Objective:To screen differentially expressed genes (DEGs) associated with liver cancer by bioinformatics analysis method, and to investigate the mechanism of the minichromosome maintenance protein 2 ( MCM2) and replication factor C subunit 4 ( RFC4) genes in liver cancer in vitro. Methods:Gene expression profiling data of 80 and 36 hepatocellular carcinoma tissues and 307 and 13 cirrhotic tissues were obtained from the GSE25097 and GSE98620 datasets of the gene expression analysis (GEO) database, respectively. Gene expression profiling data of 374 liver cancer tissues and 50 normal liver tissues were downloaded from the cancer genome atlas (TCGA) database. Limma and DESeqs R software were used to process the gene expression profiling data, construct protein-protein interaction networks, and analysis the relevance of these genes to survival. Weighted gene co-expression network analysis was performed to screen out the core genes. Liver cancer SMMC7721 cells were transfected with MCM2 blank plasmid (MCM2 control group), MCM2 overexpression plasmid [MCM2 WT1 group, MCM2 WT2 (2-fold WT1) group], RFC4 blank plasmid (RFC4 control group), and RFC4 overexpression plasmid [RFC4 WT1 group, RFC4 WT2 (2-fold WT1) group], respectively. The expression of MCM2 and RFC4 in liver cancer cell lines and their transfection levels were detected by real-time fluorescence quantitative reverse transcription PCR and Western blotting. The effects of MCM2 and RFC4 on the proliferation of hepatocellular carcinoma cells were detected by MTT assay and cell cloning assay, respectively. The effects of MCM2 and RFC4 on the migration of liver cancer cells were determined by the scratch assay. The effects of MCM2 and RFC4 on liver cancer cell invasion were detected by Transwell assay.Results:By bioinformatic analysis, 9 HCC DEGs were selected, including ubiquitin conjugating enzyme E2 T ( UBE2T), aurora kinase A ( AURKA), targeting protein for Xklp2 ( TPX2), MCM2, RFC4, ribonucleoside-diphosphate reductase subunit M2 ( RRM2), serine peptidase inhibitory factor Kazal type 1 ( SPINK1), collagen type XV alpha 1 chain ( COL15A1) and C-C motif chemokine 25 ( CCL25). Among the six genes associated with clinical stages, the MCM2 and RFC4 genes were found to be strongly associated with prognosis in liver cancer. The relative protein expression of MCM2 and RFC4 in HepG2 cells (1.83±0.07, 1.44±0.09) and SMMC7721 cells (1.74±0.05, 1.43±0.08) was higher than that in MIHA cells (1.00±0.02, 1.00±0.03), and all the differences were statistically significant (all P<0.05). The relative gene expression of MCM2 and RFC4 in HepG2 cells (14.30±0.12, 5.10±0.18) and SMMC7721 cells (10.60±0.11, 7.60±0.07) was higher than that in MIHA cells (1.00±0.05, 1.00±0.03), and all the differences were statistically significant (all P<0.05). Compared with the MCM2 control group, the absorbance values [(0.28±0.01 and 0.21±0.01) vs 0.18±0.03], the number of clonal cells [(717±12 and 782±29) cells vs (389±17) cells], the percentage migration [(0.43±0.02 and 0.68±0.01) vs 0.15±0.06], and the number of cellular invasions [(933±21 and 821±11) cells vs (409±16) cells] were higher in the MCM2 WT1 and MCM2 WT2 groups, and the differences were all statistically significant (all P<0.05). Compared with the RFC4 control group, the absorbance values [(0.30±0.02 and 0.21±0.01) vs 0.17±0.02], the number of cloned cells [(571±11 and 728±9) cells vs (373±23) cells], the percentage migration [(0.75±0.11 and 0.67±0.04) vs 0.34±0.07], and the number of cell invasion [(835±26 and 818±18) cells vs (629±12) cells] were higher in the RFC4 WT1 and the RFC4 WT2 groups, and the differences were statistically significant (all P<0.05). Conclusions:MCM2 and R FC4 genes play a role in promoting tumorigenesis and growth in hepatocellular carcinoma.

