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Submitted to: Journal of Animal Science
Publication Type: Abstract Only Publication Acceptance Date: 5/12/2009 Publication Date: 7/12/2009 Citation: Ramsay, T.G. 2009. Albumin induced cytokine expression in porcine adipose tissue explants. Journal of Animal Science 87, Suppl 2, p. 239. Interpretive Summary: Cytokines have critical roles in the regulation of animal growth and health. Recent studies have demonstrated that adipose tissue is a significant source of cytokines which can subsequently act to alter adipose tissue growth and metabolism. This may be of importance in the survival of the neonatal pig when adipose tissue is limited. The present study was performed to determine why in vitro experiments to assess these adipose tissue derived cytokines has been so difficult to interpret. The data from this study indicate that a protein (albumin) commonly used in the medium to lubricate the tissue and to absorb fatty acids released by the tissue can acutely and dramatically alter the expression of a number of adipose derived cytokines. This has required the development of alternative methods to assess cytokine production by pig adipose tissue in culture wherein all conditions may be controlled. Technical Abstract: Albumin has historically been included in medium designed for use with adipose tissue when evaluating metabolism, gene expression or protein secretion. However, recent studies with mouse adipocytes (Ruan et al., J. Biol. Chem. 278:47585-47593, 2003) and human adipose tissue (Schlesinger et al., Amer. J. Physiol. 291:27-33, 2006) have demonstrated an acute cytokine response by cells derived from adipose tissue to albumin. The present study was designed to determine whether or not albumin can alter cytokine expression in porcine adipose tissue explants relative to explants not exposed to albumin. Subcutaneous adipose tissue explants (100 mg) were prepared from dorsal subcutaneous adipose tissue of 21 day old pigs using a Stadie-Riggs microtome. Slices were placed in Hanks balanced salt solution prior to albumin exposure. Triplicate slices were blotted and frozen in liquid nitrogen (controls). Additional slices were incubated in medium 199 supplemented with 100 mM Hepes and 0.5% albumin at 37ºC. Triplicate tissue slices were removed from the medium at 1, 2 or 4 hours following exposure to the albumin containing medium, blotted and frozen in liquid nitrogen. Samples were extracted for RNA using Qiagen columns and RNA (1 ug) was reverse transcribed and used in real-time PCR analysis to assess the expression of the adipokines leptin, adiponectin, tumor necrosis factor alpha (TNF-alpha), interleukin 6 (IL6) and IL15 relative to cyclophilin. Expression of leptin and adiponectin were not altered by exposure to 0.5% albumin (P > 0.05). However TNF alpha mRNA abundance was elevated 1436 ± 134 % within 1hr of exposure to albumin (P< 0.001). Similarly, IL6 was elevated by greater than 1246 ± 286% (P < 0.001) within 1hr of incubation. In contrast, IL15 mRNA abundance was reduced by 62 ± 8% (P <0.01) within 4hr. These data indicate that albumin alters the expression of specific adipocytokines. Secondly the data indicate that in vitro analysis of porcine adipocytokine expression may be confounded if albumin is utilized in the medium. |
