Ronghua Jin

Interferon-induced transmembrane protein-3 genetic variant rs12252-C is associated with severe influenza in Chinese individuals.

The SNP rs12252-C allele alters the function of interferon-induced transmembrane protein-3 increasing the disease severity of influenza virus infection in Caucasians, but the allele is rare. However, rs12252-C is much more common in Han Chinese. Here we report that the CC genotype is found in 69% of Chinese patients with severe pandemic influenza A H1N1/09 virus infection compared with 25% in those with mild infection. Specifically, the CC genotype was estimated to confer a sixfold greater risk for severe infection than the CT and TT genotypes. More importantly, because the risk genotype occurs with such a high frequency, its effect translates to a large population-attributable risk of 54.3% for severe infection in the Chinese population studied compared with 5.4% in Northern Europeans. Interferon-induced transmembrane protein-3 genetic variants could, therefore, have a strong effect of the epidemiology of influenza in China and in people of Chinese descent.

Oncogenic activation of glypican‐3 by c‐Myc in human hepatocellular carcinoma

Glypican‐3 (GPC3) is a heparan sulfate proteoglycan that has an important role in cell growth and differentiation, and its function in tumorigenesis is tissue‐dependent. In hepatocellular carcinoma (HCC), the overexpression of GPC3 has been demonstrated to be a reliable diagnostic indicator. However, the mechanisms that regulate the expression and function of GPC3 remain unclear. The oncoprotein c‐Myc is a transcription factor that plays a significant role in more than 50% of human tumors. We report here that GPC3 is a transcriptional target of c‐Myc and that the expression of c‐Myc is also regulated by GPC3, thus forming a positive feedback signaling loop. We found that the overexpression of c‐Myc could induce GPC3 promoter‐dependent luciferase activity in luciferase reporter experiments. Furthermore, mutational analysis identified c‐Myc‐binding sites within the GPC3 promoter. The exogenous overexpression of c‐Myc increased the endogenous messenger RNA (mRNA) and protein levels of GPC3. Chromatin immunoprecipitation experiments revealed the binding of c‐Myc to the endogenous GPC3 promoter, indicating that c‐Myc can directly transcriptionally activate GPC3. Interestingly, GPC3 can also elevate c‐Myc expression. Overexpression of GPC3 increased c‐Myc protein levels, whereas the knockdown of GPC3 reduced c‐Myc expression levels. Lastly, the elevated levels of c‐Myc correlate with the overexpression of GPC3 in human HCC samples. Conclusion: These data provide new mechanistic insight into the roles of GPC3 and of c‐Myc in the development of HCC. (HEPATOLOGY 2012)

Long-Term Exposure of Mice to Nucleoside Analogues Disrupts Mitochondrial DNA Maintenance in Cortical Neurons

Nucleoside analogue reverse transcriptase inhibitor (NRTI), an integral component of highly active antiretroviral therapy (HAART), was widely used to inhibit HIV replication. Long-term exposure to NRTIs can result in mitochondrial toxicity which manifests as lipoatrophy, lactic acidosis, cardiomyopathy and myopathy, as well as polyneuropathy. But the cerebral neurotoxicity of NRTIs is still not well known partly due to the restriction of blood-brain barrier (BBB) and the complex microenvironment of the central nervous system (CNS). In this study, the Balb/c mice were administered 50 mg/kg stavudine (D4T), 100 mg/kg zidovudine (AZT), 50 mg/kg lamivudine (3TC) or 50 mg/kg didanosine (DDI) per day by intraperitoneal injection, five days per week for one or four months, and primary cortical neurons were cultured and exposed to 25 µM D4T, 50 µM AZT, 25 µM 3TC or 25 µM DDI for seven days. Then, single neuron was captured from mouse cerebral cortical tissues by laser capture microdissection. Mitochondrial DNA (mtDNA) levels of the primary cultured cortical neurons, and captured neurons or glial cells, and the tissues of brains and livers and muscles were analyzed by relative quantitative real-time PCR. The data showed that mtDNA did not lose in both NRTIs exposed cultured neurons and one month NRTIs treated mouse brains. In four months NRTIs treated mice, brain mtDNA levels remained unchanged even if the mtDNA levels of liver (except for 3TC) and muscle significantly decreased. However, mtDNA deletion was significantly higher in the captured neurons from mtDNA unchanged brains. These results suggest that long-term exposure to NRTIs can result in mtDNA deletion in mouse cortical neurons.

