OBJECTIVE: To study the effect and regulatory mechanism of high-mobility group box 1 (HMGB1) on cellular proliferation during liver regeneration after partial hepatectomy (PH) in mice. METHODS: Using the classic mouse model of 70% PH,21 male BALB/c mice were randomly divided into the sham operation and the six PH groups (n=3). Mice in the sham operation group, the abdominal cavities were open surgically to expose but without removal of the liver,while mice in the PH group received 70% PH and were killed at 6 h,and at 1,2,3,5 and 7 days after the operation,respectively. In addition,ethyl pyruvate (EP) was used as the specific inhibitor of HMGB1 for intervention. In this experiment,9 male BALB/c mice were randomly divided into sham operation,PH and ethyl pyruvate (EP) intervention (n=3) groups. Mice in EP intervention group were intraperitoneally injected with EP at 40 mg/(kg·d) after 70% hepatectomy,and were killed 2 days after operation. Total protein and nuclear or cytoplasmic proteins were extracted from liver tissues of mice from each group,and the protein expression levels of HMGB1,TLR4,RAGE,NF-κB p65,Cyclin D1 and PCNA were detected by Western blot. RESULTS: Within one week after mice had PH,the protein expression level of HMGB1 in cellular nuclei reached the peak on the 2nd day after operation (P < 0.05),then decreased gradually. Changes of protein expression levels in the cytoplasm was consistent with that trend. In addition,the protein expression levels of TLR4,RAGE,NF-κB p65,Cyclin D1 and PCNA in the PH group were also significantly higher than those in the sham operation group (P < 0.05). After the intervention of EP,the expression levels of HMGB1 in the nuclear protein of liver tissues and the expression levels of TLR4 and RAGE in the total protein did not change significantly. However,the expression levels of HMGB1 in the cytoplasmic protein and the protein expression levels of NF-κB p65,Cyclin D1 and PCNA decreased significantly (P < 0.05). CONCLUSION: HMGB1 played a regulatory role in cellular proliferation during mice liver regeneration,and this protein may be used as a potential important intervention target for the regulation of liver regeneration.

OBJECTIVE: To explore the relationships between urine arsenic levels and hemoglobin A_1C(HbA_1C) in type 2 diabetic mellitus (T2DM) patients who were exposed to low levels of arsenic. METHODS: A case-series study involving 401 patients with T2DM and with exposure to arsenic in a district of Guangzhou were selected as the study subjects in 2018. The basic data of patients were obtained by questionnaire and physical examination,and the biological samples (the blood and urine samples) of patients were collected to detect blood glucose,blood lipid,HbA_1C and urine arsenic. According to the levels of arsenic in urine,the patients were divided into three groups. t-test,χ² test and analysis of variance (ANOVA) were used to compare the differences of indicators such as blood glucose,blood lipid,HbA_1C and urine arsenic among the groups. Multivariate logistic regression analyses were conducted to evaluate the influencing factors of HbA_1C in patients with T2DM. RESULTS: A total of 401 patients exposed to low-dose arsenic were included. The median of urine arsenic was 38.31 μg/L,and the interquartile range was 48.25 μg/L. There was a linear increasing trend of HbA_1C with changes of urine arsenic in different urine arsenic level groups,and the difference between the high and the low level groups was statistically significant (P < 0.05). With the increase of urine arsenic levels between the groups,the number of normal control group (HbA_1C < 7.0%) decreased gradually,the number of abnormal control group (HbA_1C ≥ 7.0%) increased gradually,the difference was statistically significant (P < 0.05). Multivariate regression analyses show that urine arsenic was the influencing factor of HbA_1C,OR(95%CI) of the high arsenic group was 2.077 (1.049,4.110),the difference of HbA_1C between the high and the low level groups was statistically significant (P < 0.05). CONCLUSION: Urine arsenic is related to the level of HbA_1C in patients with T2DM,which may be the influencing factor of HbA_1C,and its mechanism needs to be further explored.

