L-citrulline is a nonprotein amino acid that plays an important role in human health and has great market demand. Although microbial cell factories have been widely used for biosynthesis, there are still challenges such as genetic instability and low efficiency in the biosynthesis of L-citrulline. In this study, an efficient, plasmid-free, non-inducible L-citrulline-producing strain of Escherichia coli BL21(DE3) was engineered by combined strategies. Firstly, a chassis strain capable of synthesizing L-citrulline was constructed by block of L-citrulline degradation and removal of feedback inhibition, with the L-citrulline titer of 0.43 g/L. Secondly, a push-pull-restrain strategy was employed to enhance the L-citrulline biosynthesis, which realized the L-citrulline titer of 6.0 g/L. Thirdly, the NADPH synthesis and L-citrulline transport were strengthened to promote the synthesis efficiency, which achieved the L-citrulline titer of 11.6 g/L. Finally, fed-batch fermentation was performed with the engineered strain in a 3 L fermenter, in which the L-citrulline titer reached 44.9 g/L. This study lays the foundation for the industrial production of L-citrulline and provides insights for the modification of other amino acid metabolic networks.
Citrulline/biosynthesis* ; Escherichia coli/genetics* ; Metabolic Engineering/methods* ; Fermentation ; NADP/biosynthesis*

ObjectiveTo explore the possible action mechanism of Qingshi Anti-itch Ointment （青石止痒软膏， QAO） in the treatment of psoriasis. MethodsForty mice were randomly divided into four groups， blank group， model group， calcipotriol group and QAO group， with 10 mice in each group. Except for the blank group， psoriasis was induced by applying imiquimod cream to the dorsal skin. After modeling for 6 hours daily， the calcipotriol group and QAO group were treated with 0.5 g of calcipotriol ointment or 0.5 g of QAO， respectively， applied to the treated dorsal skin. The blank group and the model group received no treatment. The skin lesions were observed， and the psoriasis area and severity index （PASI） score was assessed every other day. After 7 days， Hematoxylin and Eosin （HE） staining was performed on dorsal skin tissue to observe pathological changes. The levels of interleukin 1β （IL-1β） and interleukin 18 （IL-18） were determined by enzym-linked immunosorbent assay （ELISA）. The protein levels of Caspase-1，Pro-Caspase-1， gasdermin D （GSDMD） and gasdermin-D-N （GSDMD-N） were detected by Western Blot （WB）. The protein levels of GSDMD were observed by immunohistochemistry. ResultsCompared with the blank group， the model group mice showed redness， erythema， and white scales on their skin， with histological observations indicating epidermal thickening， elongated spines， and infiltration of inflammatory cells. The PASI scores of the skin tissue on days 1， 3， 5， and 7 were elevated； the IOD and AOD values of GSDMD protein increased； the protein levels of Caspase-1， Pro-Caspase-1，GSDMD， GSDMD-N， and IL-1β and IL-18 were significantly elevated （P＜0.05 or P＜0.01）. Compared with the model group， the QAO group and calcipotriol group showed lighter skin lesions； the PASI scores on day 5 and day 7 in the QAO group， and on day 3， 5， and 7 in the calcipotriol group， were reduced； the IOD and AOD values of GSDMD protein， and the protein level of Caspase-1， GSDMD， and GSDMD-N， as well as level of IL-18 and IL-1β decreased in both groups； in the calcipotriol group， Pro-Caspase-1 protein level also decreased （P＜0.05 or P＜0.01）. Compared with the calcipotriol group， the QAO group showed slightly redder skin， more obvious thickening of the stratum corneum， and less capillary dilation； the PASI scores on day 3 and day 7 increased， while the score on day 5 was reduced； the protein level of Pro-Caspase-1， GSDMD， GSDMD-N， and the level of IL-18 and IL-1β were increased in the QAO group （P＜0.05）. ConclusionQAO can effectively relieve psoriasis dermatitis in mice. Its potential mechanism may be related to the regulation of the Caspase-1/GSDMD protein pathway， down-regulation of IL-18 and IL-1β levels， and alleviation of pyroptosis.

