Is microRNA-33 an Appropriate Target in the Treatment of Atherosclerosis?

The maintenance of cholesterol homeostasis is a complicated process involving regulation of cholesterol synthesis, dietary uptake and bile acid synthesis and excretion. Reverse cholesterol transport, described as the transfer of cholesterol from non-hepatic cells, including foam cells in atherosclerotic plaques, to the liver and then its excretion in the feces is important part of this regulation. High-density lipoproteins are the key mediators of reverse cholesterol transport. On the other hand, microRNA-33 was identified as a key regulator of cholesterol homeostasis. Recent studies indicate the impact of microRNA-33 not only on cellular cholesterol efflux and HDL production but also on bile metabolism in the liver. As proper coordination of cholesterol metabolism is essential to human health, discussion of recent findings in this field may open new perspectives in the microRNA-dependent treatment of a cholesterol imbalance.

The Impact of Short-Term Shark Liver Oil Supplementation on the Fatty Acid Composition of Erythrocyte Membranes.

Fatty acid (FA) balance is strictly related to human health. The composition of fatty acids in lipid membranes seems to be influenced by diet. Shark liver oil (SLO) supplementation has been widely used recently in the prevention and treatment of human diseases. We analyzed the impact of short-term SLO supplementation on certain biochemical parameters and erythrocyte FA composition in a group of young healthy women. Our results showed that 6 weeks of SLO supplementation led to a significant decrease in C-reactive protein levels in sera and intracellular cholesterol levels in peripheral blood mononuclear cells. SLO supplementation caused a significant increase in the content of the polyunsaturated omega-3 FAs: docosahexaenoic acid, docosapentaenoic acid and α-linolenic acid. In the group of omega-6 FAs, we observed a significant elevation of arachidonic and dihomo-gamma-linoleic acid content. Due to these alterations, the omega-3 index increased significantly from 3.6% (before) to 4.2% (after supplementation). We also observed the impact of SLO supplementation on the membrane fluidity index. The ratio between saturated and unsaturated FAs decreased significantly from 13.1 to 9.9. In conclusion, our results show that even short-term SLO supplementation can improve human erythrocyte fatty acid composition and other parameters that may have health-promoting consequences.

Hepatitis C Virus Uses Host Lipids to Its Own Advantage.

Lipids and lipoproteins constitute indispensable components for living not only for humans. In the case of hepatitis C virus (HCV), the option of using the products of our lipid metabolism is "to be, or not to be". On the other hand, HCV infection, which is the main cause of chronic hepatitis, cirrhosis and hepatocellular carcinoma, exerts a profound influence on lipid and lipoprotein metabolism of the host. The consequences of this alternation are frequently observed as hypolipidemia and hepatic steatosis in chronic hepatitis C (CHC) patients. The clinical relevance of these changes reflects the fact that lipids and lipoprotein play a crucial role in all steps of the life cycle of HCV. The virus circulates in the bloodstream as a highly lipidated lipo-viral particle (LVP) that defines HCV hepatotropism. Thus, strict relationships between lipids/lipoproteins and HCV are indispensable for the mechanism of viral entry into hepatocytes, viral replication, viral particles assembly and secretion. The purpose of this review is to summarize the tricks thanks to which HCV utilizes host lipid metabolism to its own advantage.

Changes in miR-122 and Cholesterol Expression in Chronic Hepatitis C Patients after PegIFN-Alpha/Ribavirin Treatment.

The hepatitis C virus (HCV) is known as a main etiological cause of chronic hepatitis. HCV infection disturbs cholesterol metabolism of the host, which is frequently observed in patients suffering from chronic hepatitis C (CHC). The course of viral infections remains under strict control of microRNA (miRNA). In the case of HCV, miR-122 exerts a positive effect on HCV replication in vitro. The purpose of this study was to investigate the impact of peginterferon alpha (pegIFN-α) and ribavirin treatments on the expression of miR-122 and the cholesterol level in the peripheral blood mononuclear cells (PBMCs) of CHC patients. We report here that the level of miR-122 expression in the PBMCs decreased after the antiviral treatment in comparison to the pretreated state. Simultaneously, the level of cholesterol in the PBMCs of CHC patients was higher six months following the treatment than it was pretreatment. Consequently, it seems that the decrease of miR-122 expression in the PBMCs of CHC patients is one of the antiviral effects connected with the pegIFN-alpha/ribavirin treatments.

