Down-regulation of IRES containing 5'UTR of HCV genotype 3a using siRNAs

<p>Abstract</p> <p>Background</p> <p>Hepatitis C virus (HCV) is a major causative agent of liver associated diseases leading to the development of hepatocellular carcinoma (HCC) all over the world and genotype-3a responsible for most of the cases in Pakistan. Due to the limited efficiency of current chemotherapy of interferon-α (IFN-α) and ribavirin against HCV infection alternative options are desperately needed out of which the recently discovered RNAi represent a powerful silencing approach for molecular therapeutics through a sequence-specific RNA degradation process to silence virus infection or replication. HCV translation is mediated by a highly conserved internal ribosome entry site (IRES) within the 5'UTR region making it a relevant target for new drug development.</p> <p>Materials and methods</p> <p>The present study was proposed to assess and explore the possibility of HCV silencing using siRNA targeting 5'UTR. For this analysis full length HCV 5'UTR of HCV-3a (pCR3.1/5'UTR) was tagged with GFP protein for <it>in vitro </it>analysis in Huh-7 cells. siRNA targeting 5'UTR were designed, and tested against constructed vector in Huh-7 cell line both at RNA and Protein levels. Furthermore, the effect of these siRNAs was confirmed in HCV-3a serum infected Huh-7 cell line.</p> <p>Results</p> <p>The expression of 5'UTR-GFP was dramatically reduced both at mRNA and protein levels as compared with Mock transfected and control siRNAs treated cells using siRNAs against IRES of HCV-3a genotype. The potential of siRNAs specificity to inhibit HCV-3a replication in serum-infected Huh-7 cells was also investigated; upon treatment with siRNAs a significant decrease in HCV viral copy number and protein expression was observed.</p> <p>Conclusions</p> <p>Overall, the present work of siRNAs against HCV 5'UTR inhibits HCV-3a expression and represents effective future therapeutic opportunities against HCV-3a genotype.</p

microRNA-122 stimulates translation of hepatitis C virus RNA

Hepatitis C virus (HCV) is a positive strand RNA virus that propagates primarily in the liver. We show here that the liver-specific microRNA-122 (miR-122), a member of a class of small cellular RNAs that mediate post-transcriptional gene regulation usually by repressing the translation of mRNAs through interaction with their 3′-untranslated regions (UTRs), stimulates the translation of HCV. Sequestration of miR-122 in liver cell lines strongly reduces HCV translation, whereas addition of miR-122 stimulates HCV translation in liver cell lines as well as in the non-liver HeLa cells and in rabbit reticulocyte lysate. The stimulation is conferred by direct interaction of miR-122 with two target sites in the 5′-UTR of the HCV genome. With a replication-defective NS5B polymerase mutant genome, we show that the translation stimulation is independent of viral RNA synthesis. miR-122 stimulates HCV translation by enhancing the association of ribosomes with the viral RNA at an early initiation stage. In conclusion, the liver-specific miR-122 may contribute to HCV liver tropism at the level of translation

Behavioural Lifestyle Intervention Study (BLIS) in patients with type 2 diabetes in the United Arab Emirates: A randomized controlled trial

BACKGROUND: Lifestyle modification is a cornerstone of the management of type 2 diabetes. However, in the United Arab Emirates, a country where type 2 diabetes is highly prevalent, non-compliance with a healthy lifestyle has been reported in many diabetic Emirati patients. The use of behavioural theories in lifestyle counselling is believed to facilitate behavioural changes, nevertheless, there are no published data regarding the use of structured behavioural lifestyle programmes tailored to suit Emirati culture. The primary objective of this study was to develop a behavioural lifestyle programme and to evaluate its effectiveness in improving glycaemic control in Emirati patients with type 2 diabetes. METHODS: The Behavioural Lifestyle Intervention Study (BLIS) was a translational randomized controlled trial in which patients (n = 35) were randomly assigned to the intervention or control group. Patients in the intervention group went through a six-month behavioural lifestyle programme composed of 8 sessions, whereas patients in the control group received standard care. Cognitive behavioural theory was the underpinning theory for the lifestyle intervention. HbA1c levels were the trial’s primary outcome measure, and the main dietary factor targeted for change was carbohydrate intake. They were measured at baseline, 3 months and 6 months and were assessed using one-way ANOVA at a significance level of P < 0.05. All of the patients were then followed up at 1 year on all outcome measures. RESULTS: At 6 months, the HbA1c levels of the patients (n = 18) in the intervention group were significantly reduced (−1.56 ± 1.81, P < 0.05), whereas no significant change was observed in the patients of the control group. Similarly, both carbohydrate intake from cereals and total carbohydrate intake (in grams) were reduced (p < 0.05) in the intervention group, by 32.92 ± 54.34 and 20.94 ± 56.73, respectively. At 1 year, the patients in the intervention group maintained a significant reduction in HbA1c levels (−1.12 ± 1.46, p < 0.05), whereas no change was observed in the control group. CONCLUSION: The behavioural lifestyle intervention programme was effective in improving glycaemic control and compliance with carbohydrate intake goals in Emirati patients with type 2 diabetes. Larger randomized controlled trials are needed to validate these results and to identify key behavioural strategies that will improve compliance to lifestyle modifications in real life. TRIAL REGISTRATION: Clinicaltrials.gov trial identifier NCT0238693

A922 Sequential measurement of 1 hour creatinine clearance (1-CRCL) in critically ill patients at risk of acute kidney injury (AKI)

Meeting abstrac

Vampires in the village Žrnovo on the island of Korčula: following an archival document from the 18th century

Središnja tema rada usmjerena je na raščlambu spisa pohranjenog u Državnom arhivu u Mlecima (fond: Capi del Consiglio de’ Dieci: Lettere di Rettori e di altre cariche) koji se odnosi na događaj iz 1748. godine u korčulanskom selu Žrnovo, kada su mještani – vjerujući da su se pojavili vampiri – oskvrnuli nekoliko mjesnih grobova. U radu se podrobno iznose osnovni podaci iz spisa te rečeni događaj analizira u širem društvenom kontekstu i prate se lokalna vjerovanja.The main interest of this essay is the analysis of the document from the State Archive in Venice (file: Capi del Consiglio de’ Dieci: Lettere di Rettori e di altre cariche) which is connected with the episode from 1748 when the inhabitants of the village Žrnove on the island of Korčula in Croatia opened tombs on the local cemetery in the fear of the vampires treating. This essay try to show some social circumstances connected with this event as well as a local vernacular tradition concerning superstitions