Objective:To investigate whether IL-7-secreting oncolytic herpes simplex virus(HSV)could activate CD8+T cells and inhibit the growth of hepatocellular carcinoma.Methods:The expression of IL-7 was detected by Western blot.The in vitro cleavage of tumor cells by tumor oncolytic virus HSV and HSV-IL-7 were detected by crystal violet staining.The tumor inhibition ability of HSV-IL-7 and HSV were detected in subcutaneous transplanted tumor model.Levels of IL-7,IFN-γ and TNF-α in serum and tumor tissues were determined by ELISA.The infiltration of CD8+T cells in tumor tissues was detected by immunohistochemistry.Flow cytometry was used to detect Granzyme B secretion in CD8+T cells infiltrated by tumor.Results:Tumor cells infected with HSV-IL-7 expressed high level of IL-7.Both HSV and HSV-IL-7 can effectively lyse B16-F10,CT-26 and H22 tumor cell lines in a dose-dependent manner in vitro.HSV-IL-7 could significantly inhibit the growth of H22 hepatoma cells in vivo(P<0.01)and prolong the survival time of tumor-bearing mice(P<0.001).HSV-IL-7 could significantly increase the IL-7 content in tumor sites(P<0.000 1),and effectively increase the number of tumor infiltrating CD8+T cells(P<0.001).HSV-IL-7 significantly enhanced Granzyme B secretion of tumor-infiltrating CD8+T cells and IFN-γ and TNF-α in tumor tissues(P<0.000 1).Conclusion:HSV-IL-7 has well tumor inhibition activity in vivo and in vitro.It also can activate the anti-tumor activity of CD8+T cells in vivo by secreting IL-7,inhibit tumor growth and prolong the survival time of tumor-bearing mice.

Asymptomatic spleen-stomach diseases refer to diseases without related symptoms and signs of abdo-minal pain， bloating， diarrhea an others in patients， but showing lesions or pathological changes discovered by modern medical techniques such as endoscopy， CT， MRI. The four examination techniques of traditional Chinese medicine （TCM） are based on symptoms and signs of patients， which are the advantage of TCM but also have certain limitations. In the context of the increasingly modernized diagnosis and treatment in TCM， it is proposed to expand the application of the four examination techniques from three aspects including microcosmic syndrome differentiation， data sharing， and artificial intelligence in asymptomatic spleen-stomach diseases， in order to achieve the goals of dynamically observing the disease process， collecting disease data in multiple dimensions， and intelligently processing disease data. This will strengthen the modern requirements of early diagnosis and treatment in TCM， and highlight the advantages of TCM in “treating disease before it arises and treating the symptoms beforehand”.

Immune escape is one of the ten hallmarks of tumors， which plays an important role in the occurrence and development of tumors. Immune escape refers to a process where tumor cells remodel and edit the immune system through the model of immune clearance， immune balance， and immune escape to "transform" the immune cells into immunosuppressive cells in the tumor microenvironment， so as to support immune escape. The five-stage evolution is the summary of tumor pathogenesis by professor LI Jie. He believes that the gradual development of tumors follows the core pathogenesis of "deficiency-cold-toxin-obstruction-collapse"， in which "depression" runs through the whole process， and cancer toxin is the key. Based on immune editing， this paper combined phenotypic characteristics of tumor cells with the core pathogenesis of the five-stage evolution of professor LI to reveal the biological basis of malignant tumor five-stage evolution. The results indicate that the prominent change from deficiency to cold is the reduction of immune surveillance and the prominent change from toxin to obstruction is immune escape. The final stage of collapse is the outcome of immune failure. Depression is the booster of tumor immune editing. Therefore， the method of reinforcing the healthy Qi and removing toxins was proposed to regulate the immune editing and cut off the five-stage evolution of tumors. Supplementing Qi and warming Yang can reinforce the healthy Qi and restore immune surveillance. Removing toxins and dredging can reverse toxins and immune escape. The harmonizing method can maintain the dynamic balance of immune cells/immunosuppressive cells. Resolving depression can truncate tumor immune editing. Those methods can provide a certain reference for the treatment based on microscopic syndrome differentiation in traditional Chinese medicine （TCM）. In future studies， it is necessary to further explore the specific mechanism of the regulation of immune editing with the methods of supplementing Qi and warming Yang， removing toxins and dredging， their combination， and resolving depression， so as to find out specific Chinese medicines and targets and provide more sufficient evidence for the regulation of tumor immune editing by TCM.

Objective : To investigate the regulatory role of ubiquitin-specific protease 10 ( USP10) on the protein expression of Krüppel-like factor 4 (KLF4) and its impact on the proliferation and invasion ability of hepatocellular carcinoma (HCC) cells． Methods : The protein expression differences of USP10 and KLF4 in normal liver cell line L02 and HCC cell lines，including HepG2，HUH7，HCCLM3 were detected by immunoblotting ( Western blot ) methods．HCCLM3 and HUH7 cells were selected，and lentiviral particles overexpressing or silencing USP10 ( oe- USP10 or sh-USP10) was transfected into the cells，and they were designated as the oe-USP10 group and oe-NC group，respectively．Immunoprecipitation ( Co-IP) experiments were conducted to examine whether USP10 could di- rectly interact with KLF4 in HCCLM3 or HUH7 cells．The Co-IP assay was repeated in HCC cells transfected with oe-USP10 or sh-USP10，with the addition of the proteasome inhibitor MG132，which used to detect the ubiquitina- tion level of KLF4 protein in the transfected HCC cells．The pcDNA3. 1 vector containing overexpressed KLF4 or its negative control plasmid (pc-KLF4 or pc-NC) was co-transfected into cells of the sh-USP10 group or sh-NC group. These cells were designated as the sh-NC + pc-NC group，sh-USP10 + pc-NC group，sh-NC + pc-KLF4 group，and sh-USP10 + pc-KLF4 group．The cell proliferation activity of each group was measured using the CCK-8 assay，and the cell invasion ability was assessed using the Transwell assay. Results : Compared to L02 cells，the protein expres- sion of USP10 and KLF4 significantly decreased in HepG2，HUH7，HCCLM3，and other cells (P＜0. 05) ．In HC- CLM3 and HUH7 cells，USP10 protein directly interacted with KLF4．Furthermore，treatment with MG132 resulted in a time-dependent increase in KLF4 protein expression in HCCLM3 and HUH7 cells．Silencing USP10 increased the ubiquitination of KLF4 in HCCLM3 or HUH7 cells，while overexpressing USP10 decreased the ubiquitination level of KLF4 in cells．Compared to the sh-NC + pc-NC group，both the proliferation activity and invasion ability of HCCLM3 and HUH7 cells significantly increased in the sh-USP10 + pc-NC group (P ＜0. 01) ，while they signifi- cantly decreased in the sh-NC + pc-KLF4 group and sh-USP10 + pc-KLF4 group (P＜0. 05) ．Compared to the sh- USP10 + pc-NC group，the proliferation activity and invasion ability of cells significantly decreased in the sh-USP10 + pc-KLF4 group (P＜0. 05) ． Conclusion USP10 can promote the stability of KLF4 protein through deubiquiti- nation in HCC cell lines，thereby inhibiting the proliferation and invasion of tumor cells.