ABSTRACT: Long-Term Programming of Antigen-Specific Immunity from Gene Expression Signatures in the PBMC of Rhesus Macaques Immunized with an SIV DNA Vaccine
Project description:This study describes differential miRNA expression in intact colon tissue during acute SIV infection of rhesus macaques. Nine miRNAs were found to be significantly affected by infection, with 5 down-regulated and 4 up-regulated miRNAs. The expression of one upregulated miRNA was further characterized and found to be significantly elevated specifically in response to SIV replication and not immune activation/inflammation accompanying SIV infection.
Project description:The primary objective of this study was to evaluate response to a SIV DNA-based vaccine that was adminstered via vivo electroporation (EP) in rhesus macaques to further understand the molecular correlates of protection against SIV. In this study, rhesus macaques were immunized with a DNA vaccine including individual plasmids encoding SIV gag, env, and pol alone, or in combination with a molecular adjuvant, plasmid DNA expressing the chemokine ligand 5 (RANTES), followed by EP. At eight month post-vaccination, animals were challenged with SIV. Standard immunological assays, flow-based activation analysis without ex vivo restimulation and high-throughput gene expression analysis were performed to determine the host response to each vaccine regimen.
Project description:This study describes differential miRNA expression in intact colon tissue during acute SIV infection of rhesus macaques. Nine miRNAs were found to be significantly affected by infection, with 5 down-regulated and 4 up-regulated miRNAs. The expression of one upregulated miRNA was further characterized and found to be significantly elevated specifically in response to SIV replication and not immune activation/inflammation accompanying SIV infection. We performed TaqMan Low Density Array based high throughput miRNA analysis on intact colon tissue from 10 acutely SIV-infected and 5 uninfected control macaques. All SIV-infected animals were inoculated intravenously with 100TCID50 of SIV. Out of the ten, one animal each was at 7, 8 and 10DPI (days post infection), 3 each at 13 and 21DPI, and 1 at 29DPI. microRNA reverse transcription and preamplification was performed according to the manufacturerM-bM-^@M-^Ys recommendation. Data analysis was performed using RQ Manager 1.2.2 and DataAssist v3.01 software. Data was normalized using Global normalization method and multiple comparisons correction was performed using Benjamini-Hochberg method.
Project description:The primary objective of this study was to evaluate response to a SIV DNA-based vaccine that was adminstered via vivo electroporation (EP) in rhesus macaques to further understand the molecular correlates of protection against SIV. In this study, rhesus macaques were immunized with a DNA vaccine including individual plasmids encoding SIV gag, env, and pol alone, or in combination with a molecular adjuvant, plasmid DNA expressing the chemokine ligand 5 (RANTES), followed by EP. At eight month post-vaccination, animals were challenged with SIV. Standard immunological assays, flow-based activation analysis without ex vivo restimulation and high-throughput gene expression analysis were performed to determine the host response to each vaccine regimen. The overall study was designed to evaluate the response to a SIV-DNA vaccination administered to animals via intramuscular electroporation. Chinese rhesus macaques were divided into three treatment groups (n=6 animals per group): Control (no vaccination), DNA vaccine alone (pCSIVgag, pCSIVpol, pCSIVenv), DNA vaccine with RANTES adjuvant (pCSIVgag, pCSIVpol, pCSIVenv, pmacRANTES). Eight months following the last vaccination, animals were infected with 25 MID of SIVmac251 and response to infection was monitored. RNA for microarray analysis was isolated from fresh PBMCs that were isolated from individual animals and treated overnight with a pool of overlapping SIV pol peptides or mock treated. Samples for microarray analysis were taken longitudinally at 8 months post-vaccination (pre-SIV challenge; biological n=5-6 per group for each treatment; technical n=2 for each sample) and at day 10 post-SIV challenge (n=5-6 per group for each treatment; technical n=2 for each sample).