The Correlation between miR-122 and Lipoprotein Lipase Expression in Chronic Hepatitis C Patients.

Chronic HCV infection is strictly associated with host lipid/lipoprotein metabolism disorders. The study aimed to analyze the relationship between viral load, lipid profile, IFNγ, and the expression of miR-122 and LPL in the liver and PBMCs. Sera, PBMCs, and matching liver biopsies from 17 chronic hepatitis C patients were enrolled in this study. Collected data shows that liver (not PBMCs) miR-122 expression is positively correlated with HCV RNA load and IFNγ and reversely with LPL expression in CHC patients. Presented, for the first time, in this study, the reverse correlation of miR-122 and LPL expression in liver; miR-122 and LPL seem to be important factors of CHC infection.

Predominance of genotype P[9]G3 in rotavirus gastroenteritis in Polish children.

INTRODUCTION: Rotavirus (RV) infection is the most common cause of gastroenteritis in children. This paper identifies the most common genotypes of rotaviruses isolated from children hospitalized with gastroenteritis and attempts to determine any relationship between infection with a certain rotavirus genotype. MATERIAL AND METHODS: The investigated group consisted of 68 consecutive children with rotavirus gastroenteritis (confirmed by an agglutination test). Rotavirus genotype was determined in stool samples obtained from each child. RESULTS: The P[9]VP4 genotype was observed in 41/61 positive samples (over 67.2%) that were permanently associated with the G3 VP7 genotype. Moreover, G3 was determined as the most commonly isolated G type (77.94%). As well as the P[9]G3 type, G3 was also found in the P[4] type (5 cases). Twenty-six out of 61 (42.6%) children in whom rotavirus genotype was determined were co-infected with pathogenic bacteria. No statistical correlation was observed between rotavirus P[9]G3 gastroenteritis and digestive tract co-infection with pathogenic bacteria (p > 0.05). Elevated ALT activity was found in 34/59 (57.6%) cases of rotavirus gastroenteritis. Elevated ALT serum level was found to correlate with P[9]G3 rotavirus genotype but concomitant infections did not. CONCLUSIONS: The most common genotype of rotaviruses observed in our group of children, P[9]G3, has rarely been described. Co-infection of the digestive tract with pathogenic bacteria and elevated serum ALT concentrations were found to be the most frequent phenomena. A correlation between P[9]G3 rotavirus genotype and elevated serum ALT level was found, but no significant relationship was identified between concomitant infections and P[9]G3 genotype.

Ribavirin priming has no beneficial effects for chronic hepatitis C patients.

AIM: The aim of this study is to assess the efficacy of an initial dose of ribavirin administered before a 48-week course of treatment with peg-IFN + ribavirin in treatment-naïve patients and in patients after previous failure of CHC treatment. MATERIAL AND METHODS: A total of 103 patients with chronic hepatitis C infected with genotype 1 HCV were qualified to the study. Study patients were randomised to receive one of two treatments: A- RBV for 4 weeks followed by combined therapy with peg-IFN alpha-2a +RBV for 48 weeks (n = 73), or B- combined therapy with peg-IFN alpha-2a +RBV for 48 weeks (n = 30). RESULTS: SVR 24 was observed in 44% patients in group A and in group 40% patients in group B (40%), p > 0.05. Comparing subgroups of the naive patients, it was found that the SVR24 value was higher in group A than group B (57% vs. 47%, p > 0.05). In the re-therapy subgroups, higher treatment response rates in patients not responding earlier was found in group A than group B (39% vs. 16%, p > 0.05). CONCLUSION: No significant advantage was found in the use of a priming method over a standard regimen. However, it could be recommended in patients with a total lack of response to peg-IFN and ribavirin when no other therapeutic options are available.

