Genetic basis for variation in plasma IL-18 levels in persons with chronic hepatitis C virus and human immunodeficiency virus-1 infections.

Inflammasomes are multi-protein complexes integrating pathogen-triggered signaling leading to the generation of pro-inflammatory cytokines including interleukin-18 (IL-18). Hepatitis C virus (HCV) and human immunodeficiency virus (HIV) infections are associated with elevated IL-18, suggesting inflammasome activation. However, there is marked person-to-person variation in the inflammasome response to HCV and HIV. We hypothesized that host genetics may explain this variation. To test this, we analyzed the associations of plasma IL-18 levels and polymorphisms in 10 genes in the inflammasome cascade. About 1538 participants with active HIV and/or HCV infection in three ancestry groups are included. Samples were genotyped using the Illumina Omni 1-quad and Omni 2.5 arrays. Linear regression analyses were performed to test the association of variants with log IL-18 including HCV and HIV infection status, and HIV RNA in each ancestry group and then meta-analyzed. Eleven highly correlated single-nucleotide polymorphisms (r2=0.98-1) in the IL-18-BCO2 region were significantly associated with log IL-18; each T allele of rs80011693 confers a decrease of 0.06 log pg ml-1 of IL-18 after adjusting for covariates (rs80011693; rs111311302 ß=-0.06, P-value=2.7 × 10-4). In conclusion, genetic variation in IL-18 is associated with IL-18 production in response to HIV and HCV infection, and may explain variability in the inflammatory outcomes of chronic viral infections.

Preoperative radiotherapy in the management of retroperitoneal liposarcoma.

BACKGROUND: Histological subtype influences both prognosis and patterns of treatment failure in retroperitoneal sarcoma. Previous studies on the efficacy of neoadjuvant radiotherapy (NRT) have incorporated multiple histological types with heterogeneous tumour biology. The survival impact of NRT specifically for patients with retroperitoneal liposarcoma is poorly defined. METHODS: Patients who underwent resection with curative intent for retroperitoneal liposarcoma and who received NRT or surgery alone were identified in the US National Cancer Data Base (2004-2013). Cox regression was used to identify co-variables associated with overall survival. NRT and surgery-alone cohorts were matched 1 : 1 by propensity scores based on the survival hazard on Cox modelling. Overall survival was compared by Kaplan-Meier estimates. RESULTS: A total of 2082 patients with retroperitoneal liposarcoma were identified; 1908 underwent surgery alone and 174 received NRT before surgical resection. Median tumour size was 22·0 cm and 34·9 per cent of tumours were high grade. In the unmatched cohort, NRT was not associated with improved overall survival (χ2 = 3·49, P = 0·062). In the propensity score-matched cohort, NRT was associated with an improvement in survival (median overall survival 129·2 versus 84·3 months; P = 0·046; hazard ratio (HR) 1·54, 95 per cent c.i. 1·01 to 2·36). This effect appeared most pronounced for tumours with adjacent organ invasion (median overall survival not reached versus 63·8 months; P = 0·044; HR 1·79, 1·01 to 3·19). CONCLUSION: NRT improved survival in patients undergoing surgery for retroperitoneal liposarcoma, particularly those with high-risk pathological features.

Association of IFNL3 and IFNL4 polymorphisms with liver-related mortality in a multiracial cohort of HIV/HCV-coinfected women.