Objective:To explore the expression and clinical significance of fibrinogen-like protein 2(FGL2)in gastrointesti-nal stromal tumor(GIST).Methods:Immunohistochemical EnVision two-step method was used to detect expression of FGL2 protein in 158 GIST patients,and correlation between FGL2 protein and clinicopathological parameters was analyzed,and relationship be-tween FGL2 protein and the count of CD3+T,CD8+T,CD20+T and CD68+T of tumor-infiltrating lymphocytes(TILs)in GIST was inves-tigated.Results:Expression of FGL2 protein in GIST was negatively correlated with tumor size,NIH risk grade,necrosis and mitotic image count(P<0.05),there was no significant difference in gender and tumor location(P>0.05).FGL2 expression was associated with low CD3+T cell counts in GIST(P<0.05),while there was no significantly difference with CD8+T,CD20+T and CD68+T cell counts(P>0.05).Conclusion:FGL2 protein is involved in occurrence and development of GIST,which expression level is partly pre-dictive of the malignant potential of GIST.FGL2 protein may participate in occurrence and development of GIST by regulating tumor microenvironment.

Background: Colorectal cancer is one of the most commonly diagnosed cancer and leading causes of cancer death worldwide. Fibrinogen-like protein 2 (FGL2), a member of the fibrinogen family, has been demonstrated as a regulator of immune cell functions in tumor microenvironment and might facilitate tumor progression. Aims: To study the expression and clinical significance of FGL2 in colon cancer. Methods: One hundred and fifty colon cancer patients diagnosed from January 2018 to January 2022 at the Second Hospital of Tianjin Medical University were enrolled in this study. The paraffin-embedded tissues were collected for detection of FGL2 protein and the surface markers of tumor infiltrating immune cells (TIICs), including CD4, CD8, CD20, CD68, and CD56 by using immunohistochemistry. The correlations of FGL2 expression level with the clinicopathological parameters and TIICs counting were analyzed. Results: Expression of FGL2 was upregulated in 68.7% of the colon cancer cases. Its expression level was correlated positively with the tumor size and TNM staging (all P<0.05), while no correlations were found between FGL2 expression level and gender, age, tumor differentiation, and presence of vascular invasion (all P>0.05). Meanwhile, the expression level of FGL2 was associated with the cell counting of CD4

Autophagy is a special cellular process, which can participate in regulating cell survival, growth, differentiation and homeostasis maintenance by transporting damaged organelles and macromolecular substances to lysosomes for degradation. Autophagy plays a very important role in all aspects of life process. Research results show that autophagy plays an important role in tumor occurrence, development and metastasis. The synergistic effect of autophagy and vascular endothelial growth factor promotes tumor angiogenesis and cell repair, and may play an important role in the development of tumor resistance to anti-vascular drug therapy. Targeted therapy with autophagy as the target may be a new direction for anti-tumor molecular targeted therapy in the future, aiming to provide multi-target collaborative therapy to benefit patients.

Objective:To investigate the role of autophagy mediated by mTOR signaling pathway in the inhibition of osteogenic differentiation of human bone marrow mesenchymal stem cells (hBMSCs) induced by cadmium.Methods:HBMSCs were divided into 0, 2.5 or 5.0 μmol/L groups according to the exposure dose of cadmium chloride (CdCl 2), and each group was treated for 1 day, 4 days and (or) 7 days. The ALP activity and mRNA and protein expression levels of osteogenesis markers (ALP, RUNX2 and OSTERIX), autophagy-related proteins (LC3 and Beclin-1) and mTOR signaling pathway related proteins (mTOR, p-mTOR and p-p70S6K) expression, alkaline phosphatase staining and alizarin red staining were detected. MHY 1485 was selected as the signaling pathway activator. The control group, CdCl 2 group (5.0 μmol/L), MHY 1485 group and CdCl 2+MHY 1485 combined treatment group were set. After 7 days of treatment, the expression levels of autophagy related proteins and mTOR signaling pathway related proteins of hBMSCs in each group were detected. Results:There was no significant difference in ALP activity between 0, 2.5 and 5.0 μmol/L groups on day 1 and 4 ( P>0.05); On day 7, compared with the 0 μmol/L group, the ALP activity, expression of osteogenic markers (ALP, RUNX2, OSTERIX) and mTOR signaling pathway related proteins (mTOR, p-mTOR, p-p70S6K) expression decreased in the 2.5 and 5.0 μmol/L group ( P<0.05). Compared with the 0 μmol/L group, the staining of the 2.5 and 5.0 μmol/L groups became lighter, and the formation of ALP and mineralized nodules was reduced. Compared with the CdCl 2 group, the autophagy related protein expression in the CdCl 2+MHY 1485 combined treatment group decreased, and the mTOR signaling pathway related protein expression increased. The difference was statistically significant ( P<0.05). Conclusion:The inhibition of osteogenic differentiation of hBMSCs by cadmium may be related to autophagy mediated by mTOR signaling pathway.

