[Effect of extracellular matrix components on adhesion of bone marrow multipotent mesenchymal stromal cells to polytetrafluoroethylene].

Search for new bioengineering materials for creation of small-diameter vascular grafts is currently a priority task. One of the promising trends of creating tissue engineering constructions is coating the internal layer of implants made of polytetrafluoroethylene (PTFE) with autologous mesenchymal multipotent stromal cells. In the study we assessed the ability of separate components of the extracellular matrix such as fibronectin, type I collagen and type IV collagen to influence adhesion, proliferation and morphology of mesenchymal multipotent stromal cells being cultured on PTFE. Bone marrow multipotent stromal cells taken from second-passage Wistar rats in the amount of 106 per 1 cm2 were applied onto PTFE. We used the following variants of preliminary treatment of the material prior to seeding: fibronectin with type I collagen, fibronectin with type IV collagen, fibronectin with a mixture of type I and IV collagens, as well as a control group without coating. After six weeks of cell growth on PTFE patches the samples were subjected to fixation in 10% formalin followed by haematoxylin-eosin stain and morphometric assessment of adhered cells by calculation using the software AxioVision (Carl Zeiss), assessing the number of cells, area of the cellular monolayer, dimensions and ratios of the area of separate cells and the area of cellular nuclei. The maximal area of the monolayer from mesenchymal multipotent stromal cells on the PTFE surface was revealed while culturing with a mixture of fibronectin and type I and IV collagens. Cell colonization density while treatment of the synthetic material with mixtures of fibronectin with type I collagen, type IV collagen and type I and IV collagens demonstrated the results exceeding the parameters of the control specimen 5-, 2.5- and 7-fold, respectively. Hence, extracellular matrix components considerably increase enhance adhesion of cells to PTFE, as well as improve formation of a monolayer from mesenchymal multipotent stromal cells.

Therapy of Chronic Cardiosclerosis in WAG Rats Using Cultures of Cardiovascular Cells Enriched with Cardiac Stem Cell.

We developed a protocol for preparing cardiac cell culture from rat heart enriched with regional stem cells based on clonogenic properties and proliferation in culture in a medium with low serum content. Experiments on WAG rats with experimental ischemic myocardial damage showed that implantation of autologous regional stem cells into the left ventricle reduced the volume of cicatricial tissue, promoted angiogenesis in the damaged zone, and prevented the risk of heart failure development.

[Study of patency of vascular grafts manufactured by means of electrospinning].

UNLABELLED: In vivo experiments were carried out to study functioning of vascular grafts manufactured by means of electrospinning from solutions of polycaprolactone (PCL) and hexafluoroisopropanol (HFIP), PCL with 10% gelatine and a low-permeability inner layer (LPIL) 10 µm thick and PCL with 10% gelatine and LPIL (10 µm) wherein as polymeric base instead of PCL copolymer of lactic and hydroxyacetic acids (polylactide-co-glycolide, PLGA) was used. The grafts were implanted into the infrarenal portion of the aorta to 45 rats, 15 rats for each type of the graft. Patency of artificial vessels was assessed by means of magnetic resonance tomography and diagnostic ultrasound Dopplerography at 2, 4 and 20 weeks (5 animals for each time point). The state of the graft and surrounding tissues was analysed by means of intraoperative assessment, survey microscopy and survey fluorescent microscopy. CONCLUSION: The obtained findings demonstrated that vascular grafts made by electrospinning technique with a low-permeability inner layer are less prone to formation of the neointima and stenosing as compared with grafts having no such layer.

[Use of laser technologies in treatment of chronic venous insufficiency in patients with a wide ostial segment of the main trunks of subcutaneous veins].

Presented herein is the authors' experience in endovascular laser obliteration of the major trunks of the grate saphenous vein (GSV) with a wide ostial segment (measuring from 15 to 34 mm) in patients presenting with chronic venous insufficiency. Group One patients (n=32) underwent crossectomy followed by endovasal laser obliteration (EVLO) of the GSV's trunk on the femur. Group Two patients (n=46) were not subjected to crossectomy, whereas obliteration of the GSV's trunk was carried out immediately from the ostium. In Group One we managed to achieve obliteration of the GSV's trunk in 32 patients (100%) with no additional interventions, and in Group Two this was achieved in 42 (91.3%) patients. Four patients (8.7%) required performing a secondary procedure of EVLO after which obliteration of the trunk was achieved in all patients of Group Two. There was no evidence of deep-vein thrombosis.

Expression of mRNA of apolipoprotein E, apolipoprotein A-IV, and matricellular proteins in the myocardium and intensity of fibroplastic processes during experimental hypercholesterolemia.

The expression of mRNA of matricellular proteins (osteopontin, and lumican), apolipoproteins E and A-IV, and microsomal triglyceride transfer protein, and the intensity of fibroplastic processes were studied in the myocardium of rats during experimental chronic hypercholesterolemia. We have found that the development of chronic hypercholesterolemia was followed by an increase in volume density of interstitial connective tissue in the myocardium reflecting the activation of fibroplastic processes. A slight positive correlation was observed between the connective tissue density in the myocardium and expression of osteopontin mRNA (r=0.408) and lumican mRNA (r=0.470). Myocardium remodeling during hypercholesterolemia is realized against the background of increased expression of apolipoproteins E and A-IV mRNA and microsomal triglyceride transfer protein mRNA involved in transport and metabolism of lipoproteins in several tissues and probably play a pivotal role in the regulation of lipoprotein transport and metabolism in the myocardium. We concluded that the increase in the expression of apolipoproteins (E and A-IV) and microsomal triglyceride transfer protein play adaptive and compensatory role and is related to the increase in lipoprotein utilization by macrophages.

Stimulation of angiogenesis in rat ischemic limb by intramuscular implantation of mononuclear fraction cells from autologous bone marrow.

We studied angiogenic effects of two variants of indirect revascularization (transplantation of mononuclear fraction cells from autologous bone marrow alone or in combination with laser tunneling) after modeled ischemia of the lower limbs in rats. Doppler sonography and immunofluorescence studies showed that intramuscular implantation of isolated fraction of autologous bone marrow mononuclear cells in combination with laser tunneling of the muscles is most effective and can be recommended as a method of angiogenesis stimulation in non-reconstructable distal vascular pathology of the extremities.

Morphological and molecular analysis of angiogenesis after intramyocardial transplantation of autologous bone marrow mononuclear cells.

We studied the peculiarities of angiogenesis in the postinfarction period after transmyocardial laser revascularization and intramyocardial implantation of mononuclear bone marrow cells into the pericicatrical zone of the left ventricular myocardium in dogs. Morphological manifestation of angiogenesis in the myocardium after application of laser and cell technologies are angiomatosis, formation of large thin-wall vessels and sinusoids. The angiogenic effect of implanted mononuclear bone marrow cells is determined by high content (43-47%) of CD31+ cells in both adherent and nonadherent fractions. More pronounced angiogenic potential of nonadherent cells is determined by intensive expression of cytokine VEGF-B and D mRNA essential for arterial vessels growth. Immunohistochemical studies showed that about 90% cells of the nonadherent fraction are endothelial precursors expressing endothelial cell markers isolectin B4 and VEGF-R2. It was found that the use of adherent mononuclear bone marrow cells during the postinfarction period induces ossification of the epicardium and subepicardial myocardium layer, formation of cartilage plates, and focal calcification. Implantation of nonadherent mononuclear bone marrow cells into transmyocardial laser channels did not induce ectopic ossification of the myocardium.

Angiomatosis and ectopic ossification in the dog myocardium after intramyocardial implantation of autologous bone marrow mononuclears in experimental coronary disease.

Disseminated ossification of the myocardium and severe angiomatosis were detected in sites of implantation of unseparated bone marrow mononuclear fraction cells in dogs with experimental chronic coronary disease. Minor immunocytochemical differences in cells of the mononuclear fraction adhering and not adhering to plastic were found. Significant differences in the expression of mRNA of chondro-osteogenesis genes (aggrecan, lumican, and osteopontin) in adherent and nonadherent cells were detected. The expression of aggrecan gene mRNA was 3-fold lower, of lumican gene 6-fold, and of osteopontin gene 11-fold lower in nonadherent cells compared to adherent fraction.

Morphological evaluation of cytoarchitectonics of aortic graft at the biotechnological stage with analysis of changes in laser-induced fluorescence spectra.

We performed a fluorescent microscopic examination of human and animal aortic grafts at different stages of decellularization. Treatment of aortic grafts with trypsin/EDTA solution for 48 h leads to their complete decellularization and preserved the connective tissue fiber backbone, which gains a netlike structure. The use of this protocol of decellularization leads to disappearance of subintimal calcium deposits in human aortic grafts. Differences in laser-induced fluorescence spectra of aortas before decellularization and at different stages of this process were revealed. Our findings suggest that the use of fluorescence induced by excimer lasers is promising for identification of the composition of biological tissues, analysis of their state, and the presence of changes.