African Americans coinfected with HIV and hepatitis C virus (HCV) have lower liver-related mortality than Caucasians and Hispanics. While genetic polymorphisms near the IFNL3 and IFNL4 genes explain a significant fraction of racial differences in several HCV-related outcomes, the impact of these variants on liver-related mortality has not been investigated. We conducted a cohort study of HIV/HCV-coinfected women followed in the multicentre, NIH-funded Women's Interagency HIV Study (WIHS) to investigate whether 10 polymorphisms spanning the IFN-λ region were associated with liver-related mortality by dominant, recessive or additive genetic models. We also considered whether these polymorphisms contributed to previously reported differences in liver-related death by race/ethnicity (ascertained by self-report and ancestry informative markers). Among 794 coinfected women, there were 471 deaths including 55 liver-related deaths during up to 18 years of follow-up. On adjusted analysis, rs12980275 GG genotype compared to AG+AA hazards ratios [(HR) 0.36, 95% CI 0.14-0.90, P = 0.029] and rs8109886 AA genotype compared to CC+AC (HR 0.67, 95% CI 0.45-0.99, P = 0.047) were most strongly associated with liver-related death although these associations were no longer significant after adjusting for race/ethnicity (HR 0.41, 95% CI 0.16-1.04, P = 0.060 and HR 0.78, 95% CI 0.51-1.19, P = 0.25, respectively). African American women had persistently lower liver-related death independent of IFN-λ variants (HRs ≤ 0.44, P values ≤ 0.04). The lower risk of death among African American HIV/HCV-coinfected women is not explained by genetic variation in the IFN-λ region suggesting, that other genetic, behavioural and/or environmental factors may contribute to racial/ethnic differences in liver-related mortality.

Admixture analysis of spontaneous hepatitis C virus clearance in individuals of African descent.

Hepatitis C virus (HCV) infects an estimated 3% of the global population with the majority of individuals (75-85%) failing to clear the virus without treatment, leading to chronic liver disease. Individuals of African descent have lower rates of clearance compared with individuals of European descent and this is not fully explained by social and environmental factors. This suggests that differences in genetic background may contribute to this difference in clinical outcome following HCV infection. Using 473 individuals and 792,721 single-nucleotide polymorphisms (SNPs) from a genome-wide association study (GWAS), we estimated local African ancestry across the genome. Using admixture mapping and logistic regression, we identified two regions of interest associated with spontaneous clearance of HCV (15q24, 20p12). A genome-wide significant variant was identified on chromosome 15 at the imputed SNP, rs55817928 (P=6.18 × 10(-8)) between the genes SCAPER and RCN. Each additional copy of the African ancestral C allele is associated with 2.4 times the odds of spontaneous clearance. Conditional analysis using this SNP in the logistic regression model explained one-third of the local ancestry association. Additionally, signals of selection in this area suggest positive selection due to some ancestral pathogen or environmental pressure in African, but not in European populations.

Reducing infection transmission in solid organ transplantation through donor nucleic acid testing: a cost-effectiveness analysis.

For solid organ transplant (SOT) donors, nucleic acid-amplification testing (NAT) may reduce human immunodeficiency virus (HIV) and hepatitis C virus (HCV) transmission over antibody (Ab) testing given its shorter detection window period. We compared SOT donor NAT + Ab versus Ab alone using decision models to estimate incremental cost-effectiveness ratios (ICERs; cost per quality-adjusted life year [QALY] gained) from the societal perspective across a range of HIV/HCV prevalence values and NAT costs. The cost per QALY gained was calculated for two scenarios: (1) favorable: low cost ($150/donor)/high prevalence (HIV: 1.5%; HCV: 18.2%) and (2) unfavorable: high cost ($500/donor)/low prevalence (HIV: 0.1%; HCV: 1.5%). In the favorable scenario, adding NAT screening cost $161 013 per QALY gained for HIV was less costly) for HCV, and cost $86 653 per QALY gained for HIV/HCV combined. For the unfavorable scenario, the costs were $15 568 484, $221 006 and $10 077 599 per QALY gained, respectively. Universal HCV NAT + Ab for donors appears cost-effective to reduce infection transmission from SOT donors, while HIV NAT + Ab is not, except where HIV NAT is ≤$150/donor and prevalence is ≥1.5%. Our analyses provide important data to facilitate the decision to implement HIV and HCV NAT for deceased SOT donors and shape national policy regarding how to reduce infection transmission in SOT.

Molecular characterization of intrahepatic and extrahepatic hepatitis B virus (HBV) reservoirs in patients on suppressive antiviral therapy.