E2F1 is involved in DNA single-strand break repair through cell-cycle-dependent upregulation of XRCC1 expression.

The X-ray repair cross complementing group 1 (XRCC1) protein is involved in DNA base excision repair and its expression varies during the cell cycle. Although studies have demonstrated that rapid XRCC1-dependent single-strand break repair (SSBR) takes place specifically during S/G(2) phases, it remains unclear how it is regulated during the cell cycle. We found that XRCC1 is a direct regulatory target of E2F1 and further investigated the role of XRCC1 in DNA repair during the cell cycle. Saos2 primary osteosarcoma cells stably transfected with inducible E2F1-wt or mutant E2F1-132E were treated with hydroxurea (HU) for 36h and were subsequently withdrawn HU for 2-24h to test whether cell-cycle-dependent DNA SSBR requires E2F1-mediated upregulation of XRCC1. We found that SSBR activity, as determined using a qPCR-base method, was correlated with E2F1 levels at different phases of the cell cycle. XRCC1-positive (AA8) and negative (EM9) CHO cells were used to demonstrate that the alterations in SSBR were mediated by XRCC1. The results indicate that E2F1-mediated regulation of XRCC1 is required for cell-cycle-dependent SSBR predominantly in G(1)/S phases. Our observations have provided new mechanistic insight for understanding the role of E2F1 in the maintenance of genomic stability and cell survival during the cell cycle. The regulation of XRCC1 by E2F1 during cell-cycle-dependent SSBR might be an important aspect for practical consideration for resolving the problem of drug resistance in tumor chemotherapies.

A new approach utilizing real-time qPCR to detect in vitro base excision repair.

DNA lesions in mammalian cells may be induced by reactive oxygen species, alkylation, and ionizing radiation. This damage can then be repaired via the base excision repair (BER) pathway, which includes single strand break repair (SSBR). Thus, the BER (SSBR) pathway plays a critical role in maintaining genomic integrity, and may help us to better understand the mechanisms of aging, tumor formation, and degenerative diseases. AP site (apurinic/apyrimidinic site) or damaged base excision, nucleotide insertion and ligation are included in the BER (SSBR) pathway, which are facilitated by different enzymes at each step. Most previous in vitro BER studies have used modified radiolabeled (32)P oligonucleotide substrates. Which is a very conventional method for in vitro BER assay. However, the use of radioactive isotope material was limited in various laboratories which are unable to handle radioactive hazard. In this study, we developed a novel technique using real-time quantitative PCR (qPCR) to quantify BER activity in in vitro assays. Single strand breaks, DNA ligase activity, and glycosylase activity were detected to establish the feasibility and advantages of this qPCR technique for in vitro BER profiling.

Cytokine levels contribute to the pathogenesis of minimal hepatic encephalopathy in patients with hepatocellular carcinoma via STAT3 activation.

Hepatocellular carcinoma (HCC) patients were grouped according to the degree of encephalopathy, with healthy volunteers as controls. We investigated clinical presentation, protein and mRNA expression of 14 cytokines, and activation of six STAT proteins, the downstream signaling mediators. Levels of all 14 cytokines were significantly elevated in HCC patients with clinical hepatic encephalopathy. Statistical analysis showed that levels of IL-1β, IL-6, IFNγ, IL-17α, IFNλ2 and IFNλ3 were correlated with minimal hepatic encephalopathy (MHE). Multivariate regression analysis identified serum IL-6, IFNλ3 and IL-17α as independent risk factors for MHE. Increased mRNA levels of IL-6 and IFNγ were associated with MHE. Among the STAT proteins examined, only STAT3 was elevated in MHE. Treatment with a STAT3 inhibitor protected neurons from cytokine-induced apoptosis in vitro. In conclusion, this study identified potential biomarkers for MHE in HCC. The cytokines investigated may induce neural apoptosis via STAT3 in the pathogenesis of MHE in HCC.

