The relationship between NTCP gene varieties and the progress of liver disease after HBV infection: an updated systematic review and meta-analysis.

BACKGROUND: The aim of this study was to analyze the relationship between sodium taurocholate cotransporting polypeptide (NTCP) gene varieties and hepatitis B virus (HBV) infection and the progress of HBV-related liver disease. METHODS: PubMed, EMBASE, Web of Science and Cochrane library were used to search eligible studies. STATA software was performed to combine results. Pooled odds ratios (OR) was used to assess the potential genetic relationships. RESULTS: A total of 18 eligible case-control studies with 24960 cases and 28342 controls were included in this meta-analysis. The A allele of rs2296651 polymorphism was found to be significantly linked to a protection of HBV infection in the whole combined analysis (P = 0.000). Meanwhile, this allele was significantly associated with a decreased risk of hepatocellular carcinoma (HCC) (A vs. G: OR = 0.668, 95% CI: 0.571-0.782, P = 0.000), and was significantly associated with HBV nature clearance (A vs. G: OR = 0.744, 95% CI: 0.585-0.946, P = 0.016; AA+GA vs. GG: OR = 0.775, 95% CI: 0.613-0.980, P = 0.033; GA vs. GG: OR = 0.748, 95% CI: 0.588-0.952, P = 0.018). However, rs4646287 genetic varieties had no statistical differences in all models with HBV infection or HBV-related disease progress, liver cirrhosis, acute-on-chronic liver failure and HCC, as well as rs7154439, rs4646285, rs4646296. CONCLUSIONS: Rs2296651 polymorphism (A allele) may protect from HBV infection and the progress of HBV-related disease (HBV-related HCC). Future research about other single nucleotide polymorphisms (SNPs) (rs4646287, rs7154439, rs4646285, rs4646296) of NTCP may be needed to clarify the relationship of NTCP gene varieties with HBV infection and HBV-related disease.

THRSP identified as a potential hepatocellular carcinoma marker by integrated bioinformatics analysis and experimental validation.

Hepatocellular carcinoma (HCC) is the most common malignant liver tumor with high mortality and poor prognosis worldwide. This study aimed to identify hub genes and investigate the underlying molecular mechanisms in HCC progression by integrated bioinformatics analysis and experimental validation. Based on the Gene Expression Omnibus (GEO) databases and The Cancer Genome Atlas (TCGA), 12 critical differential co-expression genes were identified between tumor and normal tissues. Via survival analysis, we found higher expression of LCAT, ACSM3, IGF1, SRD5A2, THRSP and ACADS was associated with better prognoses in HCC patients. Among which, THRSP was selected for the next investigations. We found that THRSP mRNA expression was negatively correlated with its methylation and closely associated with clinical characteristics in HCC patients. Moreover, THRSP expression had a negative correlation with the infiltration levels of several immune cells (e.g., B cells and CD4+ T cells). qRT-PCR verified that THRSP was lower expressed in HCC tissues and cell lines compared with control. Silencing of THRSP promoted the migration, invasion, proliferation, and inhibited cell apoptosis of HCCLM and Huh7 cell lines. Decreased expression of THRSP promoted HCC progression by NF-κB, ERK1/2, and p38 MAPK signaling pathways. In conclusion, THRSP might serve as a novel biomarker and therapeutic target of HCC.

The Biomarkers for Predicting Viral HepatitisAssociated Hepatocellular Carcinoma.

BACKGROUND: Hepatocellular carcinoma (HCC) is the fifth most common cancer in the world, and more than half of the newly diagnosed cases are chronic hepatitis B patients. Due to the lack of specific clinical manifestations, many patients are already at an advanced stage at the time of diagnosis and therefore have missed the best time for treatment. Organs in a pathological state usually secrete specific substances into the blood, which can indirectly indicate the pathological state of the organ, so some biological markers in the blood can be used as a tool to predict the incidence of HCC. METHODS: The Research articles related to HCC were collected by searching PubMed databases with the keywords "hepatocellular carcinoma", "serum biomarker", "hepatitis B", "prediction" and "prognosis", and Additional articles were identified by manual search of references found in the primary articles, followed by a summary and review. RESULTS: Viral hepatitis is the main cause of HCC worldwide, and this phenomenon is particularly prominent in Asian and African populations. A variety of serological markers including M2BPGi, IL-6 and COMP can be used to predict the incidence of long-term HCC in patients. The risk of HCC is dynamic rather than constant, and dynamic detection will help improve prediction accuracy. For hepatitis B patients, HBV DNA load and HBcr Ag are important predictive markers of HCC. CONCLUSION: For a high-risk population of HCC, early risk prediction is helpful to guide clinical work, and timely adjustments of the screening frequency and treatment plan are helpful to prolong the survival time of HCC patients.