OBJECTIVE: The aim of this study was to investigate whether 1,25-dihydroxyvitamin D₃[1,25-(OH)₂D₃] would inhibit oxidative damage which was induced by PM_2.5 in human bronchial epithelial cells (HBE). METHODS: HBE cells were divided into four groups:vehicle (ethanol,0.1%) control,1,25-(OH)₂D₃-treated group,ethanol + PM_2.5-treated group,and PM_2.5+1,25-(OH)₂D₃-treated group. HBE cells were pretreated with ethanol (0.1%) or 1×10^-9 mol/L of 1,25-(OH)₂D₃ for 24 h. Afterwards,the cell culture medium was replaced with medium containing PM_2.5 (200 μg/mL) and ethanol (0.1%) or PM_2.5 (200 μg/mL) and 1×10^-9 mol/L 1,25-(OH)₂D₃. After another 24 hours of incubation of the cultures,cells were harvested for determination of cell viability,MDA concentrations,ratios of reduced glutathione (GSH) to oxidized glutathione (GSSG),and expression levels of transcription factor NF-E2-related factor 2 (Nrf-2) and heme oxygenase-1 (HO-1) protein. These expressions were detected using CCK-8 assay kit,cell MDA assay kit,GSH/GSSG-Glo^TM assay kit,and Western blot,respectively. RESULTS: In the PM_2.5-treated groups,the cell viabilities were obviously decreased to 80.8% as compared with the control. After 48-h treatment of 1,25-(OH)₂D₃, the viability of PM_2.5-treated HBE cells was further reduced to 75.8%. After PM_2.5 exposure,MDA concentrations were significantly increased (P < 0.05),and the ratios of GSH/GSSG were found to be decreased significantly (P < 0.01) as compared with the control. After 48 h-treatment with 1,25-(OH)₂D₃, the anti-oxidative levels in the PM_2.5-exposed group were significantly improved (P < 0.05),together with decreased MDA concentrations and increased GSH/GSSG ratios. Protein analyses reveal that PM_2.5 could increase levels of Nrf-2 and HO-1 proteins in HBE cells. The 48 h-intervention of 1,25-(OH)₂D₃ could up-regulate VDR level and slightly increased levels of Nrf-2 and HO-1 in PM_2.5-treated HBE cells. CONCLUSIONS:PM_2.5 exposure induced oxidative damage in HBE cells,together with elevated lipid peroxidation,decreased GSH/GSSG ratios,and increased levels of antioxidant proteins including Nrf-2 and HO-1 proteins. However,1,25-(OH)₂D₃ exposure attenuated PM_2.5-induced oxidative stress,probably through regulation of Nrf-2/HO-1 pathway via VDR. Furthermore,1,25-(OH)₂D₃ exposure attenuated PM_2.5-induced oxidative stress,probably related to VDR and Nrf-2/HO-1 signal pathways. Finally,1,25-(OH)₂D₃ exposure decreased cell viability,probably attributed to cell cycle arrest,and PM_2.5-induced cell apoptosis which was promoted by 1,25-(OH)₂D₃.

OBJECTIVE: To investigate expression of heparanase (HPA),epithelial marker E-cadherin, interstitial marker N-cadherin and vimentin protein in human gastric cancers and their relationship with clinical-pathological indicators of human gastric cancer. METHODS: Expressions of HPA, E-cadherin, N-cadherin and vimentin proteins in 91 cases of human gastric cancers were detected by immunohistochemical methods. The relationships between the positive expression rates of these proteins and different pathological indicators of human gastric cancer were detected using chi-square test. Furthermore,the relationships between HPA and E-cadherin,HPA and N-cadherin,vimentin protein were analyzed using Spearman rank correlation. RESULTS: The positive expression rates of HPA,E-cadherin,N-cadherin and vimentin proteins in human gastric cancer tissues were 75.82%, 51.65%, 54.95% and 23.08%, respectively. Positive expression rates of HPA, E-cadherin,N-cadherin and vimentin proteins in 91 cases of human gastric cancer tissues related to the age,sex,and size of gastric cancer patients (P > 0.05),but were related to the location and degree of differentiation of human gastric cancer. It was related to the degree of lymph node metastasis and invasion (P < 0.05). Positive expression rates of HPA,N-cadherin and vimentin proteins in the gastric cardia were significantly higher than those in the gastric body,pyloric and antral gastric cancers. Poorly differentiated,lymph node metastatic and invasive gastric cancer were significantly higher than those in well differentiated,non-lymph node metastatic and initial gastric cancer. Positive expression rates of E-cadherin protein in pyloric and antral gastric cancer were significantly higher than those in cardiac and gastric body gastric cancers. Patients with high school differentiation and without lymph node metastasis were significantly higher than those with low differentiation and lymph node metastasis. Positive expression rates of E-cadherin protein in the depth of invasion were significantly lower than that in the initial site. The Spearman grade correlation analyses show that the positive expression rates of HPA were negatively correlated with that of E-cadherin protein in human gastric cancer tissues (r=0.341,P < 0.05),but positively correlated with the positive expression rates of N-cadherin (r=0.366, P < 0.05) and vimentin (r=0.284,P < 0.05). CONCLUSION: In human gastric cancers,the positive expression rates of HPA,N-cadherin and vimentin proteins were high in poorly differentiated,lymph node metastasis and invasive depth of gastric cancer,while the positive expressions of E-cadherin proteins were high in moderately differentiated gastric cancers, non-lymph node metastasis and initial sites. There was a positive correlation between HPA and N-cadherin proteins,and a positive correlation between HPA and vimentin proteins,and a negative correlation between HPA and E-cadherin protein expression. It can be seen that HPA protein was related to the increased expression of stromal marker N-cadherin and vimentin protein, and to the loss or decrease of epithelial marker E-cadherin protein, therefore HPA may induce epithelial-mesenchymal transformation in human gastric cancer tissue.

