Changes Of δPKC Expression During Exercise Preconditioning-induced Early And Late Myocardial Protection In Rats

Objective: A short time, high intensity of intermittent exercise causes the transitory and relative myocardial ischemia and anoxia, which exerts protective effects on the following long time myocardial ischemia and anoxia, this phenomenon's called exercise preconditioning (EP). At present, although EP powerful myocardial protection has been confirmed, its mechanism still needs more researches. Protein kinase C (PKC) isozymeδPKC, as an important medium of cell signal transduction, becomes the focus of EP mechanism research. Researches have already proved theδPKC has myocardial protection and close relationship with exercise. At present, researchers reported little on the changes ofδPKC expression during exercise preconditioning-induced early and late myocardial protection. Therefore, by means of establishing rats EP model and then observing the changes ofδPKC expression during exercise preconditioning-induced early and late myocardial protection by using immunohistochemistry method, this thesis will explore the relationship betweenδPKC and EP myocardial protection effects and mechanism, which will provide new experimental basis for EP protection mechanism researches and new ideas for exercise promoting heart health researches.Methods: 150 SD rats, are randomly divided into the control group (Group C), exhaustive exercise group (Group EE), early exercise preconditioning Group (Group EEP), late exercise preconditioning Group (Group LEP), early exercise preconditioning plus exhaustive exercise group (Group EEP+EE) and late exercise preconditioning plus exhaustive exercise group (Group LEP+EE). Group C: general feeding; Group EE: one time exhaustive exercise causing acute myocardial injury; Group EEP and Group LEP: one short time high-intensity intermittent exercise on running machine establishing rats EP model; Group EEP+EE: exercise according to the EP model, after 30mins'rest, doing exhaustive exercise; Group LEP+EE: exercise according to the EP model, after 24hs'rest, doing exhaustive exercise. Group C, Group EE, Group EEP Group EEP+EE and Group LEP+EE: sacrificed 30mins after exercise; Group LEP: sacrificed 24hs after exercise. With the method of immunochemiluminometry, HE Staining and myocardial ischemia and anoxia staining (Hematoxylin Basic Fuchsin Picric Acid Staining, HBFP Staining),the research will respectively detect the changes of cardiac troponin I (cTnI) content in rats, and observe the changes of rats myocardial structure and myocardial ischemia and anoxia, and comprehensively evaluate the myocardial injury; With the method of immunohistochemistry, the research will show the expression of myocardial cellδPKC and p-δPKC; with the image analysis technique on the expression ofδPKC and p-δPKC, the research will do the half quantitative analysis.Results: 1. Comprehensive detection on rats myocardial injury: The morphological structure of myocardial cells in group C is normal, no ischemia or anoxia; compared with group C, cTnI in group EE significantly increased (P<0.05), myocardial cells is distorted and its cross striation vagued,showing block or round red spots of myocardial ischemia and anoxia changes; compared with group EE, cTnI significantly decreased in group EEP+EE and group LEP+EE (P<0.05), in group EEP+EE, the changes of myocardial ischemia and anoxia decreased, in group LEP+EE, myocardial cells was distorted, the structural integrity was damaged, and changes of myocardial ischemia and anoxia increased. 2. The immunohistochemistry and images analysis ofδPKC in myocardial cells: group C: immune positive signal ofδPKC was in tan particles, scattering in a few myocardial cells cytoplasm; group EE: immune positive signal ofδPKC enhanced, tan particles in cell cytoplasm bigger and more, in puce; group EEP and group LEP: immune positive signal ofδPKC enhanced compared with group C, most cells observed scattering; group EEP+EE and group LEP+EE immune positive signal ofδPKC enhanced compared with group EE in puce. Area and intergral optical density (IOD) ofδPKC immune response positive signal: compared with group C, in group EE, group EEP, group EEP+EE, group LEP and group LEP+EE, expression area and IOD both significantly increased(P<0.05); compared with group EE, in group EEP and group LEP, expression area and IOD significantly increased(P<0.05); compared with group EEP, in group LEP, expression area and IOD is a little low, having no significant difference (P>0.05);compared with group EEP+EE, in group LEP+EE, expression area and IOD is a little low, having no significant difference ( P>0.05 ) . 3. The Immunohistochemistry and images analysis of myocardial cells p-δPKC: group C: immune positive signal of p-δPKC in myocardial cell was in tan particles, pointlike scattering in myocardial cells cytoplasm, a few on intercalated disc; in group EE, immune positive signal expressed on intercalated disc and cell membrane was more, no on karyotheca; in group EEP, Immune positive signal expressed on intercalated disc and cell membrane was more, appearing obvious hyperchromatosis; compared with group EE, in group EEP+EE immune positive signal was obvious more, appearing obvious hyperchromatosis, most of which is on the intercalated disc and cell membrane; compared with group EEP, in group LEP, immune positive signal was obvious less, no hyperchromatosis, most of which was on the intercalated disc and cell membrane; compared with group EE, in group LEP+EE, immune positive signal was obvious less, no hyperchromatosis, most spotlike scattering in cytoplasm, a few on the intercalated disc and cell membrane. Area and IOD of p-δPKC immune response positive signal: compared with group C, in group EEP and group EEP+EE, expression area and IOD both increased, in group LEP+EE, expression area and IOD both significantly decreased(P<0.05); compared with group EE, in group EEP+EE, expression area and IOD increased, in group LEP+EE, expression area and IOD decreased significantly(P<0.05); compared with group EEP, in group LEP, expression area and IOD significantly decreased(P<0.05);compared with group EEP+EE, in group LEP+EE, expression area and IOD significantly decreased(P<0.05).Conclusion: 1. During the early protective phase of EP, the expression ofδPKC and p-δPKC increased, and p-δPKC translocated to intercalated disc and nucleus, which indicates thatδPKC mediates the early cardioprotective effect induced by EP; 2. During the late protective phase of EP, the expression ofδPKC increased, p-δPKC restored to normal, but still expressing on intercalated disc, suggests thatδPKC maybe play a role in the late cardioprotective effect induced by EP ; 3. Exercise preconditioning has early protective effect on acute myocardial injury induced by exhaustive exercise. Late protective effects of exercise preconditioning on acute myocardial injury induced by exhaustive exercise have yet to be further confirmed.