AC093797.1 as a Potential Biomarker to Indicate the Prognosis of Hepatocellular Carcinoma and Inhibits Cell Proliferation, Invasion, and Migration by Reprogramming Cell Metabolism and Extracellular Matrix Dynamics.

Purpose: The risk signature composed of four lncRNA (AC093797.1, POLR2J4, AL121748.1, and AL162231.4.) can be used to predict the overall survival (OS) of patients with hepatocellular carcinoma (HCC). However, the clinical significance and biological function of AC093797.1 are still unexplored in HCC or other malignant tumors. In this study, we aimed to investigate the biological function of AC093797.1 in HCC and screen the candidate hub genes and pathways related to hepatocarcinogenesis. Methods: RT-qPCR was employed to detect AC093797.1 in HCC tissues and cell lines. The role of AC093797.1 in HCC was evaluated via the cell-counting kit-8, transwell, and wound healing assays. The effects of AC093797.1 on tumor growth in vivo were clarified by nude mice tumor formation experiments. Then, RNA-sequencing and bioinformatics analysis based on subcutaneous tumor tissue was performed to identify the hub genes and pathways associated with HCC. Results: The expression of AC093797.1 decreased in HCC tissues and cell lines, and patients with low expressed AC093797.1 had poor overall survival (OS). AC093797.1 overexpression impeded HCC cell proliferation, invasion, and migration in vitro and suppressed tumor growth in vivo. Compared with the control group, 710 differentially expressed genes (243 upregulated genes and 467 downregulated genes) were filtered via RNA-sequencing, which mainly enriched in amino acid metabolism, extracellular matrix structure constituents, cell adhesion molecules cams, signaling to Ras, and signaling to ERKs. Conclusion: AC093797.1 may inhibit cell proliferation, invasion, and migration in HCC by reprograming cell metabolism or regulating several pathways, suggesting that AC093797.1 might be a potential therapeutic and prognostic marker for HCC patients.

Enhanced Efficacy of Direct-Acting Antivirals in Hepatitis C Patients by Coadministration of Black Cumin and Ascorbate as Antioxidant Adjuvants.

The widespread adaptation of a new generation of direct-acting antiviral agents (DAAs) unveils a superlative effect in the eradication of the hepatitis C virus (HCV). However, this therapy has been reported to exhibit vigorous side effects that pose a risk in fleet recovery. This study was conducted to investigate the efficacy of DAAs: sofosbuvir (SOF) and ribavirin (RBV), along with black cumin (BLC) and ascorbate (ASC), as adjuvants on hematological parameters; oxidative stress markers such as total antioxidant status (TAS), superoxide dismutase (SOD), reduced (GSH) and oxidized (GSSG) glutathione (GSH), gamma-glutamyl transferase (GGT), and malondialdehyde (MDA); liver function markers such as aspartate transaminase (AST), alanine aminotransferase (ALT), bilirubin, and alkaline phosphatase (ALP); and viral load with determined genotypes. HCV-infected patients (n = 30) were randomly divided into two equal groups: control group (n = 15) and treatment group (n = 15). The control group was subjected only to SOF and RBV (400 mg each/day). Synergistically, the treatment group was administered with adjuvant therapy of BLC (250 mg/day) and ASC (1000 mg/day) along with DAAs (400 mg each/day) for 8 weeks. All selected patients were subjected to sampling at pre- and posttreatment stages for the assessment of defined parameters. The data revealed that the BLC/ASC adjuvant therapy boosted the efficacy of DAAs by reducing the elevated levels of liver markers such as AST, ALT, ALP, and bilirubin in the treatment group compared with those in the control group (P > 0.05). The adjuvant therapy synchronously showed an ameliorating effect on hematological parameters. The SOF/RBV with adjuvant therapy also demonstrated an increasing effect in the activity of SOD, TAS, and GSH and a decreasing effect for GSSG, GGT, and malondialdehyde (MDA; P > 0.05) followed by curtailing a RT-PCR-quantified viral load. Our findings provide evidence that systemic administration of BLC/ASC efficiently alleviates hematological, serological, and antioxidant markers as well as the viral load in hepatitis C patients. This highlights a potentially novel role of BLC and ASC in palliating hepatitis C.

Identification of a novel four-lncRNA signature as a prognostic indicator in cirrhotic hepatocellular carcinoma.

