J Vet Sci.  2016 Mar;17(1):35-44. 10.4142/jvs.2016.17.1.35.

The association of Hsp90 expression induced by aspirin with anti-stress damage in chicken myocardial cells

• Zhang XH
• Zhu HS
• Qian Z
• Tang S
• Wu D
• Kemper N
• Hartung J
• Bao ED

• KMID: 2363333
• DOI: http://doi.org/10.4142/jvs.2016.17.1.35

Abstract

The protective effect of aspirin during exposure to heat stress in broiler chickens was investigated. We assayed pathological damage, expression and distribution of Hsp90 protein and hsp90 mRNA expression in chicken heart tissues after oral administration of aspirin following exposure to high temperature for varying times. Heat stress induced increases in plasma aspartate aminotransferase, creatine kinase and lactate dehydrogenase activities while causing severe heart damage, which was characterized by granular and vacuolar degeneration, nuclear shrinkage and even myocardium fragmentation in cardiac muscle fibers. After aspirin administration, myocardial cells showed fewer pathological lesions than broilers treated with heat alone. A high positive Hsp90 signal was always detected in the nuclei of myocardial cells from broilers treated with aspirin, while in myocardial cells treated with heat alone, Hsp90 in the nuclei decreased, as did that in the cytoplasm. Aspirin induced rapid and significant synthesis of Hsp90 before and at the initial phase of heat stress, and significant expression of hsp90 mRNA was stimulated throughout the experiment when compared with cells exposed to heat stress alone. Thus, specific pre-induction of Hsp90 in cardiovascular tissue was useful for resisting heat stress damage because it produced stable damage-related enzymes and fewer pathologic changes.

Keyword

Hsp90; aspirin; chicken; heart; heat stress damage

MeSH Terms

Animals
Anti-Inflammatory Agents, Non-Steroidal/pharmacology
Aspirin/*pharmacology
Cell Nucleus/genetics
Chickens
Gene Expression Regulation/*drug effects
HSP90 Heat-Shock Proteins/*genetics
Hot Temperature
Myocytes, Cardiac/*drug effects/enzymology/pathology
Stress, Physiological/*drug effects
Aspirin
Anti-Inflammatory Agents, Non-Steroidal
HSP90 Heat-Shock Proteins

Figure

• 

  Fig. 1 Temporal changes in enzyme activities in the serum of experimental chickens. (A) The levels of aspartate aminotransferase (AST) showed an obvious increase from 10 h of heat exposure in the heat stress (HS) group to the end of the test, while the AST in the aspirin administered before heat stress (ASA+HS) and aspirin administration (ASA) groups stayed relatively stable. (B) The levels of creatine kinase (CK) showed an increasing tendency after 3 h of HS in the HS group, while no difference was detected in the CK of the ASA+HS and ASA group (except for a decline at 10 h). (C) The levels of lactate dehydrogenase (LDH) showed a significant increasing tendency in the HS group, with that of the ASA-HS group showing a marked rise after 5 h of HS, then returning to the basal level. No difference was detected for LDH in the ASA group, except for an increase at 7 h. Differences between levels of enzymes in every group at different times and that at 0 h in the HS group are indicated by *p < 0.05, and by **p < 0.01. U/L, unit of enzyme activity/L.

• 

  Fig. 2 Pathological changes in the myocardial fibers of tested chickens. (A–D) HS group. (E–H) ASA+HS group. (A) Heart tissue at 0 h of HS. (B) After 2 h of HS, the myocardial fibres showed the decreasing in size and granular degeneration characterised by numerous tiny granules in the cytoplasm (arrow). (C) After 5 h, vacuolar degeneration characterized by tiny water droplets in the cytoplasm, and partly dissolved muscle fibrils (arrow) was observed occasionally. (D) After 10 h, necrosis characterized by pyknosis (arrow) was evident. (E) At 0 h of HS, hemangiectasis and hyperemia were observed. (F) After 2 h, myocardial fibers sizes enlarged. (G) After 5 h, granular degeneration and vacuolar degeneration were observed occasionally (arrow). (H) After 10 h, swollen myocardial cells characterized by numerous cytoplasmic particles (arrow) were observed. H&E stain. Scale bar = 10 µm.

• 

  Fig. 3 Distribution Distribution of Hsp90 in the myocardial cells of tested chickens. (A–D) HS group. (I–L) ASA group. (E–H) ASA+HS group. (A) At 0 h of HS, Hsp90 was constitutively expressed in the nucleus and cytoplasm (arrow). (B) After 2 h, Hsp90 expression increased obviously. (C) After 5 h, the maximum intensity of Hsp90 signal weakened. (D) After 24 h, Hsp90 signal showed the second peak. (E) Before HS (0 h), cellular Hsp90 signal was extremely strong. (F) After 2 h of HS, extremely high Hsp90 expression was still observed. (G) After 5 h, cytoplasmic Hsp90 signal decreased while the signal in the nucleus did not (arrow). (H) After 24 h, Hsp90 signal was still present in the nucleus (arrow). (I) At 0 h of test, strong Hsp90 signal was detected in both the nucleus and cytoplasm. (J) The Hsp90 density decreased after 2 h. (K) Hsp90 level rebounded after 5 h, especially in the nucleus (arrow). (L) After 24 h, Hsp90 expression persisted in the cytoplasm and nucleus (arrow). Immunohistochemical staining with hematoxylin counterstain. Scale bar =10 µm.

• 

  Fig. 4 The location of Hsp90 in the blood vessels of the tested chickens. (A) Immunoreactive Hsp90 was observed in the endotheliocyte as well as in the vascular wall of heart artery in the HS group after 1 h of heat stress. (B) Hsp90 was identified in the vein wall cells of the HS group after 1 h of heat stress. (C) The strong, positive signal of the heart aortic endotheliocyte in the ASA+HS group was maintained at 24 h of heat stress. Immunohistochemical staining with hematoxylin counterstain. Scale bar =10 µm.

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  Fig. 5 Relative expression of hsp90 mRNA in different groups. *p < 0.05, **p < 0.01 for the expression of hsp90 mRNA in the HS, ASA+HS and ASA groups at different times compared with that at 0 h in the HS group. Values indicated are the mean ± SD.

• 

  Fig. 6 Levels of Hsp90 in the heart tissues of different groups of broilers. The difference between the expression level of Hsp90 in the HS, ASA+HS and ASA groups at different times of HS and that of 0 h in the HS group. *p < 0.05, **p < 0.01.

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