Project description:The study describes miRNA expression in colonic epithelium of chronic SIV-infected rhesus macaques. We profiled and characterized miRNA/mRNA expression exclusively in colonic epithelium (CE) of 12 chronically SIV-infected and 8 control rhesus macaques (RMs). About 46 miRNAs were differentially expressed (DE) (20-up and 26-down) in CE during chronic SIV infection. Using TargetScan, we bioinformatically crossed the predicted targets of DE miRNAs to genome-wide transcriptomic data and identified several critical miRNA-mRNA pairings that suggested miRNA-mediated regulation of aberrant epithelial gene expression in CE. Immunofluorescence, luciferase reporter and miRNA overexpression studies confirmed the ability of miR-130a and miR-212 to bind the 3’ UTR and downregulate protein expression of occludin (OCLN) and peroxisome proliferator activator receptor gamma (PPAR), respectively, two proteins with pivotal roles in epithelial barrier function. MiR-130a and miR-212 overexpression in Caco-2 cells significantly reduced transepithelial electrical resistance (TEER). Interestingly, delta-9-tetrahydrocannabinol (9-THC) treatment restored TEER to levels observed with control miRNA mimic. Finally, ex-vivo 9-THC treatment of colon tissue from chronically SIV-infected RMs significantly increased PPAR gene expression. Our findings suggest that dysregulated miR-130a and miR-212 expression in CE during chronic HIV/SIV infection can facilitate epithelial barrier disruption by downregulating OCLN and PPAR protein expression. Most importantly, our results highlight the beneficial effects of cannabinoids on epithelial barrier function in not just HIV/SIV but potentially other chronic intestinal inflammatory diseases.
Project description:Lassa fever (LF) is a rodent-borne viral disease that can be fatal for human beings. In this study, an attenuated Lassa vaccine candidate, ML29, was tested in SIV-infected rhesus macaques for its ability to elicit immune responses without instigating signs of virulent disease. ML29 is a reassortant between Lassa and Mopeia viruses that causes a transient infection in non-human primates and confers sterilizing protection from lethal Lassa viral challenge. However, since the LF endemic area of West Africa also has high HIV seroprevalence, it is important to determine whether vaccination could be safe in the context of AIDS. SIV-infected and uninfected rhesus macaques were vaccinated with the ML29 virus and monitored for classical and non-classical signs of arenavirus disease. Classical disease signs included viremia, rash, weight loss, high liver enzyme levels, and virus invasion of the central nervous system. Non-classical signs derived from profiling the blood transcriptome of virulent and non-virulent arenavirus infections included increased expression of interferon response genes and decreased expression of COX2, IL-1?, coagulation intermediates and nuclear receptors needed for stress signaling. Here it is demonstrated that SIV-infected and uninfected rhesus macaques responded similarly to ML29 vaccination, and that none developed signs of arenavirus disease or persistence. Furthermore, 5 of 5 animals given a heterologous challenge with a lethal dose of LCMV-WE survived without developing disease signs. 30 RNA samples from Monkey PBMC: 4 uninf. Monkey PBMC, 8 SIV-infected Monkey PBMC(From 8 Monkeys), 5 SIV+ML29-sc infected week1(Monkey PBMC), 5 SIV+ML29-sc infected week2(Monkey PBMC), 1 SIV+ML29-ig infected week1(Monkey PBMC), 1 SIV+ML29-ig infected week2(Monkey PBMC), 2 SIV+Arm-sc infected week1(Monkey PBMC), 2 SIV+Arm-sc infected week2(Monkey PBMC), 1 only ML29-iv infected week1(Monkey PBMC), 1 only ML29-iv infected week2(Monkey PBMC)
Project description:The study describes miRNA expression in intact duodenum following chronic delta 9 tetrahydrocannabinol (Δ9-THC) administration to SIV-infected rhesus macaques. Chronic Δ9-THC administration to uninfected macaques significantly and positively modulated intestinal miRNA expression by increasing the total number of differentially expressed miRNAs from 14 to 60 days post infection (DPI). At 60DPI, ~28% of miRNAs showed decreased expression in VEH/SIV compared to none in the THC/SIV group. Furthermore, compared to the VEH/SIV group, THC selectively upregulated the expression of miR-10a, miR-24, miR-99b, miR-145, miR-149 and miR-187 previously shown to target proinflammatory molecules. NOX4, a potent reactive oxygen species generator was confirmed as a direct miR-99b target. A significant increase in NOX4+ crypt epithelial cells was detected in VEH/SIV compared to the THC/SIV group. We speculate that miR-99b-mediated NOX4 downregulation may protect the intestinal epithelium from oxidative stress-induced damage.