Objective:To construct high level prokaryotic cell expression vector carrying human upstream regulatory element binding protein1 (hureb1) gene for producing GST hureb1 fusion protein and generating anti hureb1 antibody Methods: After the Xho I/Not I fragment from hureb1 open reading frame was recombined into the prokaryotic expression vector pGEX 4T 2, the vector was transformed into E coli BL21(DE3) strain and induced with IPTG to express the fusion protein, GST hureb1 The crudely isolated fusion protein was applied to immunize rabbit and mouse for generating anti hureb1 polyclonal antibody and monoclonal antibody respectively Results:The prokaryotic expression vector expressing GST hureb1 fusion protein was constructed and induced to produce high level GST hureb1 that was detected up to 33 45% of the total bacterial protein expressed The GST hureb1 fusion protein immunizing animals generated high titer and specific anti hureb1 antibodies Conclusion:Recombinant hureb1 protein and anti hureb1 antibodies can be used to analyze the biological functions of hureb1

Objective:To investigate the effects of hydrogen rich-saline (HRS) on intestinal mucosal barrier in rat with intestinal ischemia/reperfusion injury (IIRI).Methods:Twenty-four healthy male Sprague-Dawley rats, aged 8 weeks, were randomly divided into 3 groups (8 in each group) by random number table method: sham group, model group and HRS group.Rats in HRS group were intraperitoneally injected with HRS (10 mL/kg) at 30 min of ischemia, and the same amount of normal saline was intraperitoneally injected in model group.After 45 min of ischemia and 6 h of reperfusion, rats were sacrificed.Serum and ileum were collected for further detection.Tumor necrosis factor alpha (TNF-α), interleukin (IL)- 1β and IL-17A expression levels in serum were detected by conducting enzyme-linked immunosorbent assay (ELISA). The localization expressions of tight junction protein Occludin was detected by immunohistochemical staining (IHC), while the localization expression of tight junction protein zonula occluden-1 (ZO-1) were detected by immunofluorescence staining (IF). The protein expression of Occludin, ZO-1, and Lysozyme were detected by performing Western blot.The mRNA expression of Lysozyme and α-defensin were detected by real-time PCR (qPCR).Results:ELISA results proved that the levels of serum TNF-α and IL-1β in HRS group rats were significantly lower than those in model group [(62.02±29.97) ng/L vs.(113.40±44.58) ng/L, (21.68±0.35) ng/L vs.(28.29±3.49) ng/L], while the level of IL-17A increased [(28.18±5.28) ng/L vs. (15.10±3.60) ng/L] (all P<0.05). IHC staining: compared with model group, the expression of Occludin in HRS group was uniform and continuous, and the staining was darker.IF results: compared with model group, the fluorescence signal intensity of ZO-1 in HRS group rats significantly increased, and the distribution was clear and continuous.Wes-tern blot results: compared with model group, the expression levels of Occludin and ZO-1 proteins in HRS group rats remarkably increased (0.79±0.06 vs. 0.54±0.04, 0.91±0.11 vs. 0.51±0.13), while Lysozyme protein decreased (1.50±0.40 vs. 2.99±0.80) (all P<0.05). qPCR results revealed that the expression level of Lysozyme mRNA in HRS group rats was lower than that in model group (1.64±0.33 vs. 2.20±0.40), while α-defensin mRNA obviously increased (0.82±0.19 vs. 0.47±0.13) (all P<0.01). Conclusions:HRS protects intestinal mucosal barrier by inhibiting the expression of tight junctions and the secretion of antimicrobial peptides in rat suffering from IIRI.

