GW4064, a farnesoid X receptor agonist, upregulates adipokine expression in preadipocytes and HepG2 cells.

AIM: To investigate the effect of GW4064 on the expression of adipokines and their receptors during differentiation of 3T3-L1 preadipocytes and in HepG2 cells. METHODS: The mRNA expression of farnesoid X receptor (FXR), peroxisome proliferator-activated receptor-gamma 2 (PPAR-γ2), adiponectin, leptin, resistin, adiponectin receptor 1 (AdipoR1), adiponectin receptor 2 (AdipoR2), and the long isoform of leptin receptor (OB-Rb) and protein levels of adiponectin, leptin, and resistin were determined using fluorescent real-time PCR and enzyme linked immunosorbent assay, respectively, on days 0, 2, 4, 6, and 8 during the differentiation of 3T3-L1 preadipocytes exposed to GW4064. Moreover, mRNA expression of AdipoR2 and OB-Rb was also examined using fluorescent real-time PCR at 0, 12, 24, and 48 h in HepG2 cells treated with GW4064. RESULTS: The mRNA expression of FXR, PPAR-γ2, adiponectin, leptin, resistin, AdipoR1, AdipoR2, and OB-Rb and protein levels of adiponectin, leptin, and resistin increased along with differentiation of 3T3-L1 preadipocytes (P < 0.05 for all). The mRNA expression of FXR, PPAR-γ2, adiponectin, leptin, and AdipoR2 in 3T3-L1 preadipocytes, and AdipoR2 and OB-Rb in HepG2 cells was significantly increased after treatment with GW4064, when compared with the control group (P < 0.05 for all). A similar trend was observed for protein levels of adipokines (including adiponectin, leptin and resistin). However, the expression of resistin, AdipoR1, and OB-Rb in 3T3-L1 cells did not change after treatment with GW4064. CONCLUSION: The FXR agonist through regulating, at least partially, the expression of adipokines and their receptors could offer an innovative way for counteracting the progress of metabolic diseases such as nonalcoholic fatty liver disease.

Position 22 of the V3 loop is associated with co-receptor usage and disease progression in HIV-1 subtype B isolates.

Our goal in this study was to analyze position 22 of the V3 loop associated with co-receptor usage and disease progression in human immunodeficiency virus type 1 (HIV-1) subtype B infection. Bioinformatics approaches were used to compare the amino acid sequence and secondary structure of the V3 loop of the CCR5-tropic virus and CXCR4-tropic virus in HIV-1 subtype B. HIV-1 subtype B V3 amino acid sequence files in the FASTA format were collected from the HIV Sequence Database. The amino acid sequences of different tropism were multiple-aligned with CLUSTAL W program, and the frequencies of the amino acids at each position of the V3 loop sequences of two groups were calculated and sorted in descending order. The secondary structure of the consensus V3 amino acid sequences from CCR5-tropic and CXCR4-tropic viruses were predicted with the APSSP2 method. The amino acids at positions 11, 22, and 25 of V3 were different between the CCR5-tropic virus and CXCR4-tropic virus. The consensus amino acid frequencies were found to be 71.9% S, 66.7% A, and 56.0% D for the CCR5-tropic virus and 50.0% R, 57.1% T, and 26.2% Q for the CXCR4- tropic virus at positions 11, 22, and 25, respectively. There was a strong association between the identity of the residues at position 11, 22, and 25 of the V3 loop amino acid sequence and CD4+ T cell counts of different patients. The change of the residue at position 22 in the R5-tropic or X4-tropic viruses is expected to likely change the secondary structure to be similar to the X4-tropic or R5-tropic viruses. Our study indicates that position 22 of the V3 loop amino acid sequence is significantly associated with viral tropism and disease progression in HIV-1 subtype B.

Combination of small interfering RNAs mediates greater suppression on hepatitis B virus cccDNA in HepG2.2.15 cells.

