Proteomic study of hypothalamus in pigs exposed to heat stress

Abstract

Background: With evidence of warming climates, it is important to understand the effects of heat stress in farm animals in order to minimize production losses. Studying the changes in the brain proteome induced by heat stress may aid in understanding how heat stress affects brain function. The hypothalamus is a critical region in the brain that controls the pituitary gland, which is responsible for the secretion of several important hormones. In this study, we examined the hypothalamic protein profile of 10 pigs (15 ± 1 kg body weight), with five subjected to heat stress (35 ± 1 °C; relative humidity = 90%) and five acting as controls (28 ± 3 °C; RH = 90%).

Result: The isobaric tags for relative and absolute quantification (iTRAQ) analysis of the hypothalamus identified 1710 peptides corresponding to 360 proteins, including 295 differentially expressed proteins (DEPs), 148 of which were up-regulated and 147 down-regulated, in heat-stressed animals. The Ingenuity Pathway Analysis (IPA) software predicted 30 canonical pathways, four functional groups, and four regulatory networks of interest. The DEPs were mainly concentrated in the cytoskeleton of the pig hypothalamus during heat stress.

Conclusions: In this study, heat stress significantly increased the body temperature and reduced daily gain of body weight in pigs. Furthermore, we identified 295 differentially expressed proteins, 147 of which were down-regulated and 148 up-regulated in hypothalamus of heat stressed pigs. The IPA showed that the DEPs identified in the study are involved in cell death and survival, cellular assembly and organization, and cellular function and maintenance, in relation to neurological disease, metabolic disease, immunological disease, inflammatory disease, and inflammatory response. We hypothesize that a malfunction of the hypothalamus may destroy the host physical and immune function, resulting in decreased growth performance and immunosuppression in heat stressed pigs.

Keywords: Heat stress; Hypothalamus; Pigs; Quantitative proteomics.

Fig. 1

Clinical changes caused by heat stress. (a) Changes in pig body weight, (b) forehead temperature, and (c) rectal temperature, under heat stress

Fig. 2

Protein identification in the hypothalamus of heat-stressed (HS) and control pigs. (a) Numbers of proteins identified, (b) distribution of the DEPs among molecular weight (kD) classes, (c) coverage of DEPs by the identified peptides, and (d) distribution of DEPs containing different numbers of identified peptides

Fig. 3

Bioinformatics analysis of the DEPs identified in the hypothalamus of pigs. a Subcellular locations of the proteins with differential expression in heat-stressed (HS) and control pigs. b Thirty most related canonical pathways from Ingenuity Pathway Analysis

Fig. 4

Functional characterization of DEPs in the hypothalamus of pigs under heat stress. (a) diseases and disorders, (b) molecular and cellular functions, (c) physiological system development and functions, and (d) toxicological functions

Fig. 5

Ingenuity Pathway Analysis of differentially expressed proteins in the hypothalamus of pigs under heat stress. Red is up-regulated, green is down-regulated, and white is a protein that is involved in pathway regulation but not identified in this study. A darker color indicates a greater change in the expression level of the protein. Different shapes represent different molecular types (e.g., protein families). The lines connecting the molecules represent intermolecular relationships—the dashed lines are indirect effects and the solid lines are direct effects. The arrows represent specific molecular relationships and directions of action

Fig. 6

Western blotting identification of DEPs. a Expression of Histone H2A in the hypothalamus of pigs at day 7 of heat stress. Con = control pigs; HS = heat-stressed pigs. b Expression of Histone H2A was significantly increased in the HS group compared with the control group