Expression profiling by high throughput sequencing
Summary
Purpose: The study was designed to determine expression differences in brain regions of F344 and HIV-1 Transgenic rats by using RNA-sq analysis. Methods: 144 RNA samples (2 strains, 2 treatments, 3 regions, and 12 animals per group) were analyzed. Following deep-sequencing analysis of 50-bp paired-end reads of RNA-Seq, we used Bowtie/Tophat/Cufflinks suites to align these reads into transcripts based on the Rn4 rat reference genome and to measure the relative abundance of each transcript. MATLAB was used to conduct all statistical analysis. qRT–PCR validation was performed using TaqMan and SYBR Green assays fo soem representative genes. Results: Statistical and bioinformatic analyses on each brain region in the two strains revealed that immune response- and neurotransmission-related pathways were altered in the HIV-1Tg rats, with brain region differences. Other neuronal survival-related pathways, including those encoding myelin proteins, growth factors, and translation regulators, were altered in the HIV-1Tg rats in a brain region-dependent manner. After nicotine expousure, 20% of the altered genes in the HIV-1Tg rat were affected by nicotine in each brain region, with the expression of most restored. Analysis of the restored genes showed distinct pathways corrected by nicotine in different brain regions of HIV-1Tg rats. Conclusions: The abnormal gene expression pattern discovered in HIV-1Tg rats suggest mechanisms underlying the deficits in learning and memory and vulnerability to drug addiction and other psychiatric disorders observed in HIV positive patients. The gene expression pattern in the HIV-1Tg rats after nicotine exposure indicate that cholinergic modulators such as nicotine may have beneficial effects on HIV-1-induced neurologic deficits.
Overall design
144 RNA samples (2 strains, 2 treatments, 3 regions, and 12 animals per group) were analyzed.
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