AIM: To observe the inhibition of hepatitis B virus (HBV) replication and expression in HepG2.2.15 cells by combination of small interfering RNAs (siRNAs). METHODS: Recombinant plasmid psil-HBV was constructed and transfected into HepG2.2.15 cells. At 48 h, 72 h and 96 h after transfection, culture media were collected and cells were harvested for HBV replication assay. HBsAg and HBeAg in the cell culture medium were detected by enzyme-linked immunoadsorbent assay (ELISA). Intracellular viral DNA and covalently closed circular DNA (cccDNA) were quantified by real-time polymerase chain reaction (PCR). HBV viral mRNA was reverse transcribed and quantified by reverse-transcript PCR (RT-PCR). RESULTS: siRNAs showed marked anti-HBV effects. siRNAs could specifically inhibit the expression of HBsAg and the replication of HBV DNA in a dose-dependent manner. Furthermore, combination of siRNAs, compared with individual use of each siRNA, exerted a stronger inhibition on antigen expression and viral replication. More importantly, combination of siRNAs significantly suppressed HBV cccDNA amplification. CONCLUSION: Combination of siRNAs mediates a stronger inhibition on viral replication and antigen expression in HepG2.2.15 cells, especially on cccDNA amplification.

Influenza virus infection and risk of acute myocardial infarction.

Increasing evidences have shown that pathogens might promote atherosclerosis and trigger acute myocardial infarction (AMI). But the conclusions from various studies on the correlation between previous influenza virus (IV) infection and AMI were inconsistent. We conducted a case-control study to assess the association of previous IV infection and AMI. Questionnaire survey was conducted to collect information about demographic characteristics and heart disease risk factors. Fasting blood sample was obtained to measure IgG antibodies to influenza virus A(IV-A), influenza virus B(IV-B), cytomegalovirus (CMV), herpes simplex virus type-1 (HSV-1) and type-2 (HSV-2), adenovirus (ADV), rubella virus (RV) and Chlamydia pneumoniae (CP) and measure the level of some biochemistry markers. Compared to controls, cases were more likely to have positive IgG antibodies to IV-A and IV-B (IV-A: OR, 3.3; 95%CI, 1.5 to 7.4; IV-B: OR, 17.2; 95%CI, 7.7 to 38.0). After adjustment for potential confounding variables, the risk of AMI was still associated with the presence of IgG antibodies to IV-A (adjusted OR, 7.5; 95%CI, 1.3 to 43.0) and IV-B (adjusted OR, 27.3; 95%CI, 6.6 to 113.8). The study supported the hypothesis that previous IV infection took part in the development of atherosclerosis and trigger the occurrence of AMI.

Combination therapy of siRNAs mediates greater suppression on hepatitis B virus cccDNA in HepG2.2.15 cell.

BACKGROUND/AIMS: Hepatitis B virus (HBV) infection is a world-wide health problem. The major obstacles for current anti-HBV therapy are the low efficacy and the occurrence of drug resistant HBV mutations. Recent studies have demonstrated that combination therapy can enhance antiviral efficacy and overcome the shortcomings. Here, the inhibitory effect mediated by combination of small interfering RNAs (siRNAs) targeting different sites of HBV nuclear localization signal (NLS) was monitored in HepG2.2.15 cells. METHODOLOGY: Recombinant plasmid psil-HBV was constructed and transfected into HepG2.2.15 cells. At 48, 72 and 96 h after transfection, culture media were collected and cells were harvested for HBV replication assay. HBsAg and HBeAg in the cell culture medium were detected by enzyme-linked immunoadsorbent assay. Intracellular viral DNA and covalently closed circular DNA (cccDNA) was quantified by real-time PCR. HBV viral mRNA was reverse transcribed and quantified by reverse-transcript PCR. RESULTS: Our data demonstrated that three used siRNAs showed marked anti-HBV effects. The expression of HBsAg and the replication of HBV DNA could be specifically inhibited in a dose-dependent manner by siRNAs. Furthermore, combination of siRNAs, compared with individual use of each siRNA, exerted a stronger inhibition on antigen expression and viral replication, even though the final concentration of siRNA in the therapy was the same. More importantly, we showed that combination therapy significantly suppressed HBV cccDNA amplification. CONCLUSION: Our results revealed that combination of siRNAs mediated a stronger inhibition on viral replication and antigen expression in HepG2.2.15 cells, especially, the amplification of cccDNA.