Decreased level of intracellular cholesterol in peripheral blood mononuclear cells is associated with chronic hepatitis C virus infection.

Peripheral blood mononuclear cells (PBMCs) constitute the main extrahepatic place of, hepatitis C virus (HCV) replication. We aimed to determine the impact of CHC infection and microRNA-, 122 expression on cholesterol expression in PBMCs. HCV RNA strand, intracellular cholesterol, HMGCoA, reductase and miR-122 expression in PBMC were determined in 54 CHC patients. The study shows that significant decrease of intracellular cholesterol level in PBMC (p=0.000000), accompanied by serum hypocholesterolemia is the characteristic feature of chronic hepatitis C infection. Although, microRNA-122 expression was detectable in PBMCs of CHC patients (52.5%), the alteration of intracellular cholesterol level was independent of miR-122 expression.

Coordinated increase of miRNA-155 and miRNA-196b expression correlates with the detection of the antigenomic strand of hepatitis C virus in peripheral blood mononuclear cells.

A tight relationship has been revealed between cellular microRNAs (miRNAs) and the course of hepatitis C virus (HCV) replication in human hepatoma cells. Although the detection of the antigenomic HCV RNA strand in peripheral blood mononuclear cells (PBMCs) has provided evidence for viral replication in PBMCs, no reports have shown how miRNAs are affected upon HCV RNA synthesis in PBMCs. The aim of the present study was to assess if and how the expression levels of miRNA-155 and miRNA-196b in PBMCs are related to HCV replication in PBMCs of chronic hepatitis C (CHC) patients. Supporting analyses were performed to evaluate the expression of precursor pri-miR-155 (BIC) and Dicer protein. The genomic and antigenomic HCV RNA strands in PBMCs were detected by strand-specific qRT-PCR. The expression levels of miRNAs, BIC RNA and Dicer protein were assayed on PBMCs by qRT-PCR and Western blotting, respectively. miRNA-155 and miRNA-196b were detected in all studied PBMC samples, but their levels varied according to the presence of the antigenomic HCV RNA strand in PBMCs. Increased expression levels of miRNA-155 and miRNA-196b were associated with the presence of the antigenomic HCV RNA strand in PBMCs. In this group of patients higher frequency of BIC RNA and Dicer protein detection was also found. This study demonstrates that HCV RNA replication in PBMCs of CHC patients is connected with the increased and coordinated expression of miRNA-155 and miRNA-196b.

Mevalonate pathway modulation is associated with hepatitis C virus RNA presence in peripheral blood mononuclear cells.

Peripheral blood mononuclear cells (PBMC) constitute the main extrahepatic reservoir of hepatitis C virus (HCV). Lipid metabolism of host seems to play important role in HCV infection. The relationship between HCV presence in PBMC and the expression of mevalonate pathway has not been elucidated. The aim of this study was to investigate the association between mevalonate pathway and HCV RNA presence in PBMC after anti-HCV treatment. 67 serum and corresponding PBMC samples were collected from patients at the end of interferon alpha and ribavirin treatment. Serum total cholesterol, HDL-C and LDL-C fractions, triglycerides, as well as intracellular cholesterol and expression level of HMG-CoA reductase, geranylgeranyl pyrophosphate synthase in PBMC were measured and matched for the HCV RNA presence or absence in sera/PBMC. HCV RNA elimination from sera and PBMC was associated with higher serum cholesterol (118.5mg/dL) and LDL-C (66.42mg/dL) levels, compared to the group, where HCV RNA was detected only in PBMC (100.94 and 53.22mg/dL) or the group, where HCV RNA was found in both sera and PBMC (86.79 and 43.79mg/dL) after treatment. Increased expression of geranylgeranyl pyrophosphate synthase was found in the majority of PBMC samples that harbored HCV RNA after elimination of HCV RNA from sera. The expression of mevalonate pathway after antiviral treatment seems to be modulated depending on HCV RNA status in peripheral blood mononuclear cells.

