Ramsay Timothy G, Stoll Margo J, Blomberg Le Ann, Caperna Thomas J
Animal Biosciences and Biotechnology Laboratory, USDA/ARS, Beltsville Agricultural Research Center, USDA, Beltsville, MD 20705 USA.
J Anim Sci Biotechnol. 2016 Apr 14;7:25. doi: 10.1186/s40104-016-0081-0. eCollection 2016.
Porcine adipose tissue expresses orosomucoid (ORM1) mRNA, a protein with anti-inflammatory and immunomodulatory properties. Previous research has demonstrated that porcine ORM1 can reduce insulin stimulated glucose metabolism in porcine adipose tissue in vitro. The present study was designed to examine the preweaning ontogeny of ORM1 mRNA abundance in porcine subcutaneous adipose and to determine if ORM1 can regulate mRNA abundance of inflammatory cytokines that contribute to insulin resistance in primary cultures derived from neonatal porcine subcutaneous adipose tissue. Cultures were differentiated in vitro and subsequently the adipocyte containing cultures were incubated for 24 h with 0-5000 ng porcine ORM1/mL medium. Cultures were then harvested, total RNA extracted for use in reverse transcription and the mRNA abundance of cytokine mRNA quantified by real-time PCR.
ORM1 mRNA abundance within neonatal adipose tissue does not change from d 1 to d 21 of age and is a very small fraction relative to liver mRNA abundance. The ORM1 mRNA level in porcine adipocytes and stromal-vascular cells are similar (P > 0.05). Treatment with ORM1 did not affect TNFα (tumor necrosis factor α) mRNA level (P > 0.05), while interleukin 6 (IL6) mRNA abundance was reduced 32 % at 1,000 ng ORM1/mL (P < 0.01). However, TNFα protein content in the cell culture media was reduced by ORM1 treatment (5,000 ng/mL, P < 0.05), whereas ORM1 had no detectable effect on the media content of IL6 (P > 0.05). The reduction of macrophage migration inhibitory factor (MIF) mRNA abundance by ORM1 was dose dependent (P < 0.01). Monocyte chemotactic protein (MCP) mRNA level was reduced 27 % by 1,000 ng ORM1/mL (P < 0.05).
The data suggest that ORM1 has limited effects TNFα, IL6, MIF or MCP expression at the concentrations tested. Secondly, these cytokines do not appear to contribute to the reported insulin resistance induced by ORM1 in porcine adipose tissue in vitro as an increase in the abundance of these inflammatory cytokines would be predicted during an insulin resistant state.
猪脂肪组织表达类粘蛋白（ORM1）mRNA，这是一种具有抗炎和免疫调节特性的蛋白质。先前的研究表明，猪ORM1在体外可降低猪脂肪组织中胰岛素刺激的葡萄糖代谢。本研究旨在检测猪皮下脂肪中ORM1 mRNA丰度在断奶前的个体发育情况，并确定ORM1是否能调节源自新生猪皮下脂肪组织的原代培养物中有助于胰岛素抵抗的炎性细胞因子的mRNA丰度。培养物在体外分化，随后将含有脂肪细胞的培养物用0 - 5000 ng猪ORM1/mL培养基孵育24小时。然后收获培养物，提取总RNA用于逆转录，并通过实时PCR定量细胞因子mRNA的mRNA丰度。
新生脂肪组织中ORM1 mRNA丰度在1日龄至21日龄期间没有变化，相对于肝脏mRNA丰度而言是非常小的一部分。猪脂肪细胞和基质血管细胞中的ORM1 mRNA水平相似（P > 0.05）。用ORM1处理不影响肿瘤坏死因子α（TNFα）mRNA水平（P > 0.05），而在1000 ng ORM1/mL时白细胞介素6（IL6）mRNA丰度降低了32%（P < 0.01）。然而，ORM1处理降低了细胞培养基中TNFα蛋白含量（5000 ng/mL，P < 0.05），而ORM1对培养基中IL6含量没有可检测到的影响（P > 0.05）。ORM1对巨噬细胞迁移抑制因子（MIF）mRNA丰度的降低呈剂量依赖性（P < 0.01）。1000 ng ORM1/mL使单核细胞趋化蛋白（MCP）mRNA水平降低了27%（P < 0.05）。
数据表明，在所测试的浓度下，ORM1对TNFα、IL6、MIF或MCP的表达影响有限。其次，这些细胞因子似乎并未导致体外猪脂肪组织中报道的由ORM1诱导的胰岛素抵抗，因为在胰岛素抵抗状态下预计这些炎性细胞因子的丰度会增加。