ObjectiveTo evaluate the clinical efficacy and safety of Huaban Jiedu Formulation （化斑解毒方， HJF） in treating psoriasis vulgaris with blood-heat syndrome. MethodsA randomized， double-blind， placebo-controlled study was conducted with 60 patients diagnosed with psoriasis vulgaris of blood-heat syndrome. Patients were randomly assigned to either a treatment group or a control group， with 30 cases in each. The treatment group received HJF granules orally， one dose a day， combined with topical Qingshi Zhiyang Ointment （青石止痒软膏）， while the control group received placebo granules， one dose a day， combined with the same topical ointment. Both groups were topically treated twice daily of 28 days treatment cours. Psoriasis area and severity index （PASI）， visual analogue scale for pruritus （VAS）， traditional Chinese medicine （TCM） syndrome scores， dermatology life quality index （DLQI）， and psoriasis life stress inventory （PLSI） were assessed before treatment and on day 14 and day 28. Response rates for PASI 50 （≥50% reduction） and PASI 75 （≥75% reduction）， as well as overall clinical efficacy， were compared between groups. Serum levels of interleukin-6 （IL-6） and interleukin-17 （IL-17） were measured before and after 28 days of treatment. Adverse reactions during treatment were recorded. ResultsAfter 28 days of treatment， both groups showed significant reductions in PASI total score， lesion area score， erythema， scaling， and infiltration scores， pruritus VAS score， TCM syndrome score， DLQI， PLSI， and serum IL-6 and IL-17 levels （P＜0.05）. Compared to the control group， the treatment group had significantly greater improvements in PASI total score and erythema score， TCM syndrome score， serum IL-6 and IL-17 levels， and PASI 50 response rate after 28 days （P＜0.05）. Between-group comparisons of score differences before and after 28-day treatment revealed that the treatment group showed significantly better improvements in PASI total， lesion area score， erythema score， TCM syndrome score， DLQI， PLSI， and inflammatory markers （P＜0.05 or P＜0.01）. The total effective rate on day 14 and day 28 was 40.00% （12/30） and 83.33% （25/30） in the treatment group， versus 6.90% （2/29） and 41.38% （12/29） in the control group， respectively. The clinical efficacy in the treatment group was significantly superior to that in the control group （P＜0.05）. Mild gastric discomfort occurred in 3 patients in the treatment group and 1 in the control group. ConclusionHJF can effectively improve skin lesions and TCM symptoms relieve pruritus， enhance quality of life， and reduce inflammatory markers IL-6 and IL-17， in patients with blood-heat syndrome of psoriasis vulgaris， with a good safety profile.