High levels of divergent HIV-1 quasispecies in patients with neurological opportunistic infections in China

Despite the fact that the survival of people infected with human immunodeficiency virus (HIV) has improved worldwide because of the increasingly powerful and highly active antiretroviral therapy, opportunistic infections (OIs) of the central nervous system (CNS) remain a serious burden. HIV-1 is capable of entering the CNS through infected peripheral monocytes, but its effect on OIs of CNS remains unclear. In this study, we investigated the characteristics of HIV-1 in acquired immunodeficiency syndrome (AIDS) patients with CNS OIs. A total of 24 patients with CNS OIs and 16 non-CNS OIs (control) cases were selected. These AIDS patients were infected with HIV-1 by paid blood donors in China. HIV-1 loads in plasma and cerebrospinal fluid (CSF) were detected using RT-PCR, and the C2-V5 region of HIV-1 envelope gene was amplified from viral quasispecies isolated from CSF using nested PCR. The CSF HIV-1 load of CNS OIs was higher than that of non-CNS OIs, but plasma HIV-1 load of CNS OIs was not higher than that of non-CNS OIs. The nucleotide sequence of C2-V5 region of the HIV-1 quasispecies isolated from the CSF of CNS OIs had a high diversity, and the HIV-1 quasispecies isolated from the CSF of CNS OIs revealed R5 tropism as 11/25 charge rule. These results suggest that high levels of divergent HIV-1 quasispecies in the CNS probably contribute to opportunistic infections.

Combination of long-acting HIV fusion inhibitor albuvirtide and LPV/r showed potent efficacy in HIV-1 patients

Long acting antiretroviral drugs represent a promising approach for chronic treatment of HIV infection. Here, we study the efficacy and safety of albuvirtide (ABT), an HIV-1 fusion inhibitor with a half life of 11–12 days in human. ABT was evaluated in a 7-week, open-label and randomized trial, combining with LPV/r. Twenty HIV-1-infected adults were assigned to two dose groups, receiving ABT (160 or 320 mg) given weekly and LPV/r given twice daily. At week 7, the decline of HIV-1 RNA from baseline was 1.9 (1.3–2.3) log10 and 2.2 (1.6–2.7) log10 copies/ml, and suppression of HIV-1 RNA to below 50 copies/ml was achieved in 11.1 % (1/9) and 55.6 % (5/9) patients, for the 160 and 320 mg dose group respectively. A clear dose-efficacy correlation of ABT was demonstrated. ABT combining with LPV/r is a promising two-drug regimen to be tested in larger patient population.

Clinical correlation between HBV infection and concomitant bacterial infections

Bacterial infections are common in patients suffering viral hepatitis and critical for prognosis. However, any correlation between HBV and concomitant bacterial infections is not well characterized. A retrospective study was conducted from Jan 2012 to Jan 2014 on 1333 hospitalized patients infected with bacteria. Among them, 491 HBV-infected patients were co-infected with E. coli (268), S. aureus (61), P. aeruginosa (64) or K. pneumoniae (98). A group of 300 complication-free chronically HBV-infected patients were controls. We found that HBV DNA levels were elevated in patients with each of the bacterial infections (all P < 0.05). ALT and HBeAg were strong determinants of high HBV DNA concentration. Patterns of determinants varied in infections by Gram-positive and Gram-negative bacteria. Patients with HBV DNA ≥ 2000 IU/mL had higher rates of all four concomitant bacterial infections (all P < 0.001). All types of strains isolated from HBV-positive patients showed less resistance to tested antimicrobials. The HBV DNA serum concentrations were inversely correlated to the number of ineffective antimicrobials in E. coli, P. aeruginosa and K. pneumoniae infections (P = 0.022, 0.017 and 0.016, respectively), but not S. aureus (P = 0.194). In conclusion, bacterial infections are associated with a high level of HBV replication, which, in turn, has a significant positive impact on bacterial resistance to antimicrobials. These correlations vary between Gram-negative and Gram-positive bacteria.

Diffuse large B‑cell lymphoma with pulmonary and cerebral involvement: A case report

Primary diffuse large B-cell lymphoma (DLBCL) is the most common type of non-Hodgkin lymphoma; however, the involvement of the lung and central nervous system (CNS) in patients with DLBCL is rare. Furthermore, patients with DLBCL rarely exhibit specific clinical symptoms, which may delay definitive diagnosis. The present study reports the case of a 42-year-old man suffering from primary DLBCL with concurrent pulmonary and cerebral involvement. The patient suffered from human immunodeficiency virus infection and presented with symptoms including dry cough, thoracalgia, intermittent mild fever and mild headache. Thoracic computed tomography scans revealed multiple pulmonary masses, and brain magnetic resonance imaging scans revealed nodules in the left frontal cortex and bilateral basal ganglia. A percutaneous lung needle biopsy test confirmed the diagnosis of DLBCL. In addition, positron emission tomography revealed the involvement of other parts of the body in DLBCL. The aim of the present study was to present the clinical, radiological and histological characteristics of the patient, which may aid physicians in diagnosing pulmonary and CNS involvement in DLBCL.