BASMCs proliferation, eventually exacerbates hypertension‐induced cerebrovascular vascular remodeling.     

B‐cell capacity for differentiation changes with age

BackgroundAge‐related immune deficiencies are thought to be responsible for increased susceptibility to infection in older adults, with alterations in lymphocyte populations becoming more prevalent over time. The loss of humoral immunity in ageing was attributed to the diminished numbers of B cells and the reduced ability to generate immunoglobulin.AimsTo compare the intrinsic B‐cell capacity for differentiation into mature plasma cells (PCs), between young and old donors, using in vitro assays, providing either effective T‐cell help or activation via TLR engagement.MethodsB cells were isolated from healthy individuals, in younger (30–38 years) and older (60–64 years) donors. An in vitro model system of B‐cell differentiation was used, analysing 5 differentiation markers by flow cytometry, under T‐dependent (TD: CD40/BCR stimulation) or T‐independent (TI: TLR7/BCR activation) conditions. Antibody secretion was measured by ELISA and gene expression using qPCR.ResultsTI and TD differentiation resulted in effective proliferation of B cells followed by their differentiation into PC. B‐cell‐executed TI differentiation was faster, all differentiation marker and genes being expressed earlier than under TD differentiation (day 6), although generating less viable cells and lower antibody levels (day 13). Age‐related differences in B‐cell capacity for differentiation were minimal in TD differentiation. In contrast, in TI differentiation age significantly affected proliferation, viability, differentiation, antibody secretion and gene expression, older donors being more efficient.ConclusionAltogether, B‐cell differentiation into PC appeared similar between age groups when provided with T‐cell help, in contrast to TI differentiation, where multiple age‐related changes suggest better capacities in older donors. These new findings may help explain the emergence of autoantibodies in ageing.     

Protectin DX promotes epithelial injury repair and inhibits fibroproliferation partly via ALX/PI3K signalling pathway

Acute respiratory distress syndrome/acute lung injury (ARDS/ALI) is histologically characterized by extensive alveolar barrier disruption and excessive fibroproliferation responses. Protectin DX (PDX) displays anti‐inflammatory and potent inflammation pro‐resolving actions. We sought to investigate whether PDX attenuates LPS (lipopolysaccharide)‐induced lung injury via modulating epithelial cell injury repair, apoptosis and fibroblasts activation. In vivo, PDX was administered intraperitoneally (IP) with 200 ng/per mouse after intratracheal injection of LPS, which remarkedly stimulated proliferation of type II alveolar epithelial cells (AT II cells), reduced the apoptosis of AT II cells, which attenuated lung injury induced by LPS. Moreover, primary type II alveolar cells were isolated and cultured to assess the effects of PDX on wound repair, apoptosis, proliferation and transdifferentiation in vitro. We also investigated the effects of PDX on primary rat lung fibroblast proliferation and myofibroblast differentiation. Our result suggests PDX promotes primary AT II cells wound closure by inducing the proliferation of AT II cells and reducing the apoptosis of AT II cells induced by LPS, and promotes AT II cells transdifferentiation. Furthermore, PDX inhibits transforming growth factor‐β₁ (TGF‐β₁) induced fibroproliferation, fibroblast collagen production and myofibroblast transformation. Furthermore, the effects of PDX on epithelial wound healing and proliferation, fibroblast proliferation and activation partly via the ALX/ PI3K signalling pathway. These data present identify a new mechanism of PDX which targets the airway epithelial cell and fibroproliferation are potential for treatment of ARDS/ALI.     

PIP2 depletion and altered endocytosis caused by expression of Alzheimer's disease‐protective variant PLCγ2 R522

Variants identified in genome‐wide association studies have implicated immune pathways in the development of Alzheimer’s disease (AD). Here, we investigated the mechanistic basis for protection from AD associated with PLCγ2 R522, a rare coding variant of the PLCG2 gene. We studied the variant''s role in macrophages and microglia of newly generated PLCG2‐R522‐expressing human induced pluripotent cell lines (hiPSC) and knockin mice, which exhibit normal endogenous PLCG2 expression. In all models, cells expressing the R522 mutation show a consistent non‐redundant hyperfunctionality in the context of normal expression of other PLC isoforms. This manifests as enhanced release of cellular calcium ion stores in response to physiologically relevant stimuli like Fc‐receptor ligation or exposure to Aβ oligomers. Expression of the PLCγ2‐R522 variant resulted in increased stimulus‐dependent PIP₂ depletion and reduced basal PIP₂ levels in vivo. Furthermore, it was associated with impaired phagocytosis and enhanced endocytosis. PLCγ2 acts downstream of other AD‐related factors, such as TREM2 and CSF1R, and alterations in its activity directly impact cell function. The inherent druggability of enzymes such as PLCγ2 raises the prospect of PLCγ2 manipulation as a future therapeutic approach in AD.     