Objective:To investigate the role of autophagy mediated by mTOR signaling pathway in the inhibition of osteogenic differentiation of human bone marrow mesenchymal stem cells (hBMSCs) induced by cadmium.Methods:HBMSCs were divided into 0, 2.5 or 5.0 μmol/L groups according to the exposure dose of cadmium chloride (CdCl 2), and each group was treated for 1 day, 4 days and (or) 7 days. The ALP activity and mRNA and protein expression levels of osteogenesis markers (ALP, RUNX2 and OSTERIX), autophagy-related proteins (LC3 and Beclin-1) and mTOR signaling pathway related proteins (mTOR, p-mTOR and p-p70S6K) expression, alkaline phosphatase staining and alizarin red staining were detected. MHY 1485 was selected as the signaling pathway activator. The control group, CdCl 2 group (5.0 μmol/L), MHY 1485 group and CdCl 2+MHY 1485 combined treatment group were set. After 7 days of treatment, the expression levels of autophagy related proteins and mTOR signaling pathway related proteins of hBMSCs in each group were detected. Results:There was no significant difference in ALP activity between 0, 2.5 and 5.0 μmol/L groups on day 1 and 4 ( P>0.05); On day 7, compared with the 0 μmol/L group, the ALP activity, expression of osteogenic markers (ALP, RUNX2, OSTERIX) and mTOR signaling pathway related proteins (mTOR, p-mTOR, p-p70S6K) expression decreased in the 2.5 and 5.0 μmol/L group ( P<0.05). Compared with the 0 μmol/L group, the staining of the 2.5 and 5.0 μmol/L groups became lighter, and the formation of ALP and mineralized nodules was reduced. Compared with the CdCl 2 group, the autophagy related protein expression in the CdCl 2+MHY 1485 combined treatment group decreased, and the mTOR signaling pathway related protein expression increased. The difference was statistically significant ( P<0.05). Conclusion:The inhibition of osteogenic differentiation of hBMSCs by cadmium may be related to autophagy mediated by mTOR signaling pathway.

Objective:To investigate the effect of exercise rehabilitation combined with psychotherapy and exercise rehabilitationn alone in elderly patients with coronary heart disease(CHD)after percutaneous coronary intervention(PCI).Methods:A total of 150 elderly patients(≥65 years)admitted to our hospital with CHD after primary PCI were enrolled and randomly divided into the group A(conventional drug therapy), group B(conventional drug therapy+ exercise rehabilitation)and group C(conventional drug therapy + exercise rehabilitation+ psychotherapy), with 50 patients in each group.Patients in groups B and C were intervened at two weeks after PCI, once a week for 12 weeks.Changes of Hamilton anxiety scale(HAMD)scores, Hamilton depression scale(HAMA)score, 6 min walking test, left ventricular ejection fraction(LVEF)and metabolic equivalents(METs)were compared among the three groups.Results:Before treatment, the METs were significantly lower in the group B(3.58±0.14)compared with the groups A(3.69±0.18)and C(3.68±0.15), and were similar between groups A and C. After 12 weeks of treatment, compared with the indicators before treatment, the HAMA scores were similar in the group A(12.98±2.51 vs. 12.16±2.91, P>0.05), and were significantly decreased in groups B(12.90±2.12 vs.8.06±2.11, P<0.05)and C(13.03±2.52 vs.6.96±2.13, P<0.05); HAMD scores were all markedly decreased in all three groups(group A: 22.38±2.52 vs.20.87±2.12; group B: 22.58±2.57 vs.17.25±2.32; group C: 22.23±2.35 vs.13.39±2.25), and were decreased most in the group B, followed by groups A and B( P<0.05). Furthermore, the LVEF group A: (49.08±1.59)% vs.(52.15±1.91)%; group B: (48.99±2.11)% vs.(57.56±2.13)%; group C: (49.04±2.02)% vs.(59.92±1.93)%, 6 min walking distance test(m)(group A: 360.78±12.50 vs.370.16±12.41; group B: 359.21±10.54 vs.394.19±15.56; group C: 363.12±15.28 vs.413.29±18.15)and METs(group A: 3.69±0.18 vs.3.91±0.21; group B: 3.58±0.14 vs.4.89±0.09; group C: 3.68±0.15 vs.5.77±0.13)were significantly improved in all groups after the treatment.Among them, group C was improved most, followed by groups B and A(all P<0.05). Conclusions:Exercise rehabilitation combined with psychotherapy is of great significance to improve the physical condition of elderly patients with CHD after PCI, and it is better than the routine exercise rehabilitation alone.