OBJECTIVE To investigate the effects of borneol on pharmacodynamic and pharmacokinetic effects of Corydalis saxicola total alkaloids in depression model rats.METHODS Thirty male SD rats were divided into blank control group,negative control group,positive control group (fluoxetine 10 mg/kg,i.g.),single drug group (C.saxicola total alkaloids 210 mg/kg,i.g.) and combined drug group (C.saxicola total alkaloids 210 mg/kg+borneol 50 mg/kg,i.g.) according to the random number table method,with 6 rats in each group.By lipopolysaccharide (LPS) induction modeling,except blank control group (no model and no administration) received intraperitoneal injection of the same amount of normal saline,the rats in the other groups were intraperitoneally injected with LPS once a day to establish a rat model of depression.After 1 week of modeling,each administration group was given relevant drug intragastrically according to the corresponding dose,and blank control group and negative control group (without drug treatment) were administered intragastrically with an equal volume of solvent to dissolve the drug;continued modeling while administering the drug.After two weeks of continuous administration,the effects of C.saxicola total alkaloids versus the combination of C.saxicola total alkaloids and borneol on the behavior of depressed rats were tested by behavioral experiments;the levels of tumor necrosis factor-α,interleukin-1β and interleukin-6 in rats were determined;the histopathological changes of the hippocampus of rats were observed.Blood sample was collected from the orbit at different time points after administration on the 15th day,and the upper plasma was obtained.Ultra-performance liquid chromatography-triple quadrupole tandem mass spectrometry was established for the simultaneous determination of dehydrocarvedine,tetrahydropalmatine,coptisine,palmatine,jatrorrhizine,berberine,berberrubine and epiberberine in rat plasma.The average plasma concentration-time curve was depicted,the area under the curve (AUC) was calculated,and the pharmacokinetic parameters were analyzed by DAS 3.2.2 software.RESULTS Compared with blank control group,the negative control group had a decrease in body mass and sugar water preference rate,a decrease in the total distance of open field,a prolonged swimming immobility time,and a increased in the expression of inflammatory factors in serum (P<0.05);compared with negative control group,the single drug group and the combined drug group increased the preference rate of sugar water,increased the total distance of open field,shortened the time of swimming immobility,and decreased the expression of inflammatory factors in serum (P<0.05).There was no significant difference in the above indicators between the single drug group and the combined drug group in rats (P>0.05).Pharmacokinetic results showed that compared with single drug group,AUC0-t of coptisine,AUC0-t,AUC0-∞,tmax and cmax of jatrorrhizine,AUC0-t,AUC0-∞,t1/2 and cmax of berberrubine,and AUC0-t of epiberberine,cmax of dehydrocarvedine,cmax of palmatine were significantly increased in combined drug group,but there was no significant difference,indicating that borneol didn't have a significant effect on the efficacy of Corydalis saxicola nigra at this dose.CONCLUSIONS Both C.saxicola total alkaloids alone and in combination with borneol can improve depression-like behavior in depression model rats,reduce serum inflammatory cytokine levels,and protect hippocampal neurons.Compared with the use of Corydalis saxicola base alone,the combination with borneol do not show significant pharmacodynamic differences,bu can improve the absorption of coptisine,jatrorrhizine in model rats.

Objective:to investigate the expression of integrin αvβ3 and osteopontin(OPN)in decidual tissues in missed abortion women with polycystic ovarian syndrome(PCOS).Methods:PCOS women with missed abor-tion(PCOS missed abortion group),non-PCOS with missed abortion(non-PCOS missed abortion group)and women with normal early pregnancy induced abortion(normal control group)who were admitted to the First Affili-ated Hospital of Xi'an Jiaotong University from February 2018 to December 2022 were selected as participants,and 20 subjects in each group.The expressions of integrin αvβ3 and OPN in decidual tissues among the three groups were detected.Correlation analysis was conducted between the expression levels of integrin αvβ3 and OPN in PCOS patients with missed abortion.Results:Integrin αvβ3 and OPN were mainly expressed in the epi-thelial cells of the uterus,as well as in decidual cells.There were significant differences in the positive rates and IHC scores of integrin αvβ3 and OPN in decidual tissues among the three groups(P<0.05).The positive rates and IHC scores in PCOS missed abortion group were significantly lower compared with non-PCOS missed abor-tion group and normal control group,and the difference was statistically significant(P<0.05),but there was no significant difference between non-PCOS missed abortion group and normal control group(P>0.05).The integrin αvβ3 and OPN in decidual tissues were mainly co-located in uterine epithelial cells.There were significant differ-ences in immunofluorescence intensity of integrin αvβ3 and OPN among the three groups(P<0.05).The missed abortion group of PCOS was significantly lower than that of non-PCOS missed abortion group and normal control group,and the difference was statistically significant(P<0.01).No significant difference was found between non-PCOS missed abortion group and normal control group(P>0.05).There were significant differences in the protein expression levels and mRNA expression levels of integrin αvβ3 and OPN in decidual tissue among the three groups(P<0.05).PCOS missed abortion group was significantly lower than non-PCOS missed abortion group and normal control group,and the difference was statistically significant(P<0.01).There was no significant difference between non-PCOS missed abortion group and normal control group(P>0.05).The results of Pearson correlation analysis showed that the protein and mRNA expression levels of integrin αvβ3 and OPN in decidual tis-sue of PCOS patients with missed abortion were significantly positively correlated(r>0,P<0.01).Conclusions:The integrin αvβ3 and OPN were expressed in both epithelial cells of the uterus and decidual cells in early preg-nancy.The expressions of them were significantly decreased in decidual tissues of PCOS patients with missed a-bortion,which might be related to reduced endometrial receptivity.The role of integrin αvβ3 and OPN in the occur-rence of missed abortion needs further study.