Objective:To analyze clinical characteristics of patients with novel coronavirus Omicron variant of concern infection, and to provide practical data and experience for subsequent clinical treatment.Methods:A retrospective analysis was performed for the clinical data of 5 cases with novel coronavirus Omicron variant of concern infection treated in the First Hospital of Jiaxing from December 18, 2021 to January 28, 2022. The patients' clinical data were recorded, including gender, age, length of hospital stay, vaccination status, clinical symptoms, laboratory indicators [white blood cell count (WBC), lymphocyte count (LYM), eosinophil count (EOS), hypersensitivity C-reactive protein (hs-CRP), novel coronavirus antibody immunoglobulin (IgG and IgM)], chest CT, treatment course and disease outcome.Results:All 5 patients were male, aged 24-37 years old. Four patients were vaccinated with novel coronavirus vaccine (one patient received 3 doses of the vaccine and 3 patients received only the first 2 doses of the vaccine), and no infection was found in chest CT. Laboratory examination showed that WBC, LYM, EOS and hs-CRP levels were normal, and only showed mild symptoms of upper respiratory tract infection. One patient was not vaccinated with novel coronavirus vaccine, and signs of viral pneumonia could be seen in chest CT, laboratory examination showed that WBC and hs-CRP levels increased, suggesting that bacterial infection, fever, cough, sputum and other respiratory symptoms were obvious, and the treatment time was long. All 5 patients were treated with Chinese medicine Lotus antipyretic and Baihu Yinqiao decoction based on routine antiviral therapy.Conclusions:Patients with novel coronavirus Omicron variant of concern infection vaccinated with the novel coronavirus vaccine have milder clinical symptoms, with less obvious chest CT findings and faster recovery. Chinese medicine Lotus antipyretic and Baihu Yinqiao decoction has obvious therapeutic effect on such patients.

Objective　To observe the short-term and long-term effects of leptin on hepatic glucose oxidation and glucokinase gene expression. Methods　Rat hepatic cell line BRL was incubated with leptin of different doses (range from 10?ng/ml-200?ng/ml) for 1?h or 24?h. Glucose oxidation was determined by liquid scintillation counting. Glucokinase gene expression (corrected by β-actin) was determined by reverse transcription semi-quantitative polymerase chain reaction. Results　Treatment with leptin 10?ng/ml for 1?h had no significant effects on glucose oxidation in hepatic cells. However, at the doses ranging from 50?ng/ml to 200?ng/ml, leptin significantly inhibited glucose oxidation. These effects disappeared when the hepatic cells were exposed to leptin for 24?h. Glucokinase mRNA expression was reduced significantly after both 1?h and 24?h exposure to leptin (100?ng/ml) as compared to that of the control group. Conclusion　A low dose of leptin has no significant effect on glucose oxidation in hepatic cells. A relatively high dose of leptin has an acute inhibitory effect on the glucose oxidation in hepatic cells. This effect may likely involve the inhibition of glucokinase gene expression. The inhibitory effect on glucose oxidation is transient and disappears with prolonged exposure time.

Improving liver regeneration (LR) remains a medical issue, and there is currently a lack of safe and effective drugs for LR. Rhizoma Dioscoreae (SanYak, SY) is a traditional Chinese medicine. However, the underlying action mechanism of SY treatment for LR is yet to be fully elucidated. To explore the mechanism by which SY affects LR, we have conducted a series of methods for network pharmacological analysis, molecular docking, and in vivo experimental validation in mice. Overall, 9 compounds and 30 predicted target genes of SY were found to be associated with the therapeutic effects of LR. Compared with the model group, hematoxylin and eosin staining revealed that the mice with preoperative drug intervention possessed fewer postoperative hepatocyte bubbles and relatively regular morphology. Furthermore, the serum alanine transaminase and aspartate aminotransferase levels were reduced, immunohistochemistry revealed elevated proliferating cell nuclear antigen positivity rate, and Western blotting demonstrated that the phospho-protein kinase B (AKT)/AKT ratio was downregulated and that vascular endothelial growth factor A (VEGFA) expression levels were upregulated. This study explored dioscin, the main active ingredient of SY, and its potential therapeutic effects on LR. It repairs damaged liver following surgery and promotes liver cell proliferation. The action mechanism comprises reducing AKT phosphorylation levels and upregulating VEGFA expression levels. Thus, this study provides a new direction for further research on the mechanism of SY promoting LR.