The hepatitis B virus (HBV) replicates via an error-prone reverse transcriptase generating potential drug-resistant quasispecies. The degree of HBV variability in liver vs peripheral blood mononuclear cells (PBMC) in patients on long-term suppressive antivirals is unclear. We characterized HBV replication, drug resistance and molecular diversity in patients with plasma HBV DNA undetectable by clinical assays. Explant liver (n=9), PBMC (n=6) and plasma (n=7) from nine such patients undergoing liver transplantation were evaluated for HBV genomes by sensitive PCR/nucleic acid hybridization assay. Cases with HBV DNA in liver and PBMC were tested for covalently closed circular DNA (HBV cccDNA). HBV polymerase (P) amplicons were cloned, sequenced and both P and overlapping surface (S) gene sequences were analysed. HBV DNA was detected in 43% (3/7) of plasma, 100% (9/9) of liver and 83% (5/6) of PBMC samples. HBV cccDNA was detected in all liver and one PBMC sample. Four patients had a clinical diagnosis of resistance. HBV P gene sequencing revealed 100% wild type (wt) in plasma (2/2), 83% wt in PBMC (5/6) but livers of 3/9 (33%) contained wt and 6/9 (66%) carried resistance to lamivudine and/or adefovir. The translated S gene revealed no changes affecting HBV antigenicity. Sequences from livers with antiviral resistant mutants revealed greater interpatient quasispecies diversity. Despite apparent HBV suppression, the liver continues to support HBV replication and extrahepatic HBV can be detected. PBMC may be a sanctuary for wt virus during antiviral therapy, while the liver harbours more drug-resistant viruses. Drug resistance correlates with intrahepatic viral diversity.

The Bb fragment of complement factor B acts as a B cell growth factor.

The process of B cell growth and differentiation into plasma cells is highly regulated and may be influenced by a large number of inflammatory mediators, including complement components. We have studied the regulatory influence of Bb, a 60-kD peptide created during the cleavage of complement Factor B by Factor D and C3b. Purified Bb alone had no effect on proliferation and differentiation of human splenic or tonsillar B cells. However, when B cells were activated by Staphylococcus aureus Cowan I (SAC), Bb enhanced proliferation in a dose-dependent manner. Bb also enhanced proliferation when cocultured with SAC and suboptimal concentrations of purified 60-kD B cell growth factor (HMW-BCGF), a previously described lymphokine that is known to possess growth-promoting activity. However, Bb had no effect on cells treated with optimal concentrations of HMW-BCGF. Like HMW-BCGF, Bb's major effect was on the larger in vivo activated B cells. Half-maximal enhancement of proliferation was reached at a Bb concentration of 1-10 nM. Of note is the fact that antibody to Factor B recognized HMW-BCGF, and an mAb to HMW-BCGF also recognized Factor B and Bb, but not Ba. Moreover, radiolabeled Bb bound saturably to activated B cells and to an EBV-transformed human B cell line. The binding of Bb was inhibited by HMW-BCGF but not by Ba or IgG. Thus, Bb is antigenically and functionally related to HMW-BCGF, and can act as a B cell growth and differentiation factor at potentially physiologic concentrations. These data suggest that Bb may be important in amplifying the immune response in areas of inflammation. Since complement activation occurs at inflammatory sites long before induction of HMW-BCGF synthesis, Bb may be an early signal for the clonal expansion of antigen-activated B cells.

Glypican-3 reexpression regulates apoptosis in murine adenocarcinoma mammary cells modulating PI3K/Akt and p38MAPK signaling pathways.

Glypican-3 (GPC3) is a proteoglycan involved in proliferation and cell survival. Several reports demonstrated that GPC3 is downregulated in some tumors, such as breast cancer. Previously, we determined that GPC3 reexpression in the murine mammary adenocarcinoma LM3 cells induced an impairment of their invasive and metastatic capacities, associated with a decrease of their motility and an increase of their cell death. We demonstrated that GPC3 inhibits canonical Wnt signaling, as well as it activates non canonical pathway. Now, we identified signaling pathways responsible for the pro-apoptotic role of GPC3 in LM3 cells. We found for the first time that GPC3 inhibits the PI3K/Akt anti-apoptotic pathway while it stimulates the p38MAPK stress-activated one. We report a concomitant modulation of CDK inhibitors as well as of pro- and anti-apoptotic molecules. Our results provide new clues regarding the mechanism involved in the modulation induced by GPC3 of mammary tumor cell growth and survival.