Objective:to investigate the expression of integrin αvβ3 and osteopontin(OPN)in decidual tissues in missed abortion women with polycystic ovarian syndrome(PCOS).Methods:PCOS women with missed abor-tion(PCOS missed abortion group),non-PCOS with missed abortion(non-PCOS missed abortion group)and women with normal early pregnancy induced abortion(normal control group)who were admitted to the First Affili-ated Hospital of Xi'an Jiaotong University from February 2018 to December 2022 were selected as participants,and 20 subjects in each group.The expressions of integrin αvβ3 and OPN in decidual tissues among the three groups were detected.Correlation analysis was conducted between the expression levels of integrin αvβ3 and OPN in PCOS patients with missed abortion.Results:Integrin αvβ3 and OPN were mainly expressed in the epi-thelial cells of the uterus,as well as in decidual cells.There were significant differences in the positive rates and IHC scores of integrin αvβ3 and OPN in decidual tissues among the three groups(P<0.05).The positive rates and IHC scores in PCOS missed abortion group were significantly lower compared with non-PCOS missed abor-tion group and normal control group,and the difference was statistically significant(P<0.05),but there was no significant difference between non-PCOS missed abortion group and normal control group(P>0.05).The integrin αvβ3 and OPN in decidual tissues were mainly co-located in uterine epithelial cells.There were significant differ-ences in immunofluorescence intensity of integrin αvβ3 and OPN among the three groups(P<0.05).The missed abortion group of PCOS was significantly lower than that of non-PCOS missed abortion group and normal control group,and the difference was statistically significant(P<0.01).No significant difference was found between non-PCOS missed abortion group and normal control group(P>0.05).There were significant differences in the protein expression levels and mRNA expression levels of integrin αvβ3 and OPN in decidual tissue among the three groups(P<0.05).PCOS missed abortion group was significantly lower than non-PCOS missed abortion group and normal control group,and the difference was statistically significant(P<0.01).There was no significant difference between non-PCOS missed abortion group and normal control group(P>0.05).The results of Pearson correlation analysis showed that the protein and mRNA expression levels of integrin αvβ3 and OPN in decidual tis-sue of PCOS patients with missed abortion were significantly positively correlated(r>0,P<0.01).Conclusions:The integrin αvβ3 and OPN were expressed in both epithelial cells of the uterus and decidual cells in early preg-nancy.The expressions of them were significantly decreased in decidual tissues of PCOS patients with missed a-bortion,which might be related to reduced endometrial receptivity.The role of integrin αvβ3 and OPN in the occur-rence of missed abortion needs further study.

