Heterogeneous loss of connexin43 protein in ischemic dog hearts

Introduction: Ischemia causes cell decoupling in the myocardium. Prolonged ischemia activates proteases and causes degradation of structural proteins as well as gap junctions. There is little information about the degradation of gap junction protein during the early time period after acute ischemia, The purpose of the present study was to investigate connexin43 (Cx43) prot ein degradation and distribution patterns in the canine left ventricular wa ll during 1 to 6 hours of ischemia. Methods and Results: Ischemia of canine left ventricular myocardium was ind uced by ligation of the left anterior descending coronary artery, Following a period of in situ ischemia of up to 6 hours, samples were harvested, and standard paraffin slides were prepared for Cx43 and wheat germ agglutinin double labeling, Cx43 distribution was visualized by confocal microscopy, I n controls, homogeneous distribution of Cx43 staining was determined, Ische mia caused a loss of Cx43 with a heterogeneous pattern by mixing foci of in farcted cells among normal cardiac myocytes, To determine if the changes we re induced by heterogeneous reduction in the blood supply, an in vitro isch emic model was studied to induce more homogeneous ischemia, Western blot an alysis of these In vitro ischemic tissue samples revealed a reduction of Cx 43 protein concentration,vith a 50% decay time of 4.8 hours. Cx43 dephospho rylation was detected after 1 hour of in vitro ischemia, heterogeneous loss of Cx43 was found in the in vitro ischemic tissue, There were no significa nt changes in Cx43 staining density during the first hour of ischemia at a time when dephosphorylation of the protein was observed, After 1 hour of is chemia, Cx43 was reduced at intercalated disk areas, and, after 6 hours, mo st Cx43 disappeared at intercalated disk areas, while small amounts of Cx43 remained at side-to-side junctions. Conclusion: Cx43 undergoes both distribution and concentration changes foll owing acute cardiac ischemia, The loss of Cx43 protein is heterogeneous. Cx 43 dephosphorylation occurred within 1 hour following ischemia.