High-altitude pulmonary hypertension in cattle (brisket disease): Candidate genes and gene expression profiling of peripheral blood mononuclear cells.
High-altitude pulmonary hypertension in cattle (brisket disease): Candidate genes and gene expression profiling of peripheral blood mononuclear cells.
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DOI：
10.4103/2045-8932.93545
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发表时间：
2011-10
影响因子：
2.6
通讯作者：
Hamid R
中科院分区：
文献类型：
--
作者：
Newman JH;Holt TN;Hedges LK;Womack B;Memon SS;Willers ED;Wheeler L;Phillips JA 3rd;Hamid R
高原肺动脉高压（HAPH）是慢性肺泡缺氧的结果，导致缺氧性血管收缩和肺循环重建。牛肺动脉高压病是一种自然发生的缺氧性肺动脉高压动物模型。遗传易感牛在海拔7，000英尺以上会出现严重的肺动脉高压和右心衰竭。目前没有关于HAPH或影响严重程度的途径和基因的信息。我们假设，可以通过以下策略发现对疾病发病机制的初步见解：（1）通过单核苷酸多态性（SNP）分析揭示的功能候选物的测序，以及（2）与海拔匹配的正常对照相比，受影响牛的基因表达谱，基因集富集分析（GSEA）和免疫途径分析（IPA）。我们通过颈静脉穿刺从一群12-18月龄的黑色安格斯牛（雌雄各半）中分离血液。10头感染牛的平均肺动脉压为85.6±13 mmHg STD，10头耐药牛的平均肺动脉压为35.3±1.2 mmHg STD，P<0.001。从外周血单核细胞中，将DNA与Affyssin 10 K基因芯片SNP杂交，并将RNA用于探测Affyssin Bovine基因组阵列。使用Btau 4.0牛基因组组装重新定位SNP位点。通过Partek软件包分析mRNA数据，以鉴定两组之间具有统计学差异的表达的基因组。对精炼的表达数据进行GSEA和IPA，以鉴定关键的细胞通路并生成网络，并对通路和网络进行功能分析。通过等位基因关联鉴定了10个SNP，并对队列中的4个候选基因进行了测序。无论是内皮型一氧化氮合成酶，NADH脱氢酶，TG相互作用因子-2，也没有BMPR 2感染和耐药牛之间的差异。一个60个基因的mRNA签名被确定，区分从未受影响的牛。46个基因在受影响的牛中过表达，14个基因在受影响的牛中下调至少20%。GSEA和Incidity分析确定呼吸系统疾病、炎症性疾病和途径是与该基因签名相关的最常见疾病和障碍（P<5.14×10-14），细胞发育和细胞信号传导是最常见的细胞功能（P<1.20×10-08），IL 6、TREM、PPAR、NF κ B细胞信号传导（P<8.69×10-09）是与该基因签名相关的最常见典型途径。这项研究在分子水平上深入了解了HAPH中RNA表达的差异，并排除了四个功能候选基因。需要进一步的研究来验证和完善这些初步发现，并确定转录基因在HAPH发生中的作用。
High-altitude pulmonary hypertension (HAPH) is a consequence of chronic alveolar hypoxia, leading to hypoxic vasoconstriction and remodeling of the pulmonary circulation. Brisket disease in cattle is a naturally occurring animal model of hypoxic pulmonary hypertension. Genetically susceptible cattle develop severe pulmonary hypertension and right heart failure at altitudes >7,000 ft. No information currently exists regarding the identity of the pathways and gene(s) responsible for HAPH or influencing severity. We hypothesized that initial insights into the pathogenesis of the disease could be discovered by a strategy of (1) sequencing of functional candidates revealed by single nucleotide polymorphism (SNP) analysis and (2) gene expression profiling of affected cattle compared with altitude-matched normal controls, with gene set enrichment analysis (GSEA) and Ingenuity pathway analysis (IPA). We isolated blood from a single herd of Black Angus cattle of both genders, aged 12-18 months, by jugular vein puncture. Mean pulmonary arterial pressures were 85.6±13 mmHg STD in the 10 affected and 35.3±1.2 mmHg STD in the 10 resistant cattle, P<0.001. From peripheral blood mononuclear cells, DNA was hybridized to an Affymetrix 10K Gene Chip SNP, and RNA was used to probe an Affymetrix Bovine genome array. SNP loci were remapped using the Btau 4.0 bovine genome assembly. mRNA data was analyzed by the Partek software package to identify sets of genes with an expression that was statistically different between the two groups. GSEA and IPA were conducted on the refined expression data to identify key cellular pathways and to generate networks and conduct functional analyses of the pathways and networks. Ten SNPs were identified by allelelic association and four candidate genes were sequenced in the cohort. Neither endothelial nitric oxide synthetase, NADH dehydrogenase, TG-interacting factor-2 nor BMPR2 were different among affected and resistant cattle. A 60-gene mRNA signature was identified that differentiated affected from unaffected cattle. Forty-six genes were overexpressed in the affected and 14 genes were downregulated in the affected cattle by at least 20%. GSEA and Ingenuity analysis identified respiratory diseases, inflammatory diseases and pathways as the top diseases and disorders (P<5.14×10-14), cell development and cell signaling as the top cellular functions (P<1.20×10-08), and IL6, TREM, PPAR, NFkB cell signaling (P<8.69×10-09) as the top canonical pathways associated with this gene signature. This study provides insights into differences in RNA expression in HAPH at a molecular level, and eliminates four functional gene candidates. Further studies are needed to validate and refine these preliminary findings and to determine the role of transcribed genes in the development of HAPH.