Angiotensin receptors in hormone-independent prostate cancer cell line DU145: presence of two variants of angiotensin type 1 receptor.

BACKGROUND: Recently, the involvement of the renin-angiotensin system in the proliferation of prostate cancer has been suggested. There is increasing evidence that angiotensin II receptor type 1 (AT1) is expressed in a variety of cancer cells and tissues and may have a role in tumor growth, angiogenesis, and invasive activity of malignant lesions in vivo. The implications of data referring to an angiotensin receptor in hormone-independent human prostate cancer DU145 cells are unclear. Angiotensin II has been shown to inhibit the proliferation of DU145 cell lines. However, it is known that AT1 stimulates cell proliferation and that angiotensin II receptor type 2 (AT2) induces apoptosis and inhibits cell proliferation. MATERIAL/METHODS: The aim of our study was to investigate, by means of immunohistochemical and reverse transcriptase polymerase chain reaction assays, the type of angiotensin II receptor that is present in DU145 cells. RESULTS: In DU145-derived complementary deoxyribonucleic acid (cDNA), a polymerase chain reaction assay revealed 2 AT1-specific PCR products (93 bp and 126 bp). DU145-derived cDNA did not reveal AT2 expression at a level sufficient for detection by PCR. In cultured cells, immunohistochemical testing revealed a positive reaction in cultures immunostained with anti-AT1 antibody but not in those immunostained with AT2 antibody. CONCLUSIONS: The inhibitory effect of angiotensin II on the proliferation of DU145 cells is exerted via AT1. It is possible that presence of 2 variants of AT1 in that cancer cell line is essential to produce the biological effects of angiotensin.

Gene modulation associated with inhibition of liver regeneration in hepatitis B virus X transgenic mice.

AIM: To analyze the modulation of gene expression profile associated with inhibition of liver regeneration in hepatitis B X (HBx)-expressing transgenic mice. METHODS: Microarray technology was performed on liver tissue obtained from 4 control (LacZ) and 4 transgenic mice (HBx-LacZ), 48 h after partial hepatectomy. The significance of the normalized log-ratios was assessed for each gene, using robust t-tests under an empirical Bayes approach. Microarray hybridization data was verified on selected genes by quantitative PCR. RESULTS: The comparison of gene expression patterns showed a consistent modulation of the expression of 26 genes, most of which are implicated in liver regeneration. Up-regulated genes included DNA repair proteins (Rad-52, MSH6) and transmembrane proteins (syndecan 4, tetraspanin), while down-regulated genes were connected to the regulation of transcription (histone deacetylase, Zfp90, MyoD1) and were involved in the cholesterol metabolic pathway and isoprenoid biosynthesis (farnesyl diphosphate synthase, Cyp7b1, geranylgeranyl diphosphate synthase, SAA3). CONCLUSION: Our results provide a novel insight into the biological activities of HBx, implicated in the inhibition of liver regeneration.

[Role of lipid rafts in hepatitis C virus life cycle]. / Rola tratw lipidowych w cyklu zyciowym wirusa zapalenia watroby typu C.

Although the mechanism of HCV RNA replication is still poorly characterized, we know that HCV active replication complex contains membrane structures. Biochemical analysis showed that these membrane structures were resistant to nonionic detergents at low temperature and contained caveolin-2, characteristic of lipid rafts. Further analysis suggested that lipid rafts protect HCV replication complex against degradation. Lipid rafts provide also a mechanism of how HCV may modulate cellular processes, what can lead to disruption of antiviral immune response and then to chronic infection. Association hepatitis C virus with lipid rafts can be also a source of new antiviral therapies.

[Late spontaneous elimination of HCV-RNA from mononuclear cells derived from peripheral blood in patients with chronic hepatitis C]. / Pózna samoistna eliminacja HCV-RNA z komórek jednojadrzastych krwi obwodowej po skutecznym leczeniu przewleklego zapalenia watroby typu C.

