Adalberto Sánchez

Caracterización genómica de la integración in vitro del VIH-1 en células mononucleares de sangre periférica, macrófagos y células T de Jurkat

Introduction: Most of the infected host cell genome is available for retroviral integration; however, it has been proposed that this process does not occur at random and depends upon each type of retrovirus. Objective: The objective is to identify and characterize differences in human genome regions of peripheral blood mononuclear cells, macrophages and Jurkat T cells in which integration of HIV-1 occurs. Material and Methods: Three hundred human DNA genome sequences, previously deposited in the GenBank, were selected at random. Using program BLAST, only 264 of them were included in the study because relevant information about chromosomal position, associated genes, repetitive sequences, number of CpG islands and average replication time was available; these sequences were exported to other data bases for analysis. Results: 53% (140/264) of integrations were located on G bands. 70.45% of provirus was located in human genes and the rest was located in repetitive elements. In general the integration site selection was correlated with genomics and structural characteristics of cell chromatin including Alu-Sx and L1 sequences, gene and CpG island densities, remodeling of chromatin, and replication time. All of them would influence the efficient interaction between the pre-integration complex and target cell genomes. Conclusion: It was determined that HIV-1 integration in target cellular genomes would be conditioned by differential characteristics of associated chromatin and by epigenetic processes that would influence the selection of integration sites.

Human T-Lymphotropic virus (HTLV) type I in vivo integration in oral keratinocytes

Although the infection of HTLV-1 to cell components of the mouth have been previously reported, there was not until this report, a detailed study to show the characteristics of such infection. From 14 Tropical Spastic Paraparesis/HTLV-1-Associated Myelopathy (HAM/TSP) patients and 11 asymptomatic carrier individuals (AC) coming from HTLV-1 endemic areas of southwest Pacific of Colombia, infected oral mucosa cells were primary cultured during five days. These cell cultures were immunophenotyped by dual color fluorescence cell assortment using different lymphocyte CD markers and also were immunohistochemically processed using a polyclonal anti-keratin antibody. Five days old primary cultures were characterized as oral keratinocytes, whose phenotype was CD3- /CD4-/CD8-/CD19-/CD14-/CD45-/A575-keratin+. From DNA extracted of primary cultures LTR, pol, env and tax HTLV-1 proviral DNA regions were differentially amplified by PCR showing proviral integration. Using poly A+ RNA obtained of these primary cultures, we amplify by RT-PCR cDNA of tax and pol in 57.14% (8/14) HAM/TSP patients and 27.28% (3/11) AC. Tax and pol poly A+ RNA were expressed only in those sIgA positive subjects. Our results showed that proviral integration and viral gene expression in oral keratinocytes are associated with a HTLV-1 specific local mucosal immune response only in those HTLV-1 infected individuals with detectable levels of sIgA in their oral fluids. Altogether the results gave strong evidence that oral mucosa infection would be parte of the systemic spreading of HTLV-1 infection.