Chloroquine Inhibits Dengue Virus Type 2 Replication in Vero Cells but Not in C6/36 Cells

Dengue viruses are the most important arthropod-borne viruses in terms of morbidity and mortality in the world. Since there is no dengue vaccine available for human use, we have set out to investigate the use of chloroquine as an antiviral drug against dengue. Chloroquine, an amine acidotropic drug known to affect intracellular exocytic pathways by increasing endosomal pH, was used in the in vitro treatment of Vero and C6/36 cells infected with dengue virus type 2 (DENV-2). Real-time RT-PCR and plaque assays were used to quantify the DENV-2 load in infected Vero and C6/36 cells after chloroquine treatment. Our results showed that a dose of 50 μg/ml of chloroquine was not toxic to the cells and induced a statistically significant inhibition of virus production in infected Vero cells when compared to untreated cells. In C6/36 cells, chloroquine does not induce a statistically significant difference in viral replication when compared to untreated cells, showing that this virus uses an unlikely pathway of penetration in these cells, and results were also confirmed by the plaque assay (PFU). These data suggest that the inhibition of virus infection induced by chloroquine is due to interference with acidic vesicles in mammalian cells

Structural characterization of a novel peptide with antimicrobial activity from the venom gland of the scorpion Tityus stigmurus: Stigmurin

AbstractA new antimicrobial peptide, herein named Stigmurin, was selected based on a transcriptomic analysis of the Brazilian yellow scorpion Tityus stigmurus venom gland, an underexplored source for toxic peptides with possible biotechnological applications. Stigmurin was investigated in silico, by circular dichroism (CD) spectroscopy, and in vitro. The CD spectra suggested that this peptide interacts with membranes, changing its conformation in the presence of an amphipathic environment, with predominance of random coil and beta-sheet structures. Stigmurin exhibited antibacterial and antifungal activity, with minimal inhibitory concentrations ranging from 8.7 to 69.5μM. It was also showed that Stigmurin is toxic against SiHa and Vero E6 cell lines. The results suggest that Stigmurin can be considered a potential anti-infective drug

Antiviral activity of chloroquine against Dengue virus type 2 replication in Aotus Monkeys

FAPESP (São Paulo Foundation for Research; Grant no. 05/04450-4) ; FAPESP fellowship (Grant no. 06/55789-4).University of São Paulo. School of Medicine of Ribeirão Preto. Department of Internal Medicine. São Paulo, SP, Brazil / University of São Paulo. School of Medicine of Ribeirão Preto. Program of Graduate Studies on Applied Microbiology and Immunology. São Paulo, SP, Brazil / University of São Paulo. School of Medicine of Ribeirão Preto. Virology Research Center. São Paulo, SP, Brazil.University of São Paulo. School of Medicine of Ribeirão Preto. Department of Internal Medicine. São Paulo, SP, Brazil / University of São Paulo. School of Medicine of Ribeirão Preto. Program of Graduate Studies on Applied Microbiology and Immunology. São Paulo, SP, Brazil / University of São Paulo. School of Medicine of Ribeirão Preto. Virology Research Center. São Paulo, SP, Brazil.Ministério da Saúde. Secretaria de Vigilância em Saúde. Instituto Evandro Chagas. Centro Nacional de Primatas. Ananindeua, PA, Brasil.Ministério da Saúde. Secretaria de Vigilância em Saúde. Instituto Evandro Chagas. Centro Nacional de Primatas. Ananindeua, PA, Brasil.University of São Paulo. School of Medicine of Ribeirão Preto. Department of Internal Medicine. São Paulo, SP, Brazil / University of São Paulo. School of Medicine of Ribeirão Preto. Program of Graduate Studies on Applied Microbiology and Immunology. São Paulo, SP, Brazil / University of São Paulo. School of Medicine of Ribeirão Preto. Virology Research Center. São Paulo, SP, Brazil.Dengue virus (DENV) of the Flaviviridae family is a single positive-stranded RNA virus that is transmitted by Aedes aegypti and Aedes albopictus mosquitoes. The objective of this study was to investigate the use of chloroquine (CLQ) as an antiviral drug against dengue virus in monkeys. To analyze the action of the drug in vivo, nonhuman primates groups (Aotus azarai infulatus) were inoculated with a subcutaneous injection of a virulent strain of DENV-2, treated and untreated CLQ. Blood hematological, viremia, and serum biochemical values were obtained from 16 DENV-2-inoculated, treated and untreated; four received only CLQ and one mock-infected Aotus monkeys. Monkey serum samples (day 0–10 post-inoculation) were assayed by reverse transcription polymerase chain reaction and Cytometric Bead Array for determination of viremia and inflammatory cytokines, respectively. Additionally, body temperature and activity levels were determined. In the present work, CLQ was effective on replication of DENV-2 in Aotus monkeys; a time viremia reduction was observed compared with the controls. The concentration of tumor necrosis factor alpha and interferon gamma in the serum of the animals had a statistically significant reduction in the groups treated with CLQ after infection compared with the controls. A significant decrease in systemic levels of the liver enzyme aspartate aminotransferase (AST) was also observed in the animals treated with CLQ after infection compared with the controls. These results suggest that CLQ interferes in DENV-2 replication in Aotus monkeys