BACKGROUND: Many studies have shown that long noncoding RNAs (lncRNA) are closely associated with the occurrence and development of various tumors and have the potential to be prognostic markers. Moreover, cirrhosis is an important prognostic risk factors in patients with liver cancer. Some studies have reported that lncRNA-related prognostic models have been used to predict overall survival (OS) and recurrence-free survival (RFS) in patients with hepatocellular carcinoma (HCC). However, no one has constructed a prognostic lncRNA model only in patients with cirrhotic HCC. Thus, it is necessary to screen novel potential lncRNA markers for improve the prognosis of cirrhotic HCC patients. METHODS: The probe expression profile dataset (GSE14520-GPL3921) from the Gene Expression Omnibus (GEO), which included 204 cirrhotic HCC samples, was reannotated and the lncRNA and mRNA expression dataset was obtained. The patients were randomly assigned to either the training set (n = 103) and testing set (n = 100). Univariate cox regression and the least absolute shrinkage and selection operator (LASSO) model were applied to screen lncRNAs linked to the OS of cirrhotic HCC in the training set. The lncRNAs having significant correlation with OS were then selected and the multivariate Cox regression model was implemented to construct the prognostic score model. Whether or not this model was related to RFS in the training set was simultaneously determined. The testing set was used to validate the lncRNA risk score model. A risk score based on the lncRNA signature was used for stratified analysis of different clinical features to test their prognostic performance. The prognostic lncRNA-related protein genes were identified by the co-expression matrix of lncRNA-mRNA, and the function of these lncRNAs was predicted through the enrichment of these co-expression genes. RESULTS: The signature consisted of four lncRNAs:AC093797.1,POLR2J4,AL121748.1 and AL162231.4. The risk model was closely correlated with the OS of cirrhotic HCC in the training cohort, with a hazard ratio (HR) of 3.650 (95% CI [1.761-7.566]) and log-rank P value of 0.0002. Moreover, this model also showed favorable prognostic significance for RFS in the training set (HR: 2.392, 95% CI [1.374-4.164], log-rank P = 0.0015). The predictive performance of the four-lncRNA model for OS and RFS was verified in the testing set. Furthermore, the results of stratified analysis revealed that the four-lncRNA model was an independent factor in the prediction of OS and RFS of patients with clinical characteristics such as TNM (Tumor, Node, Metastasis system) stages I-II, Barcelona Clinic Liver Cancer (BCLC) stages 0-A, and solitary tumors in both the training set and testing set. The results of functional prediction showed that four lncRNAs may be potentially involve in multiple metabolic processes, such as amino acid, lipid, and glucose metabolism in cirrhotic HCC.

Association of IL-6, IL-10 and TGF-ß1 gene polymorphisms with brucellosis: A systematic review with meta-analysis.

BACKGROUND: Brucellosis is one of the major public health problems worldwide. Several current studies have provided data that polymorphisms in the interleukin-6 (IL-6), interleukin-10 (IL-10) and transforming growth factor beta1(TGF-ß1) gene were associated with the susceptibility to human brucellosis, but the results remain inconsistent. OBJECTIVES: The aim of present study was to investigate the relationship between IL-6 (-174 G/C), IL-10 (-1082 A/G, -819C/T) and TGF-ß1 (codon 10, codon 25) gene polymorphisms and brucellosis. METHODS: We performed a comprehensive search of the PubMed, EMBASE, Web of Science, OVID-EBMR, and the Cochrane Library up to Oct. 30, 2018. The search was designed using the following key words: "brucellosis" or" "brucella melitensis", "IL-10" or "interleukin10" or "interleukin-10", "IL-6" or "interleukin6" or "interleukin-6", "TGF-ß1" or "TGF-beta1" or "transforming growth factor ß1", "polymorphism" and "single nucleotide polymorphism (SNP)". Crude odds ratios (ORs) with 95% confidence intervals (CIs) were used to measure the strength of association between TGF-ß1, IL-10 and IL-6 polymorphisms and brucellosis risk. All the statistical analyses were conducted by Review manager 5.3 software. RESULTS: A total of 8 studies involving 1308 cases and 902 controls met the inclusion criteria for IL-6, IL-10, TGF-ß1 polymorphisms and brucellosis risk. There was a slightly trend of increasing risk of brucellosis in individuals with the G allele compared with individuals with the C allele (OR = 1.07, 95% CI: 0.85-1.33, P = 0.57) in IL-6 polymorphism. However, statistical analysis showed that these differences are not significant. Our results suggested TGF-ß1 (codon 25 G/C) GG genotype may be considered as a risk factor for brucellosis (OR = 1.67, 95% CI: 1.12-2.50, P = 0.01). Herein, we failed to find any significant association between IL-10 (-1082 A/G, -819C/T), TGF-ß1 (codon 10C/T) gene polymorphism and susceptibility to brucellosis in all gene models. CONCLUSION: IL-6 (-174 G/C), IL-10 (-1082 A/G, -819C/T), and TGF-ß1 (codon 10C/T) polymorphisms is not a risk factor for brucellosis infection. TGF-ß1 codon 25 GG genotype may be considered as a risk factor for brucellosis.

