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In this study, we estimated the distribution of DNA diploidy and aneuploidy in porcine mesenchymal stem cells (pMSCs) that were subjected to osteoblast/osteocyte and adipocyte differentiation to determine the impact of long-term in vitro culture and differentiation on the cell cycle distribution and nuclear DNA profile. This determination could be helpful to confirm or exclude the suitability of physico-chemical culture conditions for the purposes of both the maintenance of an undifferentiated state and to promote differentiation in pMSCs. Flow cytometry was applied to analyze the cell cycle and occurrence of aneuploidy/diploidy, and real-time PCR was used to quantify aP2 and osteocalcin, markers of adipocytes and osteocytes, respectively. The chi-squared test was used to compare the total rates of G0/G1-, S-, and G2/M-phase cell fractions with diploid and aneuploid DNA and the DNA index ratios between three experimental groups of pMSCs. Five weeks of in vitro culture under differentiating conditions resulted in a considerable reduction of DNA stability and a remarkable increase in the rate of cells exhibiting an aneuploid DNA stem line; however, a similar dependence was not found in the nondifferentiated MSCs. Furthermore, the cell fraction rates in each phase of the mitotic cycle and the DNA index (DI) were calculated. The results of real-time PCR for aP2 and osteocalcin proved positive MSC differentiation toward adipocytes and osteocytes. In terms of the possible use of differentiated MSC lines in tissue engineering and regenerative medicine, we propose cytokinetic diagnostics using flow cytometry as an objective and useful method for screening the tumor-forming capacity and malignancy potential of both in vitro long-term cultured MSCs and MSCs subjected to ectopic differentiation. [ABSTRACT FROM AUTHOR]

The qualification of new knowledge is one of the oldest problems in experimental medicine that provides a link between fundamental discovery, hypothesis, 'proof of concept' preclinical studies and development of clinical trials. The biggest challenge in animal models is the proper evaluation of all the aspects that are crucial in specific studied pathologies as well as the prediction of their progression. The aim of this review was to describe and discuss the rat animal model of heart transplant. The rat model of heart transplantation is an excellent yet underestimated method of research of prevention, monitoring and treatment of acute and chronic, immune and nonimmune response to organ transplantation. Despite being a technically and logistically demanding model, it provides a tool for reproducible experiments with longterm animal survival and excellent graft survival., Competing Interests: The authors report no confflict of interest., (Copyright © 2020 Polish Society of Cardiothoracic Surgeons (Polskie Towarzystwo KardioTorakochirurgów) and the editors of the Polish Journal of Cardio-Thoracic Surgery (Kardiochirurgia i Torakochirurgia Polska).)

Introduction: Heart transplant is an accepted treatment modality in end-stage heart failure. The graft coronary artery vasculopathy is a main concern to explain the heterogeneity of the rejection process according to the gender of the donor and recipient., Aim: To assess the severity and type of mechanisms leading to failure of the graft., Material and Methods: Experimental allogenic heart transplantation in the abdomen was performed on Wistar rats depending on the gender of the donor and recipient (F - female; M - male) in four groups (FF, FM, MM MF). The donor heart was implanted in the abdominal cavity of the recipient. Complete time of observation was 10 weeks. Bromodeoxyuridine was administered intraperitoneally to detect proliferating cells., Results: There was 42.5% graft survival in all experiments. The mean time of graft survival was 60 ±18, 54 ±29, 58 ±23 and 64 ±18 days (FF, FM, MM and MF) and no significant difference was found in graft survival time among the four experimental groups ( p = 0.73). None of the heart weight changes reached statistical significance., Conclusions: The use of an animal experimental model helps to understand the mechanisms leading to graft failure and to compare the changes that occur in rats to human hearts. The gender matching affects the survival of the transplanted heart and severity of the graft vasculopathy.

Background: Cardiac surgery is supposed to be a risk factor of PRA formation, however the role of PRA presence in non-transplant subjects is not known. Aim of the study was to assess PRA occurrence in patients undergoing elective cardiosurgery procedures and to evaluate its influence on the perioperative course., Material and Methods: Blood samples were obtained before operation in 44 subjects (36M/8F; 55.9 +/- 8.1 y/o) undergoing primary elective cardiosurgery procedures--CABG (n = 30), CABG + valve (n = 2) or valve procedure (n = 12). PRA results were obtained after the discharge, and patients were retrospectively divided into: Group A (n = 18) with PRA > 1%, and Group B (n = 26) with PRA. < or = 1%. PRA screening was repeated 3 months after the procedure in 41 subjects. They were divided into Group I (n = 13) with PRA > 1%, and Group II (n = 28) with PRA < or = 1%. Comparison was performed of Groups A vs. B, and I vs. II., Results: Differences in pre-operative characteristics and procedure type distribution were insignificant. Post-operative complications were more frequent in Groups A and I (pulmonary hypertension in Group I vs. II: 38 vs. 4%; p = 0.01). Duration of post-operative ICU stay was longer in Group I vs. 11 (2.9 vs. 1.9d.; p = 0.01). Overall hospital stay was longer in Group A vs. B (10.1 vs. 7.8d.; p = 0.054). Increase of PRA titers was observed in 10 subjects (3 pts. /17% from Group A, and 7 pts. /27% from Group B), exceeding 10% in 2 females after valve replacement. 6 months after procedure, detectable PRA was still observed in 7 patients., Conclusions: Cardiac surgery is not a strong causative factor of PRA formation. The presence of perceptible PRA level may be associated with increased incidence of complications and consequently prolonged in-hospital stay. Influence of PRA on peri-operative course is not dependent on the source of its increased level.