Progestin-induced caveolin-1 expression mediates breast cancer cell proliferation.

Progestin regulation of gene expression was assessed in the progestin-dependent murine tumor line C4HD which requires MPA, a synthetic progestin, for in vivo growth and expresses high levels of progesterone receptor (PR). By using suppressive subtractive hybridization, caveolin-1 was identified as a gene whose expression was increased with in vivo MPA treatment. By Northern and Western blot analysis, we further confirmed that caveolin-1 mRNA and protein expression increased in MPA-treated tumors as compared with untreated tumors. When primary cultures of C4HD cells were treated in vitro with MPA, caveolin-1 levels also increased, effect that was abolished by pre-treatment with progestin antagonist RU486. In addition, MPA promoted strong caveolin-1 promoter transcriptional activation both in mouse and human breast cancer cells. We also showed that MPA regulation of caveolin-1 expression involved in activation of two signaling pathways: MAPK and PI-3K. Short-term MPA treatment of C4HD cells led to tyrosine phosphorylation of caveolin-1 protein, where Src was the kinase involved. Additionally, we showed that MPA-induced association of caveolin-1 and PR, which was detected by coimmunoprecipitation and by confocal microscopy. Finally, we proved that MPA-induced proliferation of C4HD cells was inhibited by suppression of caveolin-1 expression with antisense oligodeoxynucleotides to caveolin-1 mRNA. Furthermore, we observed that inhibition of caveolin-1 expression abrogated PR capacity to induced luciferase activity from a progesterone response element-driven reporter plasmid. Comprehensively, our results demonstrated for the first time that caveolin-1 expression is upregulated by progestin in breast cancer. We also demonstrated that caveolin-1 is a downstream effector of MPA that is partially responsible for the stimulation of growth of breast cancer cells.

Normal human intestinal B lymphocytes. Increased activation compared with peripheral blood.

The state of activation of normal human intestinal mononuclear cells obtained from transplant donors was studied. Compared with PBMC, freshly isolated intestinal mononuclear cells expressed significantly more cell surface activation antigens on both B and T lymphocytes. Intestinal mononuclear cells contained significant numbers of immunoglobulin secreting cells immediately after cell separation. This population included CD5-positive B cells that secreted predominantly IgA. Cells from the large bowel consistently revealed higher numbers of IgA secreting cells than cells from the small bowel. Thus, intestinal B cells are markedly activated in vivo compared with PBMC and this increased activation correlates with increased spontaneous antibody secretion. B cells from the large intestine are more highly activated and secrete more antibody than do cells from the small intestine. The intestinal lamina propria lymphoid compartment exhibits a heightened state of activation that may be important for its distinct role in mucosal defense.

Disruption of the RAD52 gene alters the spectrum of spontaneous SUP4-o mutations in Saccharomyces cerevisiae.

Defects in the RAD52 gene of the yeast Saccharomyces cerevisiae confer a mutator phenotype. To characterize this effect in detail, a collection of 238 spontaneous SUP4-o mutations arising in a strain having a disrupted RAD52 gene was analyzed by DNA sequencing. The resulting mutational spectrum was compared to that derived from an examination of 222 spontaneous mutations selected in a nearisogenic wild-type (RAD52) strain. This comparison revealed that the mutator phenotype was associated with an increase in the frequency of base-pair substitutions. All possible types of substitution were detected but there was a reduction in the relative fraction of A.T----G.C transitions and an increase in the proportion of G.C----C.G transversions. These changes were sufficient to cause a twofold greater preference for substitutions at G.C sites in the rad52 strain despite a decrease in the fraction of G.C----T.A transversions. There were also considerable differences between the distributions of substitutions within the SUP4-o gene. Base-pair changes occurred at fewer sites in the rad52 strain but the mutated sites included several that were not detected in the RAD52 background. Only two of the four sites that were mutated most frequently in the rad52 strain were also prominent in the wild-type strain and mutation frequencies at almost all sites common to both strains were greater for the rad52 derivative. Although single base-pair deletions occurred in the two strains with similar frequencies, several classes of mutation that were recovered in the wild-type background including multiple base-pair deletions, insertions of the yeast transposable element Ty, and more complex changes, were not detected in the rad52 strain.(ABSTRACT TRUNCATED AT 250 WORDS)

Fulminant hepatic failure resulting from lamivudine-resistant hepatitis B virus in a renal transplant recipient: durable response after orthotopic liver transplantation on adefovir dipivoxil and hepatitis B immune globulin.