ObjectiveTo investigate the effect of total alkaloids of Corydalis saxicola on a rat model of lipopolysaccharide（LPS）-induced depression， as well as the pharmacokinetic characteristics of 8 of its major components. MethodTwenty-four male SD rats were randomly divided into normal group， model group， fluoxetine group（10 mg·kg^-1） and total alkaloids of C. saxicola group（210 mg·kg^-1）， with 6 rats in each group. In addition to the normal group， the rats were injected intraperitoneally with LPS to establish the inflammation model of depression， and the drug administration was started 1 week after modeling， and the administration groups were gavaged according to the corresponding dose， and the normal and model groups were intragastric administration with equal volume of distilled water， and the administration was performed along with the modeling. After two weeks of continuous administration， the effect of total alkaloids of C. saxicola on the behavior of depressed rats were tested by sucrose preference， forced swimming and open field experiments， the levels of tumor necrosis factor-α（TNF-α）， interleukin（IL）-1β and IL-6 in serum of rats were determined by enzyme-linked immunosorbent assay（ELISA）， the histopathological changes of rat hippocampus were observed by hematoxylin-eosin（HE） staining. After the last administration， blood was collected from orbit according to the set time， and ultra-high performance liquid chromatography-triple quadrupole tandem mass spectrometry（UPLC-QqQ-MS） was established to simultaneously detect the concentrations of dehydrocavidine， tetrahydropalmatine， coptisine， palmatine， jatrorrhizine， berberine， berberrubine and epiberberine in plasma， and drug-time curves were drawn. The pharmacokinetic parameters were analyzed by DAS 2.0 software. ResultCompared with the normal group， the model group exhibited a decrease in sucrose preference rate， total distance traveled in the open field， as well as an increase in swimming immobility time and serum inflammatory factor expression（P<0.01）. In contrast， compared with the model group， rats in each administration group showed an increase in sucrose preference rate and total distance traveled in the open field， a decrease in swimming immobility time， and a reduction in serum inflammatory factor expression（P<0.05， P<0.01）. Additionally， HE staining results revealed that neurons in the hippocampus of rats from the model group were characterized by loss， disorganization and residual vacuoles， whereas those from the total alkaloids of C.saxicola group displayed an increase in number with orderly arrangement and clear cytoplasm. Pharmacokinetic results showed that the time to peak（t_max） and half-life（t_1/2） of the 8 active ingredients were 0.19-2.06 h and 3.71-8.70 h after continuous administration of total alkaloids of C. saxicola. Among them， the area under the curve（AUC_0-∞） of tetrahydropalmatine was the highest and the t_1/2 was the shortest， and the AUC_0-∞ of coptisine， palmatine， jatrorrhizine， berberine， berberrubine and epiberberine were low. The curves of dehydrocavidine， coptisine， palmatine， berberine and epiberberine showed obvious double peak phenomenon. ConclusionTotal alkaloids of C. saxicola can improve the depression-like behavior of rats， inhibit the expression of inflammatory factors in serum， improve the pathological injury of hippocampus， and has the antidepressant effect. Meanwhile， the effective site is absorbed quickly and eliminated slowly in the depressed model rats， and the efficacy is maintained for a long time.