Improving liver regeneration (LR) remains a medical issue, and there is currently a lack of safe and effective drugs for LR. Rhizoma Dioscoreae (SanYak, SY) is a traditional Chinese medicine. However, the underlying action mechanism of SY treatment for LR is yet to be fully elucidated. To explore the mechanism by which SY affects LR, we have conducted a series of methods for network pharmacological analysis, molecular docking, and in vivo experimental validation in mice. Overall, 9 compounds and 30 predicted target genes of SY were found to be associated with the therapeutic effects of LR. Compared with the model group, hematoxylin and eosin staining revealed that the mice with preoperative drug intervention possessed fewer postoperative hepatocyte bubbles and relatively regular morphology. Furthermore, the serum alanine transaminase and aspartate aminotransferase levels were reduced, immunohistochemistry revealed elevated proliferating cell nuclear antigen positivity rate, and Western blotting demonstrated that the phospho-protein kinase B (AKT)/AKT ratio was downregulated and that vascular endothelial growth factor A (VEGFA) expression levels were upregulated. This study explored dioscin, the main active ingredient of SY, and its potential therapeutic effects on LR. It repairs damaged liver following surgery and promotes liver cell proliferation. The action mechanism comprises reducing AKT phosphorylation levels and upregulating VEGFA expression levels. Thus, this study provides a new direction for further research on the mechanism of SY promoting LR.

Improving liver regeneration (LR) remains a medical issue, and there is currently a lack of safe and effective drugs for LR. Rhizoma Dioscoreae (SanYak, SY) is a traditional Chinese medicine. However, the underlying action mechanism of SY treatment for LR is yet to be fully elucidated. To explore the mechanism by which SY affects LR, we have conducted a series of methods for network pharmacological analysis, molecular docking, and in vivo experimental validation in mice. Overall, 9 compounds and 30 predicted target genes of SY were found to be associated with the therapeutic effects of LR. Compared with the model group, hematoxylin and eosin staining revealed that the mice with preoperative drug intervention possessed fewer postoperative hepatocyte bubbles and relatively regular morphology. Furthermore, the serum alanine transaminase and aspartate aminotransferase levels were reduced, immunohistochemistry revealed elevated proliferating cell nuclear antigen positivity rate, and Western blotting demonstrated that the phospho-protein kinase B (AKT)/AKT ratio was downregulated and that vascular endothelial growth factor A (VEGFA) expression levels were upregulated. This study explored dioscin, the main active ingredient of SY, and its potential therapeutic effects on LR. It repairs damaged liver following surgery and promotes liver cell proliferation. The action mechanism comprises reducing AKT phosphorylation levels and upregulating VEGFA expression levels. Thus, this study provides a new direction for further research on the mechanism of SY promoting LR.

Improving liver regeneration (LR) remains a medical issue, and there is currently a lack of safe and effective drugs for LR. Rhizoma Dioscoreae (SanYak, SY) is a traditional Chinese medicine. However, the underlying action mechanism of SY treatment for LR is yet to be fully elucidated. To explore the mechanism by which SY affects LR, we have conducted a series of methods for network pharmacological analysis, molecular docking, and in vivo experimental validation in mice. Overall, 9 compounds and 30 predicted target genes of SY were found to be associated with the therapeutic effects of LR. Compared with the model group, hematoxylin and eosin staining revealed that the mice with preoperative drug intervention possessed fewer postoperative hepatocyte bubbles and relatively regular morphology. Furthermore, the serum alanine transaminase and aspartate aminotransferase levels were reduced, immunohistochemistry revealed elevated proliferating cell nuclear antigen positivity rate, and Western blotting demonstrated that the phospho-protein kinase B (AKT)/AKT ratio was downregulated and that vascular endothelial growth factor A (VEGFA) expression levels were upregulated. This study explored dioscin, the main active ingredient of SY, and its potential therapeutic effects on LR. It repairs damaged liver following surgery and promotes liver cell proliferation. The action mechanism comprises reducing AKT phosphorylation levels and upregulating VEGFA expression levels. Thus, this study provides a new direction for further research on the mechanism of SY promoting LR.

