A microarray gene analysis of peripheral whole blood in normal adult male rats after long-term GH gene therapy

Abstract

The main aims of this study were to determine the effects of GH gene abuse/misuse in normal animals and to discover genes that could be used as candidate biomarkers for the detection of GH gene therapy abuse/misuse in humans. We determined the global gene expression profile of peripheral whole blood from normal adult male rats after long-term GH gene therapy using CapitalBio 27 K Rat Genome Oligo Arrays. Sixty one genes were found to be differentially expressed in GH gene-treated rats 24 weeks after receiving GH gene therapy, at a two-fold higher or lower level compared to the empty vector group (p < 0.05). These genes were mainly associated with angiogenesis, oncogenesis, apoptosis, immune networks, signaling pathways, general metabolism, type I diabetes mellitus, carbon fixation, cell adhesion molecules, and cytokine-cytokine receptor interaction. The results imply that exogenous GH gene expression in normal subjects is likely to induce cellular changes in the metabolism, signal pathways and immunity. A real-time qRT-PCR analysis of a selection of the genes confirmed the microarray data. Eight differently expressed genes were selected as candidate biomarkers from among these 61 genes. These 8 showed five-fold higher or lower expression levels after the GH gene transduction (p < 0.05). They were then validated in real-time PCR experiments using 15 single-treated blood samples and 10 control blood samples. In summary, we detected the gene expression profiles of rat peripheral whole blood after long-term GH gene therapy and screened eight genes as candidate biomarkers based on the microarray data. This will contribute to an increased mechanistic understanding of the effects of chronic GH gene therapy abuse/misuse in normal subjects.

Hepatic adiponectin receptor R2 expression is up-regulated in normal adult male mice by chronic exogenous growth hormone levels

“…The raaV2/1 vector containing GH1 was constructed and produced as previously described (23). Briefly, GH1 was cloned from a polymerase chain reaction (Pcr) product using the primers 5'-GccaGaaTTcGccac caTGGcTacaGGcTcccGG-3' (forward) and 5'-cTGcGT cGacGaaGccacaGcTGcccTc-3' (reverse) (Ecori and Sali restriction sites are indicated by underlining) from the template of a puc19 plasmid dna containing GH1 (Xinxiang Medical university, Henan Province, P.r.…”

Analysis of the role of the integrin signaling pathway in hepatocytes during rat liver regeneration

“…The quality of total RNA was assessed by optical density measurement at 260/280 nm and agarose electrophoresis (180 V, 0.5 h); the sample with a 2:1 ratio of 28S RNA to 18S RNA was regarded as qualified [17]. T7-oligo DT (24) (Keck Foundation, New Haven, CT), SuperScript II RT (Invitrogen Corporation, Carlsbad, CA) and 5 μg of total RNA were used to synthesize the first strand of cDNA.…”

Impacts of DNA Microarray Technology in Gene Therapy

“…The results imply that exogenous GH gene expression in normal subjects is likely to induce cellular changes in the metabolism, signal pathways and immunity. Based on such screening, these researchers claimed eight genes as candidate biomarkers (Qin & Tian, 2010). Since genomedicines (e.g., antisense oligo deoxy nucleotides (As-ODN), RNA interference) can selectively block disease-causing genes, the responsible genes of diseases (e.g., cancer genes) have been chosen as potential targets even though undesired nonspecific side effects may tarnish the actual mechanism of gene therapy, blemishing clinical development of gene based therapeutics.…”