Madsen Sally A, Weber Patty S D, Burton Jeanne L
Immunogenetics Laboratory, Department of Animal Science, 1205E Anthony Hall, Michigan State University, East Lansing 48824, USA.
Vet Immunol Immunopathol. 2002 Jul;86(3-4):159-75. doi: 10.1016/s0165-2427(02)00020-x.
The periparturient dairy cow undergoes a plethora of physiological changes, including changes in the immune system that lead to profound effects on animal health. Of the immune cells affected at parturition, the neutrophil has been of particular interest due to its primary role in innate immune defense against mastitis. Immune functions of bovine neutrophils are known to be depressed around parturition, but it has not been discerned at what level these alterations occur, including the possibility that parturition induces changes in expression of key genes. The hypothesis of the present study was that blood neutrophils respond to the physiology of parturition by altering the expression of genes needed for normal cellular functions. The main objectives of the study were to detect and characterize parturition induced changes in neutrophil gene expression, to determine if altered gene expression was significantly associated with the main steroid hormones of bovine parturition, and to obtain putative identities of differentially expressed neutrophil genes. Differential gene expression was detected and characterized through mRNA abundance changes in neutrophils, and steroid hormone concentrations by RIA assay of periparturient serum samples. Preliminary assessment of differential gene expression was done using differential display reverse transcription polymerase chain reaction (DDRT-PCR) followed by secondary screening using high throughput cDNA dot blot hybridization. Altered gene expression was confirmed using Northern blot hybridization and detailed expression patterns characterized using quantitative slot blot analysis. The identities of two fully characterized transcripts with clear parturition induced repression (P< or =0.02) in neutrophils had high DNA sequence homology with genes that encode bovine mitochondrial cytochrome b (cytb) and rig/ribosomal protein S15 (rig/RPS15). These proteins are critical for normal respiratory metabolism and translation in cells, respectively. The gene expression profiles for cytb were significantly related to serum progesterone concentration (r=0.44) and for rig/RPS15 to progesterone and estradiol concentrations (r=0.35, 0.36, respectively). Eleven additional transcripts showed evidence of parturition induced repression in neutrophils and were putatively identified as representing genes of the citric acid cycle and various DNA binding proteins. Results of this study show for the first time that genes regulating basic life functions of bovine neutrophils may be repressed by parturition, possibly due to influences of steroid hormones.
围产期奶牛会经历大量生理变化，包括免疫系统的变化，这些变化会对动物健康产生深远影响。在分娩时受影响的免疫细胞中，中性粒细胞因其在抵抗乳腺炎的天然免疫防御中的主要作用而备受关注。已知牛中性粒细胞的免疫功能在分娩前后会受到抑制，但尚未明确这些改变发生在何种水平，包括分娩是否会诱导关键基因表达变化的可能性。本研究的假设是血液中的中性粒细胞通过改变正常细胞功能所需基因的表达来响应分娩生理。该研究的主要目标是检测并表征分娩诱导的中性粒细胞基因表达变化，确定基因表达改变是否与牛分娩的主要类固醇激素显著相关，并获得差异表达的中性粒细胞基因的推定身份。通过中性粒细胞中mRNA丰度变化检测并表征差异基因表达，通过放射免疫分析（RIA）测定围产期血清样本中的类固醇激素浓度。使用差异显示逆转录聚合酶链反应（DDRT-PCR）进行差异基因表达的初步评估，随后使用高通量cDNA斑点印迹杂交进行二次筛选。使用Northern印迹杂交确认基因表达改变，并使用定量斑点印迹分析表征详细的表达模式。在中性粒细胞中，有两个完全表征的转录本在分娩时受到明显抑制（P≤0.02），它们与编码牛线粒体细胞色素b（cytb）和rig/核糖体蛋白S15（rig/RPS15）的基因具有高度DNA序列同源性。这些蛋白质分别对细胞中的正常呼吸代谢和翻译至关重要。cytb的基因表达谱与血清孕酮浓度显著相关（r = 0.44），rig/RPS15的基因表达谱与孕酮和雌二醇浓度相关（分别为r = 0.35和0.36）。另外11个转录本显示在中性粒细胞中有分娩诱导的抑制证据，并被推定为代表柠檬酸循环和各种DNA结合蛋白的基因。本研究结果首次表明，调节牛中性粒细胞基本生命功能的基因可能会因分娩而受到抑制，这可能是由于类固醇激素的影响。
