ABSTRACT
Provirus mutations of human T-lymphotropic virus 1 (HTLV-1), mostly the lack of the 5= long terminal repeat (LTR) genomic region, have been described and associated with severe adult T cell leukemia/lymphoma (ATLL), non-sense point mutations with low proviral load, and Western blotting indeterminate results. Until now, no information concerning provirus mutations of HTLV-2 and its consequences, as well as those of HTLV-1/2 in HIV-coinfected individuals, had been described. Therefore, we searched for these mutations in provirus samples of 44 HIV/HTLV-1- and 25 HIV/HTLV-2-coinfected individuals. Using protocols well established for amplification and sequencing of segments of the LTR, env, and tax regions, we searched for defective type 1 particles that retain LTRs and lack internal sequences and type 2 particles that lack the 5=LTR region. In addition, using as references the prototypes ATK (HTLV-1) and Mo (HTLV-2), we searched for point mutations in the LTR and synonyms and nonsynonymous mutations and non-sense mutations in env and tax regions. Defective HTLV-1 and HTLV-2 provirus type 1 or 2 was detected in 31.8% of HIV/HTLV-1- and 32.0% of HIV/HTLV-2-coinfected individuals. Synonymous and nonsynonymous mutations were identified mostly in HTLV-2 and associated with lower levels of specific antibodies. No non-sense mutations that resulted in premature termination of Env and Tax proteins were detected. On the contrary, mutation in the stop codon of Tax2a produced a long protein characteristic of the HTLV-2c subtype. The clinical significance of these mutations in coinfected individuals remains to be defined, but they confirmed the lower sensitivity of serological and molecular diagnostic tests in HIV/HTLV-1/2 coinfections. IMPORTANCE HTLV-1 and HTLV-2 are endemic to Brazil, and they have different effects in HIV/AIDS disease progression. HIV/HTLV-1 has been described as accelerating the progression to AIDS and death, while HIV/HTLV-2 slows the progression to AIDS. Provirus mutations of HTLV-1 were implicated in severe leukemia development and in problems in the diagnosis of HTLV-1; in contrast, provirus mutations of HTLV-2 had not been confirmed and associated with problems in HTLV-2 diagnosis or disease outcome. Nevertheless, data obtained here allowed us to recognize and understand the false-negative results in serologic and molecular tests applied for HTLV-1 and HTLV-2 diagnosis. Defective proviruses, as well as synonymous and nonsynonymous mutations, were associated with the diagnosis deficiencies. Additionally, since HIV-1 and HTLV-1 infect the same cells (CD4 positive), the production of HIV-1 pseudotypes with HTLV-1 envelope glycoprotein during HIV/HTLV-1 coinfection cannot be excluded. Defective provirus of HTLV-2 and Tax2c is speculated to influence progression to AIDS. (AU)
Subject(s)
Human T-lymphotropic virus 1 , Human T-lymphotropic virus 2 , Acquired Immunodeficiency Syndrome , HIV , Proviruses , Coinfection , MutationABSTRACT
Objective To investigate the expression changes of provirus integration site 1 for moloney murine leukemia virus(Pim-1) gene in damaged neurons in vitro and related molecular basis of neurotrophic factors regulating Pim-1 expression and promoting the neurite regeneration of damaged neurons. Methods Neuro-2a(N-2a)cells were induced into neuron-like N-2a(N-2a-N) cells by retinoic acid,the proliferation of N-2a cells was inhibited by deferoxamine mesylate (DFO), and N-2a-N cells were injured by acrylamide. The N-2a-N cells were divided into normal control group, injury group, ciliary neurotrophic factors (CNTF) group and neuritin (Nrn1) group, with four samples in each group. The phenotype of N-2a cells and the expression of Pim-1 protein in N-2a cells were detected by immunofluorescence cytochemistry, and the expression of Pim-1 in each group was detected by Real-time PCR and Western blotting. Western blotting was used to detect the expression changes of relevant molecules involving in regulating activity of Pim-1, cells survival, apoptosis and axonal regeneration. Results Cell immunofluorescence showed that N-2a-N cells had neuronal phenotype to express β-Ⅲ tubulin and neurofilament-200, and Pim-1 protein was expressed in N-2a-N cells. N-2a