Single‐cell transcriptional profiling of human carotid plaques reveals a subpopulation of endothelial cells associated with stroke incidences

The differences in plaque histology between symptomatic and asymptomatic patients have been widely accepted. Whether there is a heterogeneity of cells between symptomatic and asymptomatic plaques remains largely unclear. To reveal the potential heterogeneity within different plaques, which may contribute to different stroke incidences, we obtained the scRNA‐seq data from symptomatic and asymptomatic patients and identified eight cell types present in plaques. Further analysis of endothelial cells (ECs) revealed three distinct EC subpopulations appeared to be endowed with specific biological functions such as antigen processing and presentation, cell adhesion, and smooth muscle cell proliferation. Of note, the differentially expressed genes of the EC 2 subpopulation showed that the genes involved in cell adhesion were up‐regulated in asymptomatic plaques compared to symptomatic plaques. Integrating the data of intraplaque haemorrhage and plaque stability, the 5th top‐enriched biological process was cell adhesion in the stable or non‐haemorrhaged plaques compared to unstable plaques or haemorrhaged plaques. Among these cell adhesion‐related genes, the intersection gene AOC3 may play a vital role in plaque haemorrhage and plaque stability. Targeting cell adhesion and the specialized genes may provide potential new therapeutic directions to prevent asymptomatic patients from stroke.     

SETD2 epidermal deficiency promotes cutaneous wound healing via activation of AKT/mTOR Signalling

ObjectivesCutaneous wound healing is one of the major medical problems worldwide. Epigenetic modifiers have been identified as important players in skin development, homeostasis and wound repair. SET domain–containing 2 (SETD2) is the only known histone H3K36 tri‐methylase; however, its role in skin wound healing remains unclear.Materials and MethodsTo elucidate the biological role of SETD2 in wound healing, conditional gene targeting was used to generate epidermis‐specific Setd2‐deficient mice. Wound‐healing experiments were performed on the backs of mice, and injured skin tissues were collected and analysed by haematoxylin and eosin (H&E) and immunohistochemical staining. In vitro, CCK8 and scratch wound‐healing assays were performed on Setd2‐knockdown and Setd2‐overexpression human immortalized keratinocyte cell line (HaCaT). In addition, RNA‐seq and H3K36me3 ChIP‐seq analyses were performed to identify the dysregulated genes modulated by SETD2. Finally, the results were validated in functional rescue experiments using AKT and mTOR inhibitors (MK2206 and rapamycin).ResultsEpidermis‐specific Setd2‐deficient mice were successfully established, and SETD2 deficiency resulted in accelerated re‐epithelialization during cutaneous wound healing by promoting keratinocyte proliferation and migration. Furthermore, the loss of SETD2 enhanced the scratch closure and proliferation of keratinocytes in vitro. Mechanistically, the deletion of Setd2 resulted in the activation of AKT/mTOR signalling pathway, while the pharmacological inhibition of AKT and mTOR with MK2206 and rapamycin, respectively, delayed wound closure.ConclusionsOur results showed that SETD2 loss promoted cutaneous wound healing via the activation of AKT/mTOR signalling.     

CircASPH promotes KGN cells proliferation through miR‐375/MAP2K6 axis in Polycystic Ovary Syndrome

Polycystic Ovary Syndrome (PCOS) is a kind of endocrine disorder which is prevalent in adult women, so exploring more biomarkers for PCOS is imperative. Recently, circular RNA and microRNA are confirmed to be related with PCOS development. Whether circular RNA ASPH (circASPH) is involved in PCOS need to be studied further. We utilized RT‐qPCR to measure the expression levels of circASPH, miR‐375 and MAP2K6 in PCOS patients and normal group. The effects of circASPH and miR‐375 on KGN cells proliferation and apoptosis were observed by CCK‐8 assay, EdU incorporation assay and apoptosis assay, separately. Then Dual‐luciferase reporter assay was carried out to verify the circASPH/miR375 axis and miR375/MAP2K6 axis. The interaction between circASPH and MAP2K6 were detected with the support of RT‐qPCR and Western blot. We found circASPH and MAP2K6 were both over‐expressed in PCOS patients, while miR‐375 was in the opposite direction. Moreover, miR‐375 was negatively regulated by circASPH, while MAP2K6 was positively regulated by circASPH. In addition, circASPH directly targeted miR‐375, which targeted MAP2K6. More than that, the knockdown of circASPH repressed KGN cells proliferation and enhanced apoptosis, while the silence of miR‐375 reversed the above effects. In conclusion, circASPH promotes KGN cells proliferation through miR‐375/MAP2K6 axis in PCOS, and they are thought‐provoking biomarkers for PCOS diagnosis and therapy.     