Human herpesvirus 8 (HHV-8) detected by nested polymerase chain reaction (PCR) in HIV patients with or without Kaposi's sarcoma. An analytic cross-sectional study

CONTEXT AND OBJECTIVE: Kaposi's sarcoma (KS) is a common neoplastic disease in AIDS patients. The aim of this study was to evaluate the frequency of human herpesvirus 8 (HHV-8) infection in human immunodeficiency virus (HIV)-infected patients, with or without KS manifestations and correlate HHV-8 detection with KS staging. DESIGN AND SETTING: Analytic cross-sectional study conducted in a public tertiary-level university hospital in Ribeirão Preto, São Paulo, Brazil. METHODS: Antibodies against HHV-8 lytic-phase antigens were detected by means of the immunofluorescence assay. HHV-8 DNA was detected in the patient samples through a nested polymerase chain reaction (nested PCR) that amplified a region of open reading frame (ORF)-26 of HHV-8. RESULTS: Anti-HHV-8 antibodies were detected in 30% of non-KS patients and 100% of patients with KS. Furthermore, the HHV-8 DNA detection rates observed in HIV-positive patients with KS were 42.8% in serum, 95.4% in blood samples and 100% in skin biopsies; and in patients without KS, the detection rate was 4% in serum. Out of the 16 serum samples from patients with KS-AIDS who were classified as stage II, two were positive (12.5%); and out of the 33 samples from patients in stage IV, 19 (57.6%) were positive. CONCLUSION: We observed an association between HHV-8 detection and disease staging, which was higher in the serum of patients in stage IV. This suggests that detection of HHV-8 DNA in serum could be very useful for clinical assessment of patients with KS and for monitoring disease progression

Bothrops jararaca

Bothrops jararaca (BJ) and Bothrops erythromelas (BE) are viper snakes found in South-Southeast and Northeast regions of Brazil, respectively. Snake venoms are bioactive neurotoxic substances synthesized and stored by venom glands, with different physiological and pharmacological effects, recently suggesting a possible preference for targets in cancer cells; however, mechanisms of snakes have been little studied. Here, we investigated the mechanism responsible for snake crude venoms toxicity in cultured cervical cancer cells SiHa and HeLa. We show that BJ and BE snake crude venoms exert cytotoxic effects to these cells. The percentage of apoptotic cells and cell cycle analysis and cell proliferation were assessed by flow cytometry and MTT assay. Detection of mitochondrial membrane potential (Rhodamine-123), nuclei morphological change, and DNA fragmentation were examined by staining with DAPI. The results showed that both the BJ and BE venoms were capable of inhibiting tumor cell proliferation, promoting cytotoxicity and death by apoptosis of target SiHa and HeLa cells when treated with BJ and BE venoms. Furthermore, data revealed that both BJ venoms in SiHa cell promoted nuclear condensation, fragmentation, and formation of apoptotic bodies by DAPI assay, mitochondrial damage by Rhodamine-123, and cell cycle block in the G1-G0 phase. BJ and BE venoms present anticancer potential, suggesting that both Bothrops venoms could be used as prototypes for the development of new therapies

The crab heparin-like compound exhibits a strong inhibitory effect on infections by dengue virus-2

This study was supported by Federal University of Rio Grande do Norte.Federal University of Rio Grande do Norte - Campus Universitário. Department of Biochemistry. Natal, RN, Brazil.Federal University of Rio Grande do Norte. Postgraduate Program in Health Sciences. Natal, RN, Brazil.Ministério da Saúde. Secretaria de Vigilância em Saúde. Instituto Evandro Chagas. Programa de Pós-Graduação em Virologia. Ananindeua, PA, Brasil.Federal University of Rio Grande do Norte - Campus Universitário. Department of Biochemistry. Natal, RN, Brazil.Federal University of Rio Grande do Norte. Department of Clinical Analysis and Toxicology. Natal, RN, Brazil.Federal University of Rio Grande do Norte - Campus Universitário. Department of Biochemistry. Natal, RN, Brazil.Federal University of Campina Grande. Center of Education and Health. Cuité, PB, Brazil.Federal University of Rio Grande do Norte - Campus Universitário. Department of Biochemistry. Natal, RN, Brazil.Background: According to the World Health Organization (WHO), two-fifths of the world population is at risk of infection by DENV. There are no safe and effective vaccines estab-lished. Sulfated glycosaminoglycans such as heparin, used as anticoagulants, inhibit the initial step of dengue viral replication. Recently, an isolated heparin analogue Goniopsis cruentata (cCTH) has presented a low anticoagulant effect with reduced bleeding risk. Methods: The antiviral activity of cCTH and heparin compounds against DENV-2 in Vero cell culture was determined by quantitative RT-PCR (qRT-PCR) and titration. For this, four trials were carried out: treatment of the cells for 2 h before viral inoculation, concomitant viral inoculation treatment, treatment after viral inoculation and virucidal assay. Subsequently, the culture superna-tants were collected for periods of 24, 48 and 72 h. Results: Our results demonstrated that cCTH and heparin showed antiviral activity against DENV-2. Conclusion: These data suggest that both compounds prevented viral replication in cultured Vero cells