Xiao Y, Rungruang S, Hall L W, Collier J L, Dunshea F R, Collier R J
School of Animal and Comparative Biomedical Sciences, University of Arizona, Tucson 85721.
Feed Technology, Charoen Pokphand Company, Bangkok, Thailand 10500.
J Dairy Sci. 2017 May;100(5):4025-4037. doi: 10.3168/jds.2016-11876. Epub 2017 Feb 23.
The objective of this study was to investigate the direct effects of feed supplements niacin and betaine on the heat shock responses of in vitro cultured cells derived from bovine mammary and uterine tissues. First, we determined the mRNA expression profiles of the niacin receptor (GPR109A) in bovine tissues (liver, skin, uterus, udder, and ovary) and in cells derived from bovine mammary epithelium (mammary alveolar cells, MAC-T; bovine mammary epithelial cells, BMEC) and endometrium (bovine endometrial cells, BEND). We found that GPR109A was distributed in all examined tissues and cells, and the highest expression was in cells from skin and udder. Second, we evaluated the effects of niacin treatment on the mRNA abundance of heat shock proteins 70 and 27 (HSP70 and HSP27) in MAC-T, BMEC, and BEND under thermoneutral conditions and heat stress, and whether these effects were associated with alterations in the mRNA expression of prostaglandin E synthesis-related genes, including cyclooxygenase 1 and 2 (COX-1 and COX-2) and microsomal prostaglandin E synthase 1 and 2 (mPGES-1 and mPGES-2). Quantitative PCR data indicated that niacin suppressed HSP70 mRNA expression in BMEC and both HSP70 and HSP27 in BEND under thermoneutral conditions. Only COX-2 expression was downregulated by niacin in BMEC; other prostaglandin E synthesis-related genes stayed unaltered in BMEC and BEND. The mRNA abundance of HSP70, COX-1, COX-2, and mPGES-1 were elevated in niacin-treated MAC-T. During heat stress, niacin increased mRNA levels of HSP70 and HSP27 in MAC-T and HSP27 in BEND, but decreased HSP70 in BMEC. Although mPGES-2 was stimulated by niacin in BEND, the mRNA expression of prostaglandin E synthesis-related genes were consistent with neither HSP70 nor HSP27 expression patterns in niacin-treated BMEC and MAC-T. These data suggest that the effects of niacin on heat shock protein expression and prostaglandin E synthesis were not well coupled in these cells. Finally, we tested the effects of betaine treatment on viability and apoptosis in BMEC. Compared with control cultures, viability was higher in betaine-treated cells at 8 h under thermoneutral conditions and at 16 h in heat stress, and apoptotic rates were lower at 8 h. Our data support a dual role for niacin in regulating heat shock protein expression in normal and heat-shocked cells derived from mammary and uterine tissues, and positive effects of betaine in regulating mammary cell viability during heat stress.
本研究的目的是调查饲料添加剂烟酸和甜菜碱对源自牛乳腺和子宫组织的体外培养细胞热休克反应的直接影响。首先，我们测定了烟酸受体（GPR109A）在牛组织（肝脏、皮肤、子宫、乳房和卵巢）以及源自牛乳腺上皮（乳腺肺泡细胞，MAC-T；牛乳腺上皮细胞，BMEC）和子宫内膜（牛子宫内膜细胞，BEND）的细胞中的mRNA表达谱。我们发现GPR109A分布于所有检测的组织和细胞中，最高表达量出现在皮肤和乳房的细胞中。其次，我们评估了烟酸处理对热中性条件和热应激下MAC-T、BMEC和BEND中热休克蛋白70和27（HSP70和HSP27）mRNA丰度的影响，以及这些影响是否与前列腺素E合成相关基因（包括环氧化酶1和2（COX-1和COX-2）以及微粒体前列腺素E合酶1和2（mPGES-1和mPGES-2））的mRNA表达变化有关。定量PCR数据表明，在热中性条件下，烟酸抑制了BMEC中HSP70的mRNA表达以及BEND中HSP70和HSP27的表达。在BMEC中，只有COX-2的表达被烟酸下调；其他前列腺素E合成相关基因在BMEC和BEND中保持不变。在烟酸处理的MAC-T中，HSP70、COX-1、COX-2和mPGES-1的mRNA丰度升高。在热应激期间，烟酸增加了MAC-T中HSP70和HSP27以及BEND中HSP27的mRNA水平，但降低了BMEC中HSP70的水平。尽管在BEND中烟酸刺激了mPGES-2，但在烟酸处理的BMEC和MAC-T中，前列腺素E合成相关基因的mRNA表达与HSP70和HSP27的表达模式均不一致。这些数据表明，烟酸对热休克蛋白表达和前列腺素E合成的影响在这些细胞中没有很好地耦合。最后，我们测试了甜菜碱处理对BMEC活力和凋亡的影响。与对照培养相比，在热中性条件下8小时以及热应激下16小时，甜菜碱处理的细胞活力更高，且在8小时时凋亡率更低。我们的数据支持了烟酸在调节源自乳腺和子宫组织的正常和热休克细胞中热休克蛋白表达方面的双重作用，以及甜菜碱在热应激期间调节乳腺细胞活力的积极作用。