Association of STAT3 and STAT4 polymorphisms with susceptibility to chronic hepatitis B virus infection and risk of hepatocellular carcinoma: a meta-analysis.

Background: It has been reported that polymorphisms of signal transducer and activator of transcription (STAT) 3 and STAT4 might be associated with susceptibility to hepatitis B virus (HBV) infection and risk of chronic hepatocellular carcinoma (HCC). Owing to limitation of sample size and inconclusive results, we conducted a meta-analysis to clarify the association. Methods: We identified relevant studies by a systematic search of Medline/PubMed, Embase, Web of Science and the Cochrane Library up to 20 February 2019. The strength of the association measured by odds ratios (OR) with 95% confidence intervals (CIs) was studied. All the statistical analyses were conducted based on Review Manager 5.3 software. Results: A total of 5242 cases and 2717 controls from five studies were included for the STAT3 polymorphism, 5902 cases and 7867 controls from nine studies for the STAT4 polymorphism. Our results suggested that STAT3 rs1053004 polymorphism was a significant risk factor of chronic HBV infection (C vs. T: OR = 1.17, 95% CI: 1.07-1.29, PA=0.0007; CC + CT vs. TT: OR = 1.38, 95% CI: 1.09-1.76, PA=0.008). Validation with all the genetic models revealed that rs7574865 polymorphism of STAT4 gene was closely associated with chronic HBV infection (PA<0.01) and chronic hepatitis B (CHB)-related HCC (PA<0.05). Meanwhile, the authenticity of the above meta-analysis results was confirmed by trial sequential analysis (TSA). Conclusions: The meta-analysis showed that STAT3 rs1053004 polymorphism may be the risk for developing chronic HBV infection but not associated with HCC. The present study also indicates that STAT4 rs7574865 polymorphism increased the risk of chronic HBV infection and HCC.

iTRAQ­based proteomic analysis of endotoxin tolerance induced by lipopolysaccharide.

The purpose of the present study was to investigate the differentially expressed proteins between endotoxin tolerance and sepsis. Cell models of an endotoxin tolerance group (ET group) and sepsis group [lipopolysaccharide (LPS) group] were established using LPS and evaluated using ELISA and flow cytometry methods. Differentially expressed proteins between the ET and the LPS groups were identified using isobaric tags for relative and absolute quantitation (iTRAQ) analysis and evaluated by bioinformatics analysis. The expression of core proteins was detected by western blotting. It was identified that the expression of tumor necrosis factor­α and interleukin­6 was significantly decreased in the ET group compared with the LPS group. Following high­dose LPS stimulation for 24 h, the positive rate of cluster of differentiation­16/32 in the ET group (79.07%) was lower when compared with that of the LPS group (94.27%; P<0.05). A total of 235 proteins were identified by iTRAQ, and 36 upregulated proteins with >1.2­fold differences and 27 downregulated proteins with <0.833­fold differences were detected between the ET and LPS groups. Furthermore, the expression of high mobility group (HMG)­A1 and HMGA2 in the ET group was higher compared with the LPS group following high­dose LPS stimulation for 4 h, while HMGB1 and HMGB2 exhibited the opposite expression trend under the same conditions. In conclusion, proteomics analysis using iTRAQ technology contributes to a deeper understanding of ET mechanisms. HMGA1, HMGA2, HMGB1 and HMGB2 may serve a crucial role in the development of ET.

Baseline Characteristics and Treatment Patterns of the Patients Recruited to the China Registry of Hepatitis B.

