Effects Of Heat Stress On Physiological And Biochemical Indexes And Expression Levels Of MicroRNA In Peripheral Blood

Heat-stress is one of the significant factors affecting the growth and productivity of animals, it’s valuable to study the mechanism of heat-stress in theory and practice. The studies have found that microRNAs(miRNA) involved in the regulation of various biological processes, along with the widely use of high-throughput sequencing technology, small RNA sequencing technology(miRNA-seq) can tap various functional miRNAs under different level heat-stress. Differential expression analysis of different biological processes, can interpretation the miRNAs regulation and function differences in physiological changes, and explore potential regulatory miRNAs which involved in heat-stress.The trial selected five Red Angus cattles which were healthy, maintained as the same feeding and management from seed multiplication beef cattle farm of Chongqing and their ages and weights should be similar. Measured bovine respiratory rate and rectal temperature, then collected bovine peripheral blood from jugular vein under heat-stress and non-heat-stress respectively. Using various kits to measure serum biochemical indexes, antioxidant indexes and immune-related indexes under heat-stress and non-heat-stress. Then, constructed two small RNA libraries(HH and HC), obtained different types of miRNAs by Solexa sequencing. And thus analyzed the differential expression miRNAs in2libraries, predicted target genes by Targetscan. At last analyzed and forecasted the physiological processed that these target genes involved in, explained the different functions of differentially expressed miRNAs under heat-stress.The results of physiological and biochemical indexes indicated that the respiratory rate and rectal temperature under heat-stress were significantly higher than under non-heat stress(P<0.05); The concentration of serum K+, Na+and Ca+was lower under non-heat stress(P＞0.05), the concentration of serum Cl-was significantly lower than that in non-heat stress(P<0.05), these showed that bovine internal environment was imbalance; The antioxidant indexes:T-AOC and GSH-Px were significantly lower(P<0.05) under heat stress, and SOD was extremely significantly lower than that under non-heat stress(P<0.01), it showed that bovine antioxidant capacity decreased; Serum IgA and IgG levels were significantly lower(P<0.05), IgM level was lower than that under non-heat stress(P>0.05), under heat stress, the content of IL-2, IL-6and IL-10was lower(P>0.05), however IL-4and IL-8was higher(P>0.05), these indicated that bovine immune function decreased.Total of the2small RNA libraries obtained1.91million unique reads and25.34million total reads. Excluding junk reads, non-miRNA reads et al, obtained730thousands clean unique reads, and19.59million total reads, and all the reads meet the feature of Dicer.Compared with various databases, we identified1011miRNAs, including546known miRNA in cattle,168inter-species conserved miRNAs and297new predicted miRNAs; we also found267pre-miRN As co-expressed, indicating that the regulation of miRNAs were complex. In the2libraries, total of832miRNAs co-expressed,89and90miRNAs expressed specifically in HH and HC; The average percent of miRNAs which among3-100copies is64.78%, the average proportion of the copies of top20miRNAs was70.63%, these indicated that a small number of miRNAs expressed highly in the2libraries.Analysis of the differential expression miRNAs showed that there were315miRNAs expressed differentially(P<0.01),118miRANs up-regulated and197miRNAs down-regulated under heat stress. GO and Pathway annotation of top10most differential miRNAs showed that miRNAs up-regulated expressed in heat stress were maily involved in apoptosis, cell proliferation and protein kinase cascade, miRNAs down-regulated expressed mainly involved in the regulation of immune function.This study revealed the impact of heat stress on physiological and biochemical indexes of Bos Taurus, identified candidate miRNAs expressed in bovine peripheral blood, analyzed bovine different biological regulation from the perspective of miRNA. It also provided basis for improving and researching the biological functions of various genes under different environment.