BACKGROUND: Mutations in the hepatitis B virus (HBV) genome may occur during therapy. METHODS: We report an asymptomatic HBV carrier who underwent transplantation for end-stage renal disease. She developed an HBV flare 6 months after transplantation and was placed on lamivudine. After initial rapid improvement, she relapsed clinically and virologically. She decompensated with jaundice, peripheral edema, ascites, encephalopathy, coagulopathy, and hepatorenal syndrome. A liver biopsy specimen revealed submassive necrosis. RESULTS: Emergency liver transplantation was performed: lamivudine was discontinued. Hepatitis B immunoglobulin and adefovir dipivoxil were initiated. Sixteen months after orthotopic liver transplantation, she is HBV DNA seronegative with normal liver enzymes. Sequencing of HBV polymerase gene from preliver transplantation sera did not detect the usual lamivudine resistance mutations in the YMDD motif but instead two other mutations (F514-->L, L528-->M). Lamivudine resistance was demonstrated in vitro. CONCLUSIONS: Asymptomatic HBV carriers may reactivate following renal transplantation after immunosuppression. Resistance to lamivudine may result in severe hepatic damage in immunocompromised patients.

Pathogenesis of primary biliary cirrhosis, primary sclerosing cholangitis, and autoimmune cholangiopathy.

Autoimmune liver diseases have much in common with each other, and there are clear associations with genetic haplotypes. Elegant studies have shown autoimmune liver disease induced by viruses and drugs. Although there is evidence for nonimmunological events precipitating immune disease, especially in primary sclerosing cholangitis, the precise pathways, what is bystander and what is essential, have not been determined. This article reviews some of the mechanisms involved in pathogenesis.

Altered enteroendocrine cell expression in T cell receptor alpha chain knock-out mice.

Mice lacking T cell receptor alpha chain (TCRalpha(-/-)) develop inflammation of the colon. We have examined the effect of this inflammation on the colonic epithelium by studying markers of epithelial cuff, enteroendocrine, and immune cell differentiation. Using immunohistochemical techniques, colons were compared in normal C57/BL6 and murine TCR alpha(-/-) mice aged 2 and 3 weeks and 3-11 months. TCR alpha(-/-) mice aged 3-11 months had histologic evidence of inflammation with increased expression of CD45, CD4+, CD8+, and B220+ cells and a decrease in expression of IgA+ cells. There was a decrease in the number of cholecystokinin, serotonin, and neurotensin enteroendocrine expressing cells in the colon of TCR alpha(-/-) mice. These changes were not present in 2-3-week-old suckling/weaning mice. In contrast, peptide tyrosine tyrosine (PYY), glucagon-like peptide-1, and gastrin expression did not change and small intestinal enteroendocrine cells remained unaltered. The change in colonic enteroendocrine cell expression appears to be a specific response, since only a subset of these cells was altered, and the epithelium was intact by histologic analysis. The absence of functional T cells in TCR alpha(-/-) colon has a marked effect on differentiation of a specific subpopulation of enteroendocrine cells, prior to loss of integrity of the epithelium.

Day-5 protocol liver allograft biopsies document early rejection episodes and are predictive of recurrent rejection.