Background and Aims: Chronic hepatitis B virus (HBV) infection remains a major public health problem globally. Here, we describe the baseline characteristics and treatment profiles of HBV-infected patients recruited to the China Registry of Hepatitis B. Methods: Inclusion criteria were patients with different stages of chronic HBV infection and complete key data. Exclusion criteria were patients with hepatocellular carcinoma. The baseline clinical, laboratory and treatment profiles were analyzed. Results: Finally, 40,431 patients were included. The median age was 43 years, with 65.2% being men and 51.3% being positive for hepatitis B e antigen (HBeAg). The most common initial diagnosis was chronic hepatitis B (81.0%), followed by cirrhosis (9.3%), inactive carrier of hepatitis B surface antigen (HBsAg) (6.7%), and immune tolerant phase of hepatitis B infection (3.0%). Among the 21,228 patients who were on treatment, 88.0%, 10.0% and 2.0% received nucleos(t)ide analogues (NAs), interferon or combination of NAs and interferon, respectively. The proportion of patients who received preferred NAs (entecavir or tenofovir disoproxil fumarate) had increased from 13.5% in 2003 to 79.7% in 2016. Conclusions: We concluded that middle-aged men accounted for most of the patients with chronic hepatitis B in this cross-sectional study. About half of the patients were HBeAg-positive. NAs were the most commonly used therapy, and use of the preferred NAs had steadily increased in the past decade.

Association of GSTP1 and P16 promoter methylation with the risk of HBV-related hepatocellular carcinoma: a meta-analysis.

BACKGROUND: Study on the relationship between glutathione-S-transferase Pi 1 (GSTP1) and P16 promoter region methylation and the risk of hepatitis B virus-related hepatocellular carcinoma (HBV-related HCC) has produced inconsistent results. OBJECTIVES: To assess the correlation between GSTP1 and P16 promoter methylation frequency and HBV-related HCC susceptibility. METHODS: All relevant studies were identified by searching PubMed, Embase, Web of Science, and China National Knowledge Infrastructure literature databases before December, 2017. The OR and the corresponding 95% CI were calculated to investigate the risk of GSTP1 and P16 promoter methylation rate and HBV-related HCC. Sensitivity analysis was performed and publication bias was estimated using the Begg's and Egger's test. RESULTS: Our meta-analysis identified the relationships of GSTP1 (six studies including 213 HBV-related HCC tumor tissues) and P16 (nine studies with 287 HBV-related HCC tumor tissue) promoter methylation with HCC risk. Compared with normal liver tissue and cirrhosis, the pooled ORs of GSTP1 promoter region methylation in HBV-related HCC cancer tissues were 6.05 (95% CI =1.20-30.52) and 5.21 (95% CI =2.19-12.41), respectively. Compared with paracancerous tissue, normal liver tissue, cirrhosis, and chronic hepatitis B as controls, the pooled ORs of P16 promoter region methylation in HBV-related HCC cancer tissues were 7.18 (95% CI =2.31-22.33), 24.89 (95% CI =3.38-183.03), 5.92 (95% CI =1.78-19.68), and 12.12 (95% CI =0.75-196.50). CONCLUSION: In summary, our meta-analysis found strong associations between GSTP1 and P16 gene promoter methylation and an increased HBV-related HCC susceptibility. Moreover, GSTP1 and P16 methylation in promoter region could obviously increase the risk of HBV-related HCC in patients with cirrhosis, indicating that these would be promising biomarkers for early clinical diagnosis of HBV-related HCC.

[Effects of lingqijuangan capsule medicated serum on apoptosis of activated rat hepatic stellate cells].

OBJECTIVE: To investigate the effects of LingQiJuanGan Capsule medicated serum on the apoptosis of and the expression of Bcl-2/Bax in hepatic stellate cell-T6 (HSC-T6) activated by platelet-derived growth factor (PDGF). METHODS: Preparation of drug-medicated serum. 30 SD rats were randomly divided into three groups. Serum was obtained from the SD rats administrated intragastricly with saline (10 mL/kg, group A), Fufang Biejia Ruangan Tablet solution (1.5 g/kg, group B) and LingQiJuanGan Capsule solution respectively (4.25 g/kg, group C). Detection of cell apoptosis and the expression of Bcl-2/Bax. HSC-T6 cells were activated by 10 ng/mL PDGF and incubated with drug-medicated serum at 200 mL/L. Cell apoptosis and the expression of Bcl-2/Bax were measured by flow cytometry at 0, 12, 24 and 48 h. RESULTS: Significantly higher apoptotic rates of HSC-T6 in group B and group C were detected at 12, 24, 48 h compared with that of group A (P < 0.05), wheraus no significant difference was found between group B and group C (P > 0.05). With increase of exposure time, the apoptotic rates of HSC-T6 in group B and group C increased (P < 0.05). The levels of Bcl-2 expression in HSC-T6 in group B and C were lower than that of group A (P < 0.05) at 12, 24, 48 h (No significant difference between group B and group C, P > 0.05). The expression level of Bcl-2 decreased with increased exposure time (P > 0.05) in both group B and group C. However, no such trend was observed in group A (P > 0.05). In both group B and group C, the expression level of Bax increased in a time-dependent manner. CONCLUSION: LingQiJuanGan Capsule can induce apoptosis of PDGF activated HSC-T6 and inhibit hepatic fibrosis in a time-dependent manner by influencing the expression of Bcl-2/Bax.