Objective:To study clinical efficacy of isocitrate dehydrogenase 1(IDH1)mutation on temozolomide chemotherapy for glioma patients and its impact on immune cytokines and prognosis.Methods:A total of 134 patients with glioma who underwent surgical resection and were confirmed by pathology in Dushu Lake Hospital Affiliated to Soochow University from October 2021 to October 2022 were selected as research subjects.IDH1 mutation status was measured by direct sequencing method,and IDH1 expres-sion rate in glioma tissue was determined by immunohistochemistry.All patients with brain glioma were treated with temozolomide chemotherapy.Effects of IDH1 mutation on clinical efficacy,immune cytokines(IFN-γ,IL-2,IL-4,IL-10)and prognosis of glioma were analyzed.Results:Seventy-nine cases out of 134 cases of glioma tissue had IDH1 mutations,mostly at R132,with a mutation rate of 58.96%,significantly higher than normal brain tissue(χ2=48.066,P<0.05).Immunohistochemistry showed strong positive expression of IDH1 in 56 out of 134 glioma tissues.Proportion of WHO grade Ⅳ in IDH1 mutant group was lower than IDHI wild type group[11.39%(9/79)vs 63.64%(35/55),Z=41.020,P<0.05].Proportion of low differentiation in IDH1 mutant group was higher than IDHI wild type group[50.63%(40/79)vs 20.00%(11/55),χ2=12.907,P<0.05].Total effective rate in IDH1 mutant group was higher than IDH1 wild type group[91.14%(72/79)vs 76.36%(42/55),χ2=5.575,P<0.05],IFN-γ and IL-2 levels were higher than IDH1 wild type group[(28.98±3.25)pg/ml vs(20.15±2.54)pg/ml,(33.42±4.25)pg/ml vs(25.23±3.52)pg/ml,t=16.870,11.750,P<0.05],IL-4 and IL-10 levels were lower than IDH1 wild type group[(7.90±1.02)pg/ml vs(12.38±1.66)pg/ml,(8.79±1.00)pg/ml vs(15.26±1.23)pg/ml,t=19.330,33.500,P<0.05].IDH1 mutation was positively correlated with IFN-γ and IL-2 levels after temozolo-mide chemotherapy(r=0.845,0.772,P<0.05),and negatively correlated with IL-4 and IL-10 levels after temozolomide chemotherapy(r=-0.786,-0.685,P<0.05).Survival rate after chemotherapy in IDH1 mutant group was higher than IDH1 wild type group[89.87%(71/79)vs 72.70%(40/55),Log Rank test χ2=5.208,P<0.05].Cox regression analysis found that WHO grade Ⅲ(RR=1.342),poorly differentiated(RR=1.783),IFN-γ(RR=1.808),IL-2(RR=2.112),IL-4(RR=2.342),IL-10(RR=1.342)as risk factors,temozolo-mide chemotherapy efficacy(RR=0.653),IDH1 mutation(RR=0.895)as protective factors affect prognosis of temozolomide chemo-therapy in glioma patients(P<0.05).Conclusion:IDH1 mutation is related to disease grade and differentiation degree of glioma patients,and can affect efficacy of temozolomide chemotherapy and expressions of immune cytokines,which is a protective factor for prognosis and survival after temozolomide chemotherapy.

Objective:Trigeminal neuralgia(TN)is a common neuropathic pain.Voltage-gated potassium channel(Kv)has been confirmed to be involved in the occurrence and development of TN,but the specific mechanism is still unclear.MicroRNA may be involved in neuropathic pain by regulating the expression of Kv channels and neuronal excitability in trigeminal ganglion(TG).This study aims to explore the relationship between Kv1.1 and miR-21-5p in TG with a TN model,evaluate whether miR-21-5p has a regulatory effect on Kv1.1,and to provide a new target and experimental basis for the treatment of TN. Methods:A total of 48 SD rats were randomly divided into 6 groups:1)a sham group(n= 12),the rats were only sutured at the surgical incision without nerve ligation;2)a sham+ agomir NC group(n=6),the sham rats were microinjected with agomir NC through stereotactic brain injection in the surgical side of TG;3)a sham+miR-21-5p agomir group(n=6),the sham rats were microinjected with miR-21-5p agomir via stereotactic brain injection in the surgical side of TG;4)a TN group(n=12),a TN rat model was constructed using the chronic constriction injury of the distal infraorbital nerve(dIoN-CCI)method with chromium intestinal thread;5)a TN+antagonist NC group(n=6),TN rats were microinjected with antagonist NC through stereotactic brain injection method in the surgical side of TG;6)a TN+miR-21-5p antagonist group(n=6),TN rats were microinjected with miR-21-5p antagonist through stereotactic brain injection in the surgical side of TG.The change of mechanical pain threshold in rats of each group after surgery was detected.The expressions of Kv1.1 and miR-21-5p in the operative TG of rats were detected by Western blotting and real-time reverse transcription polymerase chain reaction.Dual luciferase reporter genes were used to determine whether there was a target relationship between Kv1.1 and miR-21-5p and whether miR-21-5p directly