METHODS: The day-5 posttransplant protocol biopsy specimens and clinical courses of 27 consecutive orthotopic liver transplant recipients followed up at least 6 months were reviewed. RESULTS: Twelve (44%) of 27 patients had histologic evidence of rejection on the day-5 biopsy; 8 (67%) of these 12 patients required OKT3 for reversal of the rejection. No significant differences in biochemical liver test results, bile output, or cyclosporine levels were observed between this group and the 15 patients (56%) without histologic evidence of rejection on day 5. Eight (67%) of the 12 patients with rejection had recurrent rejection episodes, with one recurrence each in six patients, two recurrences in one patient, and three recurrences in one patient. Of the 15 patients without rejection on day 5, nine (60%) subsequently had rejection at 10 days, 14 days, and 1 1/2, 3 1/2, 4, 5, and 11 months after transplantation. Only one (11%) of these nine patients had a recurrent rejection episode. There were no differences in the incidence of posttransplant cytomegalovirus infections between the two groups. Two cases of posttransplant lymphoma were seen; they developed in two patients without rejection on the day-5 biopsy. No patients or allografts were lost to acute or chronic rejection. No complications occurred as a result of the day-5 protocol biopsy. CONCLUSION: The day-5 protocol biopsy is useful in detecting rejection episodes that may not otherwise be clinically apparent.

Effects of antiprogestins RU486 and ZK98299 on the expression of cell cycle proteins of a medroxyprogesterone acetate (MPA)-induced murine mammary tumor.

This study links the tumor inhibitory effect of anti-progestins RU486 and ZK98299 to the expression of cell cycle proteins. Medroxyprogesterone acetate-induced ductal mammary adenocarcinoma-bearing female BALB/c mice were treated daily either with RU486 or ZK98299, observing tumor growth retardation. p21 increased after 24 h treatment, peaked at day 4 and returned to control levels at day 7. Cyclin D1, cyclin E, CDK2 and CDK4, did not change during treatment. These results suggest that p21 might play a role in early inhibitory stages of tumor growth induced by RU486 and ZK98299.

Base-pair substitutions alter the site-specific mutagenicity of UV and MNNG in the SUP4-o gene of yeast.

Yeast strains carrying SUP4-o genes that have base-pair substitutions at hotspots for UV or MNNG mutagenesis were treated with these agents. In both cases, the induced mutation frequencies were substantially reduced. Furthermore, specific substitutions at positions in SUP4-o that had not been mutated by MNNG resulted in the recovery of MNNG-induced mutations at these sites. These results demonstrate that base-pair identity is an important factor determining the site-specific mutagenicity of UV and MNNG in yeast. For UV, our findings suggest that the type of lesion that is induced, but not flanking DNA sequences, plays a role in specifying mutability at the sites examined. In contrast, DNA sequence context seems to be an important factor for MNNG mutagenesis.

Metastasis suppressors: basic and translational advances.

Tumor metastasis is a main contributor to death in cancer patients. In the last years, a new class of molecules that reduces the metastatic propensity has been identified: metastasis suppressors. These proteins regulate multiple steps in the metastatic cascade, including cell invasion, survival in the vascular and lymphatic circulation, and colonization of distant organ sites. As a consequence, they are very important therapeutic targets. This review discusses our current understanding of metastasis suppressors and how this knowledge might be useful to improve the treatment of cancer patients.

Lower peripheral blood CD14+ monocyte frequency and higher CD34+ progenitor cell frequency are associated with HBV vaccine induced response in HIV infected individuals.

We evaluated immunologic predictors of response to HBV vaccine administered in the presence or absence of GM-CSF in HIV infected individuals. We measured peripheral blood hematopoietic progenitor, monocyte and myeloid-derived suppressor cell (MDSC) frequencies, and expression of GMCSF receptor on monocytes and MDSCs, at baseline and 4weeks after immunization in relation to antibody response. We observed higher baseline progenitor and lower monocyte frequencies among week 16 antibody responders. Week 4 decline in MDSC frequency was associated with week 16 antibody response, while administration of GM-CSF was associated with preservation of these cells. No significant differences in GM-CSF receptor expression were observed in the presence vs. absence of GM-CSF. These findings are consistent with a positive role of progenitor cells and a potential negative role of monocytes in vaccine response. Additionally, GM-CSF augmented the preservation of peripheral blood MDSC, which may contribute to the lack of improved vaccine responses.