OBJECTIVE: To explore the induction of antioxidant and anti-inflammation effects of 2, 3, 5, 4'-tetrahydroxystilbene-2-O-β-D-glucoside (TSG) in LPS-stimulated RAW264.7 macrophages. METHODS: RAW264.7 cells were pretreated with TSG (30,60,120 μmol/L) for 4 hours,and then exposed to 1 μg/mL LPS for 12 hours. In addition,a blank control and two positive control groups were set up. For the positive controls,cells were treated with dexamethasone (100 nmol/L) or brusatol (50 μmol/L) at 4 h prior to LPS stimulation. Morphological changes were captured under a microscope,and the activated cell rate was analyzed. Concentrations of TNF-α and IL-6 in the culture medium were assessed using specific ELISA kits. Levels of superoxide anions and reactive oxygen species within the cells were detected using flow-cytometry after staining with MitoSOX^TM and DCFH-DA,respectively. Moreover,immunofluorescence staining and western-blot methods were used to investigate protein expression and sub-cellular localization of Nrf2 in the cells. Real-time PCR was used to detect expression of Nrf2-related antioxidant enzymes,including HO-1,SOD₂,SOD₁,CAT and GPX-1. RESULTS: Compared with the control group,LPS activated macrophages exhibited elongated cell morphology with pseudopodium-like protrusions. LPS also strongly promoted the level of pro-inflammatory cytokines,including TNF-α and IL-6. However,pretreatment with TSG inhibited LPS-induced ROS production and inflammatory cytokines secretion. Interestingly,the anti-inflammatory effect of TSG was better than Dex. TSG administration promoted Nrf2 protein expression and its translocated into the nucleus. Moreover,TSG alleviated LPS-induced oxidative stress through regulating Nrf2-related antioxidant enzymes,including HO-1,SOD₂ and CAT. Brusatol,a specific inhibitor of Nrf2,decreased TNF-α and IL-6 production in LPS-activated macrophages. CONCLUSION: TSG attenuated LPS-induced oxidative stress and inflammation in RAW264.7 macrophages through activation of Nrf2 signal pathway. Therefore,TSG may be useful as a drug to prevent or to treat inflammation-associated diseases.

OBJECTIVE: To assess the toxicity pathway activation induced by PM_2.5 using data from Tox21 10K library based on the result of Ambient fine particulate matter-bound PAHs during heating period of 2017 in Shijiazhuang. METHODS: Particle samples were collected in Hebei Medical University where it is away from polluted industrial area. Concentrations of 16 US EPA priority PAHs were measured by GC-EI-MS. The deposition fractions of PM-bound PAHs in adult alveolar region of lung were determined by multiple-path particle dosimetry model. The potencies of PAHs in the AhR,Nrf2,p53 and NF-κB pathways were calculated using data from the Tox21 10K library. The assessment of risk from PM_2.5 exposure was based on the alveolar deposition fraction combined with the potency of PAHs activated toxicity pathway. RESULTS: The average concentration of benzo[a]pyrene was 9.13 ng/m³. Daily concentrations of the top three chemicals in PM-bound PAHs were benzo[b]fluoranthene,fluoranthene and pyrene,and their average concentrations were 22.88、17.86 and 14.31 ng/m³ respectively. The benzo[a]pyrene equality concentration was 17.74 ng/m³. And the NF-κB pathway was most likely activated by PM-bound PAHs during the heating period and the second one was the AhR pathway which was followed by Nrf2. The p53 pathway was the least activated. The corresponding activity-to-exposure ratio values were 1.97,1.71,0.58 and 0.28. CONCLUSION: PM-bound PAHs in Shijiazhuang might increase the risk of carcinogenesis via activation of the NF-κB and the AhR pathways.