affected the 3'-UTR terminal of KCNA1.The effect of brain stereotaxic injection was evaluated by immunofluorescence assay,and then the analogue of miR-21-5p(agomir)and agomir NC were injected into the TG of rats in the sham group by brain stereotaxic apparatus to overexpress miR-21-5p.The miR-21-5p inhibitor(antagomir)and antagomir NC were injected into TG of rats in the TN group to inhibit the expression of miR-21-5p.The behavioral changes of rats before and after administration were observed,and the expression changes of miR-21-5p and Kv1.1 in TG of rats after intervention were detected. Results:Compared with the baseline pain threshold,the facial mechanical pain threshold of rats in the TN group was significantly decreased from the 5th to 15th day after the surgery(P<0.05),and the facial mechanical pain threshold of rats in the sham group was stable at the normal level,which proved that the dIoN-CCI model was successfully constructed.Compared with the sham group,the expression of Kv1.1 mRNA and protein in TG of the TN group was down-regulated(both P<0.05),and the expression of miR-21-5p was up-regulated(P<0.05).The results of dual luciferase report showed that the luciferase activity of rno-miR-21-5p mimics and KCNA1 WT transfected with 6 nmol/L or 20 nmol/L were significantly decreased compared with those transfected with mimic NC and wild-type KCNA1 WT,respectively(P<0.001).Compared with low dose rno-miR-21-5p mimics(6 nmol/L)co-transfection group,the relative activity of luciferase in the high dose rno-miR-21-5p mimics(20 nmol/L)cotransfection group was significantly decreased(P<0.001).The results of immunofluorescence showed that drugs were accurately injected into TG through stereotaxic brain.After the expression of miR-21-5p in the TN group,the mechanical pain threshold and the expression of Kv1.1 mRNA and protein in TG were increased.After overexpression of miR-21-5p in the sham group,the mechanical pain threshold and the expression of Kv1.1 mRNA and protein in TG were decreased. Conclusion:Both Kv1.1 and miR-21-5p are involved in TN and miR-21-5p can regulate Kv1.1 expression by binding to the 3'-UTR of KCNA1.

Objective To investigate the role of Tre2-Bub2-Cdc16 1 domain family member5(TBC1D5)in the de-velopment of hepatocellular carcinoma(HCC).Methods Western blot(WB),Immunohistochemistry(IHC)and quantitative real-time PCR(qPCR)were used to verify the difference in TBC1D5 expression in clinical sam-ples.The HCC cell lines MHCC97H and Hep3B were chosen to construct the knockdown model.The effects on cell proliferation were detected by cell proliferation assay,colony formation assay and EdU assay.Wound assay and Transwell assay were used to detect cell migration and invasion.Flow cytometry was used to detect the changes of cell cycle and H2O2-induced apoptosis of HCC cells.Finally,the effects of knockdown and overexpression of TBC1D5 on JAK/STAT pathway were detected by WB.Results The results of WB,IHC and qPCR showed that the expression of TBC1D5 in HCC tissues was higher at the protein level(P＜0.000 1)and mRNA level(P＜0.01)than that in corresponding adjacent tissues.Compared with the control group,the proliferation level of HCC cells with TBC1D5 knockdown was decreased(P＜0.05),the formation of plate colony number decreased(P＜0.001),and the proportion of EdU-positive cells decreased(P＜0.001).The results of scratch assay and Tran-swell assay showed that the migration(P＜0.01)and invasion ability(P＜0.01)of HCC cells after TBC1D5 knockdown were significantly lower than those in the control group.After TBC1D5 knockdown,the cell cycle of HCC cells was slowed down(P＜0.05)and the ability to resist apoptosis was reduced(P＜0.01)than those in the control group.Compare with the control group,knockdown of TBC1D5 decreased the phosphorylation level of JAK and STAT proteins and inhibit the JAK/STAT pathway.Conclusion TBC1D5 is highly expressed in HCC.After knocking down TBC1D5,the proliferation,migration and invasion ability,cell cycle rate and anti-apoptosis ability of HCC cells decreased and may affect HCC progression through the JAK/STAT pathway.