THE AIM: To follow persistence of HCV-RNA in PBMC in patients with chronic hepatitis C (CHC). To estimate the influence of this phenomenon on the cellular immune response of peripheral blood lymphocytes. MATERIAL AND METHODS: 8 HCV-RNA in PBMC positive children, with undetectable serum HCV-RNA after antiviral treatment, have been examined every 2-3 years. The amount of IFN-gamma, IL-12 and IL-18 secreted by PBMC obtained from the children after stimulation with phytohemagglutinin (PHA) was measured. RESULTS: Spontaneous elimination of HCV-RNA from PBMC in 2 to 6 years after treatment was found in all children. In two children HCV-RNA detectable both in serum and in PBMC, without recurrence of hepatitis, was found in single examination. PBMC containing HCV-RNA secreted more IFN-gamma than PBMC lacking it (1221 +/- 458 pg/ml vs. 651 +/- 147 pg/ml; p=0.009, similar correlation was revealed with the regard of IL-12: 21.8 +/- 12.3 pg/ml vs. 5,6 +/- 3,3 pg/ml respectively; p=0.009. Production and release of IL-18 were not correlated with HCV-RNA persistence (p=0.12). CONCLUSION: Patients with CHC and persistence of HCV-RNA in PBMC require longitudinal follow-up in the respect of possible reseroconversion. PBMC containing HCV-RNA reveal enhanced cellular immune response, which most probably effects in spontaneous elimination of the virus.

A competitor DNA template for the molecular quantification of the hepatitis B virus.

The molecular determination of viral load in the serum represents the most valuable prognostic marker of HBV infection. In this paper, a new molecular assay for the quantitative measurement of HBV presence is described. It is based on PCR performing with a HBV-specific competitor DNA template. For the construction of the DNA template, a HBV DNA-originated 436 bp DNA fragment was modified by introducing a 110 bp deletion and cloned into pUC19. The resulting vector serves as the competitor DNA template in the competitive PCR. Post-PCR, the competitor DNA generates an amplified fragment of 306 bp; it could be easily distinguished from the product generated from the viral-originated DNA product (416 bp) when the same primers are used. The quantitative ratio between the two products enables the quantitative determination of viral load. The range of the HB-PCR assay is from 3 x 10(4)to 6 x 10(10) particles/ml. A serum HBV load determination performed by HB-PCR assay indicated a close correlation with the results of the Quantiplex HBV DNA assay (bDNA). The HB-PCR assay is cheap, reliable and easy to use in any laboratory working with PCR methods.

[HLA class II alleles and response to hepatitis C treatment with interferon alpha2b]. / Antygeny zgodnosci tkankowej klasy II a odpowiedz na leczenie interferonem-alpha 2b przewleklego zapalenia watroby typu C.

BACKGROUND/AIMS: The mechanisms of humoral immunological response in chronic hepatitis C are not fully understood. It would be interesting to correlate HLA II alleles with the therapeutic response to interferon alfa2b treatment in patients with chronic hepatitis C. Such correlation has not yet been described. The purpose of the study was to correlate the presence of HLA alleles in chronic hepatitis C patients with results of interferon-alfa2b therapy. METHODS: We assessed HCV-RNA presence in serum by RT-PCR and HLA-DR B*1/alleles by PCR-SSP for locus I and II in 54 patients with chronic hepatitis C. All patients were treated with interferon alfa2b for six month. Results of the therapy were evaluated 18 months after the end of treatment. RESULTS: Based on the treatment results (TR--therapeutic response) patients were retrospectively qualified into three groups: sustained responders (SR)--18.6% (10/54), relapsers (R)--48.1% (26/54), non responders (NR)--33.3% (18/54). Allele DRB*1 07 and allele DRB1* 13 were more frequent among patients in NR and TR group, respectively. Observed differences were not statistically significant (p > 0.05). CONCLUSION: In the group of 54 adults with chronic hepatitis C no correlation between the presence of HLA-DRB1* alleles and response to interferon alfa2b therapy was found.