Objective: A new ion exchange column technology was used to establish an efficient and sensitive method for the detection of inorganic arsenic. Methods: Based on the new As Specia Fast Column, the pretreatment methods, liquid phase separation and mass spectrometry determination conditions of inorganic arsenic in rice were optimized. Finally, arsenic compounds were separated by As Specia Fast Column and detected by liquid chromatography inductively coupled plasma mass spectrometry. The external standard method was used for quantitative analysis. The detection limit, precision and accuracy of the method were determined by measuring the content of arsenic compounds in rice samples and rice standard samples. At the same time, three Guangdong rice samples were selected as the experimental samples of this study, and 1 g of each sample was weighed and measured in parallel three times. The method was compared with the method of liquid chromatography-atomic fluorescence spectrometry (LC-AFS) and liquid chromatography-inductively coupled plasma mass spectrometry (LC-ICP-MS) in the national standard. Results: The inorganic arsenic in rice was extracted with 0.5% nitric acid solution at 65 ℃ for 15 h, and the pH was adjusted to alkaline. The mobile phase A (8 mmol/L HNO₃, 50 mmol/L NH₃·H₂O) and mobile phase B (40 mmol/L HNO₃, 80 mmol/L NH₃·H₂O) were used as the mobile phase gradient elution (93%) . Five arsenic compounds can reach baseline separation under the conditions of RF power of 1 500 W and atomization gas flow of 0.97 L/min. The detection limits ranged from 0.114 to 0.331 μg/L, and the inorganic arsenic content in rice samples ranged from 0.063 to 0.232 mg/kg. The results of determination of arsenic compounds in rice flour reference materials were all within the uncertainty range indicated by the standard. The recoveries were 86.7%~106.7%, and the precision was 1.9%-12.5%. Compared with national standards, the results of determination of arsenate in rice were relatively close (using this method, LC-AFS, LC-ICP-MS to detect the content of arsenate in rice samples 1 was 0.231, 0.226, 0.236 mg/kg, respectively). However, due to insufficient sensitivity, the national standard method is difficult to detect low levels of arsenic compounds (Arsenobetaine was not detected in rice sample 1). The method can detect the content of arsenobetaine in rice sample 1 was 0.023 mg/kg. Conclusion The established method can meet the requirements of inorganic arsenic determination in rice, and it is more rapid and accurate than the current national standard. It can better monitor and evaluate the content of i-As in rice, and provide accurate data for comprehensively grasping and evaluating the safety of rice consumption of residents.

Objective To explore whether receptor interacting protein (RIP)1/RIP3 pathways participate in glutamate induced cell death in HT-22 neuronal cells and investigate the potential neuroprotection ofnecrostatin-1 in glutamate induced cell death in HT-22.Methods (1) In vitro cultured mouse hippocampal neuronal HT-22 cells were divided into control group,zVAD group,necrostatin-1 (Nec-1) group,glutamate group,glutamate+zVAD group,glutamate+zVAD+Nec-1 group and glutamate+Nec-1 group;they were treated with zVAD,Nec-1 and glutamate at the final concentrations of 20 μmol/L,30 μmol/L and 3 mmol/L for 24 h.Cell viability was detected using a luminescence-based commercial kit Cell Titer-Glo (CTG).Necrotic cell death was measured by propidium iodide (PI) and HE stainings.(2) HT-22 cells were divided into control group Ⅰ,glutamate group Ⅰ and glutamate+Nec-1 group Ⅰ;MitoSox Red was used to detect mitochondrial reactive oxygen species (ROS) level.(3) HT-22 cells were divided into control group Ⅱ,glutamate group Ⅱ and glutamate+tertiary butyl-hydroxyanisole (BHA) group;the final concentration of BHA was 100 μmol/L;necrotic cell death was measured by PI and HE stainings after 24 h of treatment.(4) HT-22 cells were divided into RIP3 siRNA and control group Ⅲ,and then,they were transfected with RIP3 siRNA or negative siRNA,respectively;the RIP3 protein expression was determined by Westem blotting after 72 h of treatment.(5) HT-22 cells were divided into negative siRNA+Control,RIP3 siRNA,negative siRNA+glutamate and RIP3 siRNA+glutamate groups;the cells were transfected with RIP3 siRNA or Negative siNRA,respectively;48 h later,the glutamate groups were treated with 3 mmol/L glutamate;PI positive cells and cell viability were measured by PI and HE stainings and CTG at 24 h after glutamate treatment.Results (1) As compared with control group,percentage of PI positive cells was greatly increased and cell viability was decreased in glutamate group and glutamate+zVAD group,with statistically significant differences (P＜0.05);as compared with those in the glutamate group,percentage of percentage of PI positive cells was was significantly decreased and cell viability was statistically increased in glutamate+Nec-1 group (P＜0.05).(2) ROS level in HT-22 cells of the glutamate group was significantly increased than that in the control group Ⅰ (P＜0.05);however,ROS level in HT-22 cells of glutamate+Nec-1 group Ⅰ was significantly decreased than that in glutamate group Ⅰ (P＜0.05).(3)Percentage of PI positive cells in the glutamate group Ⅱ was significantly higher than that in the control group Ⅱ (P＜0.05),and that in the glutamate+BHA group was statistically lower than that in the glutamate group Ⅱ (P＜0.05).(4) The RIP3 protein expression in the RIP3 siRNA group was obviously down-regulated as compared with that in the control group Ⅲ.(5) As compared with those in the negative siRNA group,percentage of PI positive cells was statistically increased and cell viabilities were statistically decreased in glutamate group (P＜0.05);however,percentage of PI positive cells was significantly decreased and cell viability was significantly increased in RIP3 siNRA+glutamate group as compared with those in the glutamate group (P＜0.05).Conclusion RIP1/RIP3 pathway and ROS might mediate glutamate induced cell death in HT-22 cells.