Vascular endothelial growth factor‐C in activating vascular endothelial growth factor receptor‐3 and chemokine receptor‐4 in melanoma adhesion

Vascular endothelial growth factor‐C (VEGF‐C) binds to receptor vascular endothelial growth factor receptor‐3 (VEGFR‐3) expressed on lymphatic endothelial and melanoma cells. Binding of VEGF‐C to VEGFR‐3 enhances receptor phosphorylation that activates mitogen‐activated protein kinase (MAP‐K) and phosphatidylinositol‐3‐kinase (PI3K). These signalling pathways regulate cell migration and adhesion in response to internal or external changes.In addition, the overexpression of VEGF‐C upregulates chemokine receptor CXCR‐4 in tumours (melanoma). CXCR‐4 is expressed on cells of the immune system (natural killer cells) and facilitates the migration of leukocytes in response to the CXCL12 ligand. The latter is expressed by lymphatic endothelial cells and by stromal cells in the tumour microenvironment (TME). The gradient established between CXCR‐4 expressed on tumour cells and CXCL12 produced by stromal and lymphatic endothelial cells enhances tumour cell metastasis.3‐(4‐Dimethylamino‐naphthalen‐1‐ylmethylene)‐1, 3‐dihydroindol‐2‐one, MAZ‐51, is an indolinone‐based synthetic molecule that inhibits the phosphorylation of the tyrosine kinase receptor VEGFR‐3. CTCE‐9908, a CXCR‐4 antagonist derived from human CXCL12, hinders receptor phosphorylation and the subsequent signalling pathways that would be activated.VEGF‐C is stimulated by transforming growth factor‐beta 1 (TGF‐β1), which facilitates cell–cell and cell‐matrix adhesion by regulating cadherins through the activation of focal adhesion kinase (FAK) and mediates paxillin upregulation.Increased VEGF‐C protein levels stimulated by TGF‐β bound to VEGFR‐3 impact on intracellular pathways that promote tumour cell adhesion. In addition, increased VEGF‐C protein levels lead to enhanced CXCR‐4 protein expression. Therefore, effective blocking of VEGR‐3 and CXCR‐4 may inhibit tumour cell metastasis by hampering intracellular proteins promoting adhesion.     

Cathepsin K deficiency promotes alveolar bone regeneration by promoting jaw bone marrow mesenchymal stem cells proliferation and differentiation via glycolysis pathway

ObjectivesTo clarify the possible role and mechanism of Cathepsin K (CTSK) in alveolar bone regeneration mediated by jaw bone marrow mesenchymal stem cells (JBMMSC).Materials and MethodsTooth extraction models of Ctsk knockout mice (Ctsk ^‐/‐) and their wildtype (WT) littermates were used to investigate the effect of CTSK on alveolar bone regeneration. The influences of deletion or inhibition of CTSK by odanacatib (ODN) on proliferation and osteogenic differentiation of JBMMSC were assessed by CCK‐8, Western blot and alizarin red staining. To explore the differently expressed genes, RNA from WT and Ctsk^‐/‐ JBMMSC was sent to RNA‐seq. ECAR, glucose consumption and lactate production were measured to identify the effect of Ctsk deficiency or inhibition on glycolysis. At last, we explored whether Ctsk deficiency or inhibition promoted JBMMSC proliferation and osteogenic differentiation through glycolysis.ResultsWe found out that Ctsk knockout could promote alveolar bone regeneration in vivo. In vitro, we confirmed that both Ctsk knockout and inhibition by ODN could promote proliferation of JBMMSC, up‐regulate expression of Runx2 and ALP, and enhance matrix mineralization. RNA‐seq results showed that coding genes of key enzymes in glycolysis were significantly up‐regulated in Ctsk^‐/‐ JBMMSC, and Ctsk deficiency or inhibition could promote glycolysis in JBMMSC. After blocking glycolysis by 3PO, the effect of Ctsk deficiency or inhibition on JBMMSC’s regeneration was blocked subsequently.ConclusionsOur findings revealed that Ctsk knockout or inhibition could promote alveolar bone regeneration by enhancing JBMMSC regeneration via glycolysis. These results shed new lights on the regulatory mechanism of CTSK on bone regeneration.