Morphological assessment of actin and desmin expression at different cold myocardial ischemia times: observational study

Abstract

Background. Heart transplantation is currently the treatment of choice for patients in terminal stages of chronic heart failure. The critical shortage of donor organs and the growing need for heart transplantation necessitate the expansion of donor selection criteria, including the estimated ischemia time of the donor heart. Despite numerous studies, the issue remains regarding the safe cold ischemia time; no definite limit to the acceptable preservation time is known and no relevant pathomorphological data are available on the state of the donor heart myocardium at different time parameters. Objective. To comparatively assess the features of cardiomyocyte pathomorphology and expression of protein markers (actin and desmin) in the myocardium of a donor heart prior to the main stage of orthotopic heart transplantation. Methods. The work adopted the design of an observational clinical study, which was prospective in nature. The study used intraoperative myocardial biopsy specimens of the left atrial appendage from donors aged up to 60 years, following cold ischemia of the transplant in Bretschneider solution (Dr. Franz Köhler Chemie GmbH, Germany) lasting up to 240 minutes (Group 1, n = 10) and over 240 minutes (Group 2, n = 7). The nature of pathomorphological myocardial transformation in the left atrial appendage of the donor heart was determined at different cold ischemia times. Histological myocardial sections were stained with hematoxylin and eosin according to standard procedures. After that, they were further studied using light and polarization microscopy; the immunohistochemical method was used to analyze the expression of actin and desmin. Morphometry was performed using the ImageJ 1.48v software (USA). In the analysis of actin and desmin amount, the area of DAB(3,3′-diaminobenzidine)-positive products of the immunohistochemical reaction was estimated as a percentage of the image area. The volume density of immunohistochemically detectable actin and desmin was determined using 20 images at a magnification of 40×10. In order to study the intensity of the immune reaction, a semiquantitative method was used, which involved counting the number of cells in 25 randomly selected fields of view. The types of myocardial contracture damage were assessed via polarization microscopy. Results. Patients included in the first and second groups were comparable in terms of mean age and anthropometric indices. The mean age of patients amounted to 50 [44;59] years in Group 1 and 50 [49;50] years in Group 2, р = 0.193. The body mass index was 25 [22;27] in Group1 and 25 [21;31] in Group 2, р = 0.288. Both groups showed male predominance: 8 (80%) in Group 1 and 6 (85.7%) in Group 2, р = 0.256. The comprehensive morphological assessment of ischemic myocardial damage at different cold ischemia times revealed the uniformity and reversibility of changes in cellular structures (in both groups) that take the form of I–II class contractures, lysis changes in individual cardiomyocytes (only in Group 2), preserved immunohistochemical reactions to actin and desmin in both groups at their average intensity and the complete absence of areas showing no reaction to desmin, which gives an idea about the degree of preservation of their macromolecular structure. Conclusion. The obtained study results showed that due to having a balanced elemental composition that determines the metabolic protection of cells and their ionic balance, the Bretschneider solution effectively protects the donor heart during its transportation, with the myocardial cold ischemia lasting up to 240 min and more.

Keywords

• heart transplant
• donor heart preservation
• cold ischemia time
• actin
• desmin
• morphological diagnostics