Objective To investigate the effect of long?term exposure to environmental cadmium on eight mineral element's metabolic balance of human body. Methods To choose a high cadmium area polluted by smelting and mining north of Guangdong province and a cadmium?free area with a similar economic level, and living and eating habit of residents as a contrast from April 2011 to August 2012. Stratified random sampling and clustered sampling method were adopted to choose the non?occupationally cadmium?exposed respondents who have lived in local area for more than 15 years, older than 40 years, having local rice and vegetable as the main dietary source, with simple and relatively stable diet, and without diabetes, kidney disease, thyroid disease, liver disease or other history of chronic disease. This study included 298 respondents, of whom 155 were in cadmium exposure group and 143 in control group. Questionnaires was used to acquire their health status and their morning urine samples were collected. Electrolytically coupled plasma mass spectrometry (ICP?MS) was used to test the concentrations of sodium (Na), magnesium (Mg), phosphorus (P), potassium (K), calcium (Ca), copper (Cu), zinc (Zn) and iodine (I). The Mann?Whitney U test method was used to compare the differences of concentrations of urinary cadmium, Na, Mg, P, K, Ca, Cu, Zn, I, and the ratio of Na to K (Na/K), Ca to P (Ca/P) between exposed group and control group.χ2 test was used to compare the abnormal rate of urinary cadmium between exposed group and control group. Pearson correlation and multiple regression method were used to investigate the relationship between urinary cadmium levels, gender, age, smoking, passive smoking, and minerals. Results The urinary cadmium level P50 (P25-P75) in exposed group was 5.45 (2.62-10.68)μg/g·cr, which was higher than that of the control group, which was 1.69 (1.22-2.36)μg/g · cr (Z=-10.49, P<0.001). The abnormal rate of urinary cadmium was 51.6%(80/155), which was higher than that of the control group (2.8%(4/143)) (χ2=87.56,P<0.001). The urinary Ca, Cu, Zn, and I level P50 (P25-P75) of exposed group were 173.80 (114.40-251.70), 20.55 (14.95-28.44), 520.23 (390.25-647.15), and 246.94 (203.65-342.97)μg/g · cr, which were higher than those in control group (142.42 (96.87-179.11), 15.44 (12.26-20.98), 430.09 (309.85-568.78) and 213.85 (156.70-281.63) μg/g · cr, respectively) (Z values were-4.33,-5.04,-3.47 and-4.24, all P values<0.001). The urinary P, K level P50 (P25-P75) of exposed group were 582.50 (463.20-742.8), 890.10 (666.00-1 305.40) μg/g · cr, which were lower than control group (694.50 (546.20-851.17), 1 098.58 (904.53-1 479.18) μg/g · cr) (Z values were-3.36,-4.02, all P values <0.001). on Based the results of Pearson correlation analysis, urinary cadmium was positively correlated with urinary Ca, Cu, Zn, and I, and the correlation coefficients were 0.31, 0.61, 0.38, and 0.25, respectively(all P values<0.05). Based on the results of multiple regression analysis, urinary cadmium levels contributed most to the metabolic balance of urinary Ca, Cu, Zn and I. The standardized regression coefficients were 0.31, 0.59, 0.39, and 0.24, respectively (all P values<0.001). Conclusion Long?term environmental exposure to cadmium affected the metabolic balance of Ca, Cu, Zn and I in human body.