Involvement of the nuclear factor-κB pathway in the adhesion of neutrophils to renal tubular cells after injury induced by neonatal postasphyxial serum.

Nuclear factor κB (NF-κB) plays an important role in the regulation of inflammatory proteins. However, it is unclear whether the NF-κB/intercellular adhesion molecule-1 (ICAM-1) pathway is involved in the adhesion of neutrophils and renal injury after hypoxia-ischemia (HI) in neonates. In this report we investigated whether NF-κB and its downstream molecule ICAM-1 were involved in renal injury induced by postasphyxial serum (PS) from neonates. Human renal proximal tubular (HK-2) cells were preincubated with 10 % fetal calf serum (control), 20 % neonatal PS, or 20 % PS plus pyrolidine dithiocarbamate (PDTC). The expression of IκBα, NF-κB p65, and ICAM-1 in HK-2 cells was determined by Western blot and/or immunohistochemistry. Nuclear translocation of NF-κB p65 in HK-2 cells was detected by immunofluorescence and Western blot. The ICAM-1 mRNA was determined by RT-PCR. Then HK-2 cells were cultured with neutrophils from neonates with asphyxia. After HK-2 cells had been cultured with neutrophils, we detected myeloperoxidase (MPO) activity, the leakage rate of lactate dehydrogenase (LDH), and cell viability. We found that PS preincubation resulted in significantly decreased IκBα expression and increased expression of NF-κB and ICAM-1, and facilitated the nuclear translocation of NF-κB in HK-2 cells. PS preincubation increased MPO activity, leading to elevated leakage rates of LDH and decreased cell viability after neutrophil exposure. Furthermore, the inhibition of NF-κB activity by PDTC significantly upregulated IκBα expression, decreased NF-κB and ICAM-1 expression, downregulated the nuclear translocation of NF-κB, and decreased MPO activity. This leads to decreased leakage rates of LDH and increased cell viability after neutrophil exposure. Our findings suggest that NF-κB/ICAM-1 pathway may be involved in neutrophil-endothelial interactions and neonatal renal injury after HI.

[Influence of diazoxide on mitochondrial apoptosis in human renal tubular cells induced by serum obtained from neonates with asphyxia].

OBJECTIVE: To study the influence of diazoxide on mitochondrial apoptosis pathway of human renal proximal tubular cells (HK-2 cells). METHODS: Cultured HK-2 cells were inoculated on 6-well plates, according to stochastic tables law, and they were divided into normal serum-treated group (NSTG) , post-asphyxial serum treatment group (PSTG), and post-asphyxial serum and diazoxide treatment group (PSDTG). The serum from neonates 24 hours after asphyxia in a dilution of 20% (volume fraction) was used for challenge. Diazoxide in a final concentration of 100 mol/L, was used for intervention. The expression of caspase-3 was detected by immunohistochemical method. The translocation rate of Omi/HtrA2 and mitochondria membrane potential were determined by indirect immunofluorescence and confocal microscopy. RESULTS: Compared with that of NSTG, the expression of caspase-3 absorbance (A) value of HK-2 cells in PSTG was significantly increased (25.19 + or - 3.33 vs. 13.63 + or - 1.89, P<0.01), the translocation rate of Omi/HtrA2 of HK-2 cells in PSTG was significantly increased [(56.01 + or - 5.30)% vs.(37.59 + or - 5.60)%, P<0.01], mitochondrial membrane red/green fluorescence intensity ratio was decreased significantly (0.79 + or - 1.42 vs. 1.82 + or - 0.23, P<0.01). Compared with the PSTG, the expression value of caspase-3 of HK-2 cells in PSDTG was significantly decreased (20.17 + or - 2.19), the translocation rate of Omi/HtrA2 of HK-2 cells in PSDTG was significantly decreased [(46.91 + or - 2.70)%], and mitochondrial membrane red/green fluorescence intensity ratio increased significantly (1.47 + or - 0.14), but did not recover to the same degree as that of the NSTG (all P<0.01). CONCLUSION: The diazoxide may reduce the expression of caspase-3, intracellular translocation of Omi/HtrA2, and stability of mitochondrial membrane potential, thereby significantly alleviates HK-2 cells injury induced by post-asphyxial-serum of neonate.