cell proliferation was effectively inhibited by 50 μmol/ L DFO, and N-2a-N cell damage model was established by 1 mmol/ L acrylamide. Pim-1 gene expression showed a tendency of first decreasing, then increasing, and then decreasing after N-2aN cells were injured. Compared with the injury group, the proportion of the longest neurite in CNTF group and Nrn1 group increased significantly, the expressions of intracellular signal transducers extracellular regulated protein kinase 1/ 2 (ERK1/ 2), phosphorylated extracellular regulated protein kinase 1/ 2 (p-ERK1/ 2), signal transducers and activators of transcription 3(STAT3), phosphorylated signal transducers and activators of transcription 3 (p-STAT3), and Pim-1 were up-regulated, the expressions of apoptosis-related molecules cleaved Caspase-3 and Bax were down-regulated, the expression of anti-apoptosis-related molecule Bcl-2 was up-regulated, so the growth-associated protein 43 (GAP-43) protein involved neurite regeneration. Conclusion There is a need to repair damaged N-2a-N cells by overexpressing the Pim-1 gene. CNTF and Nrn1 can activate the ERK1/ 2 and STAT3 signaling pathways of damaged N-2a-N cells, and then up-regulate the expression of Pim-1 and GAP-43,and then promote cell neurite regeneration.
ABSTRACT
The present experiment was designed to observe the genetic variation of equine infectious anemia virus (EIAV) envelop gp 90 gene in infected horse. One horse was infected experimentally with P337-V70 strain and showed no clinical signs after being infected at twice with the same virus strain. Seventeen proviral sequences covering principal neutralizing domain (PND) of EIAV gp 90 gene were obtained from the buffy coat and liver of the horse through PCR amplification and cloning. Comparative analysis of the sequences revealed that some sequences contained the nucleotide insertions in the PND region. The insertions might be generated by direct repeat and strand displacement of sequence segment in their PND gene, showing different lenghts.
ABSTRACT
Objective In order to evaluate the therapeutic effect, to screen effective drugs for AIDS, and to find out the changes of patient's condition, it is necessary to develop an objective method to quantitate the levels of viral load of the patients before and after treatment.Method Differential PCR(D PCR) was used to co amplify the target HIV 1 gag gene and reference template ? globin gene, so as to determine the level of HIV replication quantitatively. The levels of HIV DNA in the peripheral blood mononuclear cells of 8 patients with AIDS were determined quantitatively.Results The levels of the provirus of the 8 patients ranged from 0 508 2 210 copies/?g DNA, there were differences in the levels of replication and integration. Conclusion It is the authors′ opinion that this method is easy to control with an advantage of reliability, quickness and reproducibility, It is suitable to measure clinical specimens, and provides an objective evidence for the evaluation of responses to therapeutic intervention.
ABSTRACT
Adult T-cell leukemia/lymphoma (ATLL) and cutaneous T-cell lymphoma (CTCL) have similar clinieopathology and immunophenotype, in order to differentiate the two diseases, 4 eases of ATLL associated cutaneous lesions and 18 eases of CL were studied for elinieopathology, immunophenotype and HTLV-I provirus DNA. Two eases of actinic reticuloid and two eases of lymphocytic infiltration of the skin were selected as negative control. The results showed that four patients of ATLL manifested cutaneous lesions, at same time, they had additional systemic diseases, such as generalized lymphadenopathy, increased levels of LDH and IL-2R, rosette-like cells in their peripheral blood and abnormal bone marrow. The HTLV-I provirus DNA was detected in the peripheral blood, bone marrow, cutaneous lesions and lymph node biopsy specimens of the four patients by PCR amplification of specific HTLV-I DNA fragment. 18 cases of CL were negative for HTLV-I. The study indicates that ATLL may be diagnosed if the patients are associated with CTCL-Iike cutaneous lesions, characteristic histopathological pattern and immunophenotype, rosette-like cells in the peripheral blood and positive HTLV-I provirus DNA.