OBJECTIVES: To explore new autophagy related single nucleotide polymorphisms (SNPs) on the PI3K/Akt/mTOR pathway and their association with gastric carcinogenesis. METHODS: A 1:1 matched case-control study was conducted. SNP microarray was combined with KEGG pathway,Gene Ontology and Ensemble database to screen target SNPs. Matrix-assisted laser desorption/Ionization time of flight mass spectrometry (MALDI-TOF-MS) was used to detect SNP loci in 622 patients with gastric cancer and 622 healthy individuals from the Xianyou county,Fujian province. RESULTS: SNP chip and bioinformatics analyses screened IRS1 rs10205233,PIK3CD rs3934934,PIK3R1 rs706711,PIK3R1 rs706714 and AKT1 rs35285446 as candidate sites. Validations using expanded samples found that the polymorphism of IRS1 rs10205233 (C > T) significantly reduced the risk of gastric cancer[OR(95%CI),with co-dominant and dominant models of 0.761 (0.595,0.975) and 0.764 (0.601,0.973)],respectively. Further stratification analyses show that the dominant,invisible,codominant models and alleles of the site were not statistically different among cardia cancer and non-cardia cancer patients (P > 0.05). There was no correlation between the other 4 loci and the haplotype of PIK3R1 (rs706711,rs706714) with susceptibility to gastric cancer (P > 0.05). CONCLUSION: The IRS1 rs10205233 locus was associated with gastric carcinogenesis in the Xianyou county,a high-risk area of gastric cancer in Fujian province. The T allele may be a genetic protective factor for gastric cancer.

OBJECTIVE: To explore the expression and function of solute carrier family 7 member 11 (SLC7A11) in esophagus cancer (ESCA) and its prognostic value in patients. METHODS: Differences in expression of SLC7A11 in ESCA tissues and normal esophageal tissues were analyzed using the Oncomine and GEPIA databases. GeneMANIA database was used to construct the protein-protein interaction network (PPI) which is related to SLC7A11, and functional annotation was carried out to analyze its common enrichment functions. KEGG Pathway database was used to analyze the signaling pathways involved. Kaplan-Meier curve was drawn using the R2 gene analysis visualization platform to study the association of SLC7A11 expression and overall survival (OS) of ESCA patients. RESULTS: Based on the Oncomine and GEPIA databases,the mRNA expression levels of SLC7A11 in ESCA tissues were significantly higher than that in normal tissues,the prognosis of patients in the high-expression group was worse,and their differences were statistically significant (P < 0.05). SLC7A11 interacts with 20 proteins including SLC3A2, BSG, ASNS, PHGDH, TERT and CD44, and they were enriched in amino acid transport, modified amino acid transmembrane transporter activity, leukocyte migration,peptide antigen binding and other biological functions. SLC7A11 was involved in regulation of the ferroptosis (hsa04216) signaling pathway. CONCLUSION: SLC7A11 was up-regulated in ESCA significantly, indicative of poor prognosis, involved in ferroptosis through the P53 and ROS metabolic pathways. Therefore,SLC7A11 can be considered to be biomarkers for early diagnosis,treatment and prognosis of patients with ESCA.

OBJECTIVE: To investigate the effect and molecular mechanism of mitochondrial citrate transporter SLC25A1 on sensitivity of esophageal squamous cell carcinoma to radiotherapy in vitro. METHODS: The Cancer Genome Atlas (TCGA) tumor tissue database and UALCAN interactive network tool were used to compare differences in SLC25A1 expression between 184 cases of esophageal cancer and 11 cases of normal tissues. Radioresistant esophageal carcinoma cell line and the control were established by multiple round of X-ray sublethal dose exposures. Radioresistance and radiosensitivities were characterized by the radiation clonal survival assay and the linear quadratic target model. Morphologic changes of cell apoptosis were observed by fluorescence microscopy using the Hoechst 33258 staining after multiple X-ray exposures at a total dose of 6.0 Gy. Flow cytometry was used to detect levels of reactive oxygen species after the same X-ray exposures. Western blotting was used to analyze the expression alteration of levels of phosphor-H2AX and phosphor-DNA-PKcs,and the key signal kinases in DNA repair by SLC25A1 knockdown with lentivirus vector carrying small hairpin interfering RNA. RESULTS: A comparative analysis of TCGA tumor tissue database confirmed that the expression level of SLC25A1 mRNA in esophageal cancer tissues was 1.65-folds higher than that in normal esophageal tissues (P=1.64×10^-4). The expression level of SLC25A1 protein was increased by 2.36-folds when compared with the control (P < 0.05). The apoptosis rate of esophageal cancer cell lines with stable SLC25A1 expression compared to the control was increased by 1.58-folds (P < 0.05). ROS level in the SLC25A1 knockdown radioresistant esophageal carcinoma cell line was decreased by (65.3±14.3)% (P=0.007). Levels of phosphorylation of H2AX (Ser139) and DNA-PKCs (Ser2056) in SLC25A1 knockdown radioresistant esophageal carcinoma cell lines were significantly reduced when compared with the control (all P < 0.05). CONCLUSION: SLC25A1 may be an oncogene involved in tumor pathogenesis and radiation resistance. The data suggest that it can inhibit DNA damage repair and induce cell apoptosis by down-regulating reactive oxygen species,and can serve as a potential molecular target for radiation sensitization of esophageal squamous cell carcinoma.

