Heart stem cells: hope or myth?

The search and study of endogenous heart repair remains an urgent issue in modern regenerative medicine. It is generally accepted that the human heart has a limited regenerative potential, but recent studies show that functionally significant regeneration is possible. However, the mechanisms underlying these processes remain poorly understood. In the heart, there are populations of resident mesenchymal cells that have some properties of stem cells that carry certain markers, such as c-kit+, Sca-1, etc. The ability of these cells to differentiate directly into cardiomyocytes remains controversial, but their use in clinical trials has shown improved cardiac function in patients with myocardial infarction. Currently, approaches are being developed to use, mainly, induced pluripotent stem cells as a promising regenerative therapy, but the cardioprotective role of cardiac mesenchymal cells remains the subject of active study due to their paracrine signaling

Magnetron plasma mediated immobilization of hyaluronic acid for the development of functional double-sided biodegradable vascular graft

The clinical need for vascular grafts is associated with cardiovascular diseases frequently leading to fatal outcomes. Artificial vessels based on bioresorbable polymers can replace the damaged vascular tissue or create a bypass path for blood flow while stimulating regeneration of a blood vessel in situ. However, the problem of proper conditions for the cells to grow on the vascular graft from the adventitia while maintaining its mechanical integrity of the luminal surface remains a challenge. In this work, we propose a two-stage technology for processing electrospun vascular graft from polycaprolactone, which consists of plasma treatment and subsequent immobilization of hyaluronic acid on its surface producing thin double-sided graft with one hydrophilic and one hydrophobic side. Plasma modification activates the polymer surfaces and produces a thin layer for linker-free immobilization of bioactive molecules, thereby producing materials with unique properties. The proposed modification does not affect the morphology or mechanical properties of the graft and improves cell adhesion. The proposed approach can potentially be used for various biodegradable polymers such as polylactic acid, polyglycolide, and their copolymers and blends, with a hydrophilic inner surface and a hydrophobic outer surface

Transfer of synthetic human chromosome into human induced pluripotent stem cells for biomedical applications

Alphoid(tetO)-type human artificial chromosome (HAC) has been recently synthetized as a novel class of gene delivery vectors for induced pluripotent stem cell (iPSC)-based tissue replacement therapeutic approach. This HAC vector was designed to deliver copies of genes into patients with genetic diseases caused by the loss of a particular gene function. The alphoid(tetO)-HAC vector has been successfully transferred into murine embryonic stem cells (ESCs) and maintained stably as an independent chromosome during the proliferation and differentiation of these cells. Human ESCs and iPSCs have significant differences in culturing conditions and pluripotency state in comparison with the murine naïve-type ESCs and iPSCs. To date, transferring alphoid(tetO)-HAC vector into human iPSCs (hiPSCs) remains a challenging task. In this study, we performed the microcell-mediated chromosome transfer (MMCT) of alphoid(tetO)-HAC expressing the green fluorescent protein into newly generated hiPSCs. We used a recently modified MMCT method that employs an envelope protein of amphotropic murine leukemia virus as a targeting cell fusion agent. Our data provide evidence that a totally artificial vector, alphoid(tetO)-HAC, can be transferred and maintained in human iPSCs as an independent autonomous chromosome without affecting pluripotent properties of the cells. These data also open new perspectives for implementing alphoid(tetO)-HAC as a gene therapy tool in future biomedical applications

Protective Role of Mytilus edulis Hydrolysate in Lipopolysaccharide-Galactosamine Acute Liver Injury

Acute liver injury in its terminal phase trigger systemic inflammatory response syndrome with multiple organ failure. An uncontrolled inflammatory reaction is difficult to treat and contributes to high mortality. Therefore, to solve this problem a search for new therapeutic approaches remains urgent. This study aimed to explore the protective effects of M. edulis hydrolysate (N2-01) against Lipopolysaccharide-D-Galactosamine (LPS/D-GalN)-induced murine acute liver injure and the underlying mechanisms. N2-01 analysis, using Liquid Chromatography Mass Spectrometry (LCMS) metabolomic and proteomic platforms, confirmed composition, molecular-weight distribution, and high reproducibility between M. edulis hydrolysate manufactured batches. N2-01 efficiently protected mice against LPS/D-GalN-induced acute liver injury. The most prominent result (100% survival rate) was obtained by the constant subcutaneous administration of small doses of the drug. N2-01 decreased Vascular Cell Adhesion Molecule-1 (VCAM-1) expression from 4.648 ± 0.445 to 1.503 ± 0.091 Mean Fluorescence Intensity (MFI) and Interleukin-6 (IL-6) production in activated Human Umbilical Vein Endothelial Cells (HUVECs) from 7.473 ± 0.666 to 2.980 ± 0.130 ng/ml in vitro. The drug increased Nitric Oxide (NO) production by HUVECs from 27.203 ± 2.890 to 69.200 ± 4.716 MFI but significantly decreased inducible Nitric Oxide Synthase (iNOS) expression from 24.030 ± 2.776 to 15.300 ± 1.290 MFI and NO production by murine peritoneal lavage cells from 6.777 ± 0.373 µm to 2.175 ± 0.279 µm. The capability of the preparation to enhance the endothelium barrier function and to reduce vascular permeability was confirmed in Electrical Cell-substrate