Unlabelled: The aim of the study was morphometric and mathematic analysis of calcification profiles present in the leaflets of a cryopreserved and alive heart valve depending on the diagnosed pathologic process. Sheep antibiotic sterilised and cryopreserved biological valves were implanted in tricuspid position in young sheeps for one-year period. After this time the valves removed and studied morphologically. The control group consisted of 7 intact valves, the comparative group, so called group of valves after the processing antibiotic sterilization and cryopreservation consisted of 7 valves after mentioned procedures. Histologic investigations were based on paraffin sections of formalin-fixed valve cusps, stained with H&E and Masson trichrome, calcium deposits were stained von Kossa technique. The measured values included: 1. area and equivalent diameter, 2. length, 3. breadth, 4. perimeter, 5. elongation, 6. roundness, 7. fullness coefficient., Conclusions: 1. A process of initial processing and cryopreservation of biological valve increases a dimension and disturbs a shape of microcalcifications. 2. Cryopreserved biological valves explanted after one-year implantation into an animal in a tricuspid position possess microcalcifications and calcification foci. The size of microcalcifications decreases together with an intensification of degenerative processes of the connective tissue, especially in hyalinization. Hyalinization of the biological valve tissue seems to be favorable for a valve durability and as a pathological process decreasing calcification. 3. Mathematic analysis of morphometric features defining differences in size and shape of each calcification indicate morphologic and morphometric autonomy of calcifications, characteristic for the analyzed group of valve pathologic changes.

Unlabelled: The aim of the study was the characterization of calcification in the leaflets of a cryopreserved and alive heart valve depending on the diagnosed pathologic process. Sheep antibiotic sterilised and cryopreserved biological valves were implanted in tricuspid position in young sheeps for one year period. After this time the valves removed and studied morphologically. The control group consisted of 7 intact valves, the comparative group, so called group of valves after the processing antibiotic sterilization and cryopreservation consisted of 7 valves after mentioned procedures. Histological investigations were based on paraffin sections, calcium deposits were stained von Kossa technique. The measured values included integrative parameters as: 1. area fraction, 2. number of calcifications per area, 3. anisotropy., Conclusions: 1. A process of initial processing, sterilization and cryopreservation of biological valve increases a number of microcalcifications. 2. Cryopreserved biological valves explanted after one-year implantation into an animal in a tricuspid position possess fine calcifications and calcification foci. A number and size of fine calcifications decreases together with an intensification of degeneration and regressive processes of the connective tissue, especially in hyalinization. Hyalinization of the biological valve tissue seems to be favorable for a valve durability and as a pathological process decreasing calcification. 3. Mathematic analysis of morphometric features defining density and structure of calcifications indicate similarities among cryopreservation and initial processing groups, hyalinization, inflammation, whereas in a group of calcification foci, the similarity can be noticed between inflammation and hyalinization group.

Unlabelled: The aim of the study was an evaluation of viability and damage of biological cells sterilized with antibiotics and cryopreserved a year after the implantation. Sheep antibiotic sterilised and cryopreserved biological valves were implanted in tricuspid position in young sheeps for one-year period. After this time the valves removed and studied morphologically. The control group consisted of 7 intact valves, the comparative group, so called group of valves after the processing antibiotic sterilization and cryopreservation consisted of 7 valves after mentioned procedures. Histologic investigations were based on paraffin sections of formalin-fixed valve cusps, stained with H&E and Masson trichrome. Valve viability was assessed using intravital staining with fluoresceine diacetate, whereas damaged cells were visualized by intravital staining with neutral red. Additionally, the ultrastructural studies were performed. The viability and ultrastructural results were compared with the pathologic process in the valve., Conclusions: 1. preliminary preparation with antibiotic sterilisation and cryopresrevation induces valve leaflet oedema and degenerative ultrastructural processes, colliquative cell necrosis and apoptotic morphology of a part of valve cells; 2. cryopreserved and antibiotic sterilised pulmonary valve after one-year implantation behaves lining cells, but ultrastructural changes indicates many degenerative phenomena in smaller degree than after preparation, sterilization and cryopreservation. Histopathologically the degenerative changes were prevalent.