Improving liver regeneration (LR) remains a medical issue, and there is currently a lack of safe and effective drugs for LR. Rhizoma Dioscoreae (SanYak, SY) is a traditional Chinese medicine. However, the underlying action mechanism of SY treatment for LR is yet to be fully elucidated. To explore the mechanism by which SY affects LR, we have conducted a series of methods for network pharmacological analysis, molecular docking, and in vivo experimental validation in mice. Overall, 9 compounds and 30 predicted target genes of SY were found to be associated with the therapeutic effects of LR. Compared with the model group, hematoxylin and eosin staining revealed that the mice with preoperative drug intervention possessed fewer postoperative hepatocyte bubbles and relatively regular morphology. Furthermore, the serum alanine transaminase and aspartate aminotransferase levels were reduced, immunohistochemistry revealed elevated proliferating cell nuclear antigen positivity rate, and Western blotting demonstrated that the phospho-protein kinase B (AKT)/AKT ratio was downregulated and that vascular endothelial growth factor A (VEGFA) expression levels were upregulated. This study explored dioscin, the main active ingredient of SY, and its potential therapeutic effects on LR. It repairs damaged liver following surgery and promotes liver cell proliferation. The action mechanism comprises reducing AKT phosphorylation levels and upregulating VEGFA expression levels. Thus, this study provides a new direction for further research on the mechanism of SY promoting LR.

Objective:To investigate the effects of down-regulation of Krüppel-like factor 5 (KLF5) on biological functions of rat intestinal epithelial cells IEC-6 in response to ionizing radiation.Methods:Rat intestinal epithelial IEC-6 cells were irradiated with 0, 2, 4, 8, 12, 16 Gy of X-rays and 3 h later the expression of KLF5 in IEC-6 cells was detected by Western blot. IEC-6 cells were irradiated to 12 Gy and 0, 0.5, 1, 2, 3, 5, 7 and 24 h later the expression of KLF5 in IEC-6 cells was detected by Western blot. shRNA sequences targeting rat KLF5 gene were designed, synthesized and inserted into the lentiviral vector. The recombinant lentiviral vectors were packaged in human embryonic kidney 293T cells, and the lentivirus titers were determined. IEC-6 cells were infected with the recombinant lentivirus, and the transfection efficiency was observed under fluorescence microscope. Real-time PCR and Western blot were used to detect the expressions of KLF5 mRNA and protein in the transfected cells 72 h post transfection. The consequent experiments included four groups: negative control group, shKLF5 group, radiation group and radiation + shKLF5 group. The cell viability was observed in KLF5 silencing cells irradiated with 8 Gy by using CCK-8 assay. The cell cycle distribution and apoptosis were detected in KLF5 silencing cells irradiated with 8 Gy by flow cytometry. Immunofluorescence staining was applied to visualize the γ-H2AX foci in nucleus of KLF5 silencing cells irradiated with 2 Gy.Results:The expression of KLF5 increased with the different doses. The expression of KLF5 increased first, then decreased and peaked at 5 h post-irradiation with 12 Gy. KLF5 shRNA lentiviral vectors were successfully constructed. The mRNA and protein level of KLF5 were down-regulated in recombinant lentivirus transfected IEC-6 cells 72 h after transfection. Knockdown of KLF5 markedly induced G 2/M phase arrest ( t=11.56, P<0.05), proliferation inhibition, more apoptosis rate [radiation group: (7.42±0.49)%, radiation + shKLF5 group: (12.49±0.63)%, t=10.98, P<0.05], and more γ-H2AX foci in nucleus post-irradiation than negative control ( t=22.07, 23.89, 11.24, 59.97, 20.85, P<0.05). Conclusions:The KLF5 knockdown intestinal epithelial cell line was successfully established. The down-regulation of KLF5 expression could induce cell arrest at G 2/M, suppress the proliferation of irradiated cells and improve the cell apoptosis, enhance DNA double strand breaks and prolong DNA damage repair.