OBJECTIVE: To investigate the effects of low-level lead exposure on glucose tolerance and retinal vascular permeability in mice and explore the effects of lead on diabetic retinopathy. METHODS: A lead exposure mouse model was initiated beginning with the mother generation:four groups exposed to 0,10,100,500 mg/L lead acetate. Offspring were exposed continuously until the sacrifice time. Half of mice were sacrificed at 8 weeks age,named the 8-weeks group with 12 mice (male:female:1:1) in each exposure concentration level. The graphite furnace flame atomic absorption spectrometry was used to detect lead in blood,and the intraperitoneal glucose tolerance test was used to measure glucose tolerance. The fundus vein fluorescence angiography and retinal fundus imaging technology were used to detect the vascular permeability of retinal vessels. Real-time fluorescence quantitative polymerase chain reaction (qPCR) was used to detect transcription of tight junction-related molecules. The other half of mice were sacrificed at 20 weeks age,named the 20-week group. RESULTS: The blood lead concentrations of offspring increased with the increment of lead acetate exposure level(r_{8 week}=0.887,P < 0.05;r_{20 week}=0.972,P < 0.05). The blood lead levels were higher in the 8-week than the 20-week groups when the lead acetate exposure levels were 0,10,100 mg/L. However,the 20-week group had higher blood lead than the 8-week group when the exposure was at the 500 mg/L lead acetate exposure level (P < 0.01). Compared with the control group,the blood glucose of the exposed offspring was higher within 2 hours after intraperitoneal injection of glucose,and the area under the curve of blood glucose was higher than the control mice (P < 0.05 or P < 0.01). In the 8-week group,the blood glucose and AUC of mice exposed to 10 mg/L lead acetate showed the most significantly changes compare to the controls while those at the 20-week group was at 100 mg/L. In the 8-week group,there was no vascular leakage,but only at the molecular level,the transcriptional levels of Zona occludens protein-1(ZO-1),Occludin,Claudin-1,3,5 were increased initially and then inhibited compared with control group (P < 0.05 or P < 0.01). In the 20-week group,leakages occurred in the retinal vessels of offspring exposed to lead acetate in the 100 and 500 mg/L groups,while tight junction related molecules ZO-1,Occludin,Claudin-1,3,5 transcriptions were inhibited (P < 0.01). CONCLUSION: Lead exposure increased blood glucose in the glucose tolerance test and the dose was very low before adulthood. Persistent lead exposure directly caused leakage in the retinal blood vessels which may be a potential risk factor for diseases such as diabetic retinopathy.

OBJECTIVE: In view of the low efficiency and easy to precipitate in the determination of human serum cholinesterase (ChEs) by the hydroxylamine ferric chloride method (WS/T66-1996),the method was optimized to make the determination more efficient. METHODS: The determination of AChE and BChE by the hydroxylamine ferric chloride method was verified. On this basis,the determination of BChE by the microporous plate hydroxylamine ferric chloride method was further studied. Finally,taking the improved Ellman method as a reference,the accuracy and reliability before and after optimization were verified. RESULTS: The hydroxylamine ferric chloride method for determination of AChE enzyme activities in human serum was poor,while it was ideal for determination of BChE enzyme activities. There was a positive correlation between the results of BChE enzymatic activities as determined by the hydroxylamine ferric chloride method and the improved Ellman method (r=0.652,P < 0.01). In addition,there was also a positive correlation between the results of BChE enzymatic activities as determined by the microporous plate hydroxylamine ferric chloride method and the hydroxylamine ferric chloride method (r_s=0.653,P < 0.01). Compared with the hydroxylamine ferric chloride method,the accuracy and reliability of the microporous plate hydroxylamine ferric chloride method was slightly better,mainly reflected in the lowest detection concentration and the coefficient of variation. CONCLUSION: The microporous plate hydroxylamine ferric chloride method is a high flux and economical method for the determination of BChE enzymatic activity in human serum.