[Role of Omi/HtrA2 in renal tubular cells apoptosis induced by post asphyxial serum of neonate].

OBJECTIVE: To investigate the mechanism of inducing apoptosis of Omi/HtrA2 in renal tubular cells with post asphyxial serum of neonate. METHODS: Human renal proximal tubular cell line HK-2 cell was used as target cell. They were divided into three groups: control group, asphyxia group and Ucf-101 (Omi/HtrA2 special inhibitor) treated group. The challenge concentration of serum obtained from neonates 24 hours after asphyxia was 20%, and the treatment concentration of Ucf-101 was 10 mumol/L. The Omi/HtrA2 translocation in renal tubular cells was observed with confocal microscopy, and the rate of apoptosis was detected with flow cytometer. RESULTS: It was found that Omi/HtrA2 was translocated into cytoplasm in asphyxia group, and the rate of Omi/HtrA2 translocation in HK-2 cells of asphyxia group was significantly increased [(28.1+/-3.6)% vs. (9.4+/-2.1)%, P<0.01]. Compared with the control group, after being treated with post asphyxial serum, the rate of apoptosis of HK-2 cells in asphyxia group was significantly increased [(36.3+/-4.4)% vs.(12.4+/-2.9)%, P<0.01]. Compared with asphyxia group, the rate of apoptosis in HK-2 cells in Ucf-101 treated group was significantly decreased [(27.0+/-3.9)% vs.(36.3+/-4.4)%, P<0.01]. CONCLUSION: These experimental data demonstrates that post asphyxial serum of neonate can induce apoptosis of HK-2 cells, and translocation of Omi/HtrA2 from mitochondria into cytoplasm may play an important role in its intracellular signal transduction mechanism in induction of apoptosis.

[Role and mechanism of Salvia miltiorrhiza Bunge in alleviating injury induced by hypoxia/reoxygenation in HK-2 cells].

OBJECTIVE: Salvia miltorrhiza Bunge (SMB) is a traditional Chinese herb, which is considered to promote blood flow and remove blood stasis. This study examined whether SMB can alleviate injury induced by hypoxia/reoxygenation (H/R) in human kidney proximal tubular cells-2 (HK-2 cells). METHODS: There were 3 experimental groups: control, H/R injury and SMB-treated H/R injury. H/R injury of HK-2 cells was induced by first covering the cells with and then removing liquid paraffin wax. Different concentrations of compound SMB solution (0.05%, 0.10%, 0.15% or 0.20%) were administered to the SMB-treated H/R injury group before the hypoxic injury. After 4, 12 and 24 hrs of hypoxia and 4, 12, 24 and 48 hrs of reoxygenation, morphologic changes of HK-2 cells were observed under an inverted microscope. Cell viability was measured by the MTT method. Lactate dehydrogenase (LDH) activity in the culture supernatants was assayed using biochemical methods; TNF-alpha levels were determined by radioimmunoassay (RIA). RESULTS: The number of HK-2 cells was significantly reduced in the H/R injury group after hypoxia, and reached a nadir 24 hrs after hypoxia treatment. Various concentrations of SMB-treated groups showed significantly greater number of HK-2 cells than the H/R injury group. SMB solution (0.10%) produced the best effect. The levels of LDH and TNF-alpha in the H/R injury group were significantly increased, and reached a peak between 24 hrs of hypoxia and 4 hrs of reoxygenation when compared to the control group. Pre-treating with 0.10% SMB resulted in significantly lower levels of LDH and TNF-alpha than in the untreated H/R injury group at various time points of H/R. CONCLUSIONS: SMB has protective effects against H/R injury of HK-2 cells, possibly through inhibition of inflammatory cytokines.

Study of apoptosis induced by 188Re-DTPA-DG in MCF-7 breast carcinoma and A549 pulmonary carcinoma cells.

OBJECTIVE: To evaluate apoptosis induced by rehenium-188-labeled diethylenetriamine pentaacetic acid-glucosamine (188Re-DTPA-DG) in MCF-7 breast carcinoma cells and A549 pulmonary carcinoma cells. METHODS: Through the use of flow cytometry (FCM) with CBA software to detect apoptosis, cells of both the MCF-7 and A549 cell lines were divided into groups exposed to 188Re-DTPA-DG, 188Re-perrhenate (188ReO4-), and saline, respectively. The first two groups were further divided into subgroups on the basis of their exposure to radioactivity at 37, 55.5, or 74 kBq/mL, with the saline-exposed group divided into three corresponding subgroups. Each subgroup was introduced into 5 replicate wells of a culture plate, and the morphology of the cells in each well was determined by flow cytometry at 6-hour intervals for 18 hours. In order to determine the affinity of 188Re-DTPA-DG for tumor tissue, the biodistribution of the radiolabeled agent was assessed in breast tumor-bearing nude mice. RESULTS: Change in morphology of the cell nucleus was more evident in the 188Re-DTPA-DG-treated than in the 188ReO4--treated group, and no change in nuclear morphology was seen in the saline-exposed group. The study data suggested that there was a greater ratio of apoptotic to nonapoptotic cells among the 188Re-DTPA-DG-treated than among the 188ReO4--treated or saline-exposed cells (p<0.01), and a greater change in cell-nuclear morphology in the 188Re-DTPA-DG-treated than in the 188ReO4--treated cells. Furthermore, 188Re-DTPA-DG had a more significant apoptosis-inducing effect on both MCF-7 and A549 cells than did 188ReO4-. The biodistribution study in tumor-bearing nude mice showed that the concentration of 188Re-DTPA-DG in tumor tissue was much higher than in normal tissue, that 188Re-DTPA-DG was rapidly cleared from the blood, and that the main route of its clearance was via the kidneys. CONCLUSIONS: 188Re-DTPA-DG has a significant apoptotic effect on carcinoma cells. 188Re-DTPA-DG is an effective radiopharmaceutical for intratumoral radiation therapy.

Chronic hepatitis B serum promotes apoptotic damage in human renal tubular cells.

AIM: To investigate the effect of the serum of patients with chronic hepatitis B (CHB) on apoptosis of renal tubular epithelial cells in vitro and to study the role of hepatitis B virus (HBV) and transforming growth factor-beta (1) (TGF-beta (1)) in the pathogenesis of hepatitis B virus associated glomerulonephritis (HBV-GN). METHODS: The levels of serum TGF-beta(1) were measured by specific enzyme linked immunosorbent assay (ELISA) and HBV DNA was tested by polymerase chain reaction (PCR) in 44 patients with CHB ,and 20 healthy persons as the control. The normal human kidney proximal tubular cell (HK-2) was cultured together with the sera of healthy persons, CHB patients with HBV-DNA negative (20 cases) and HBV-DNA positive (24 cases) for up to 72 h. Apoptosis and Fas expression of the HK-2 were detected by flow cytometer. RESULTS: The apoptosis rate and Fas expression of HK-2 cells were significantly higher in HBV DNA positive serum group 19.01%+/-5.85% and 17.58%+/-8.35%, HBV DNA negative serum group 8.12%+/-2.80% and 6.96%+/-2.76% than those in control group 4.25%+/-0.65% and 2.33%+/-1.09%, respectively (P<0.01). The apoptosis rate and Fas expression of HK-2 in HBV DNA positive serum group was significantly higher than those in HBV DNA negative serum (P<0.01). Apoptosis rate of HK-2 cells in HBV DNA positive serum group was positively correlated with the level of HBV-DNA (r = 0.657). The level of serum TGF-beta (1) in CHB group was 163.05+/-91.35 microg/L, significantly higher as compared with 81.40+/-40.75 microg/L in the control group (P<0.01). CONCLUSION: The serum of patients with chronic hepatitis B promotes apoptotic damage in human renal tubular cells by triggering a pathway of Fas up-regulation. HBV and TGF-beta (1) may play important roles in the mechanism of hepatitis B virus associated glomerulonephritis.

[The relationship between TNF-alpha and the injury of renal tubular cells caused by anoxia/reoxygenation].

AIM: To investigate the relationship between TNF-alpha and renal tubular cell injury caused by anoxia/reoxygenation. METHODS: Human renal proximal tubular cell line HK-2 was used as model. Anoxia/reoxygenation were produced by covering/de-covering the cell culture with liquid paraffin wax. The level of TNF-alpha and the activity of lactate dehydrogenase(LDH) in the culture medium was determined by radioimmunoassay(RIA) and biochemical methods, respectively. Trypan blue exclusion was used to measure cell viability. RESULTS: Anoxia/reoxygenation could increase TNF-alpha level and LDH activity, but decrease viability of HK-2 cells. TNF-alpha level was positively correlated with LDH activity (r=0.89, P<0.05) and negatively with the cell viability (r=-0.91, P<0.05). CONCLUSION: TNF-alpha induced by anoxia/reoxygenation may participate in the process of renal tubular cell injury.