Enucleation-Induced Rat Adrenal Gland Regeneration: Expression Profile of Selected Genes Involved in Control of Adrenocortical Cell Proliferation

Abstract

Enucleation-induced adrenal regeneration is a highly controlled process; however, only some elements involved in this process have been recognized. Therefore, we performed studies on regenerating rat adrenals. Microarray RNA analysis and QPCR revealed that enucleation resulted in a rapid elevation of expression of genes involved in response to wounding, defense response, and in immunological processes. Factors encoded by these genes obscure possible priming effects of various cytokines on initiation of regeneration. In regenerating adrenals we identified over 100 up- or downregulated genes involved in adrenocortical cell proliferation. The changes were most significant at days 2-3 after enucleation and their number decreased during regeneration. For example, expression analysis revealed a notable upregulation of the growth arrest gene, Gadd45, only 24 hours after surgery while expression of cyclin B1 and Cdk1 genes was notably elevated between days 1–8 of regeneration. These changes were accompanied by changes in expression levels of numerous growth factors and immediate-early transcription factors genes. Despite notable differences in mechanisms of adrenal and liver regeneration, in regenerating adrenals we identified genes, the expression of which is well recognized in regenerating liver. Thus, it seems legitimate to suggest that, in the rat, the general model of liver and adrenal regeneration demonstrate some degree of similarity.

Nicotinamide phosphoribosyltransferase and the hypothalamic-pituitary-adrenal axis of the rat

“…RNA isolation. The methods used have been described previously (24)(25)(26)(27)(28). Total RNA was extracted from collected cells, samples of adrenal zones, entire adrenal glands, periadrenal adipose tissue, pituitaries and hypothalami using TRI Reagent and subsequently purified on columns (NucleoSpin RNA XS; Macherey-Nagel, Düren, Germany).…”

“…qPCR was performed using a LightCycler 2.0 instrument 4.05 software version (Roche Diagnostics). Using the aforementioned primers, a SYBR Green detection system was applied, as described previously (24)(25)(26)(27)(28)(29). Every 20 µl reaction mixture contained 2 µl template cDNA (standard or control), 0.5 µM specific primers and a previously determined optimum MgCl 2 concentration (3.5 µM for one reaction).…”

Visinin-like peptide 1 in adrenal gland of the rat. Gene expression and its hormonal control

“…The applied methods were described earlier [1,2,[45][46][47][53][54][55][56]. From collected cells, samples of adrenal zones and from samples of entire adrenal glands, total RNA was extracted using TRI Reagent (Sigma) and, then, purified on columns (Rnasy Mini Kit, Qiagen).…”

“…Microarray expression analysis was also described earlier [54][55][56]. RNA subjected to microarray studies was obtained from the above described experiments.…”

“…Using the above mentioned primers, SYBR green detection system was applied as described earlier [45][46][47][54][55][56]. Every of 20 l reaction mixtures contained 4 l template cDNA (standard or control), 0.5 M of specific primer and a previously determined optimum MgCl 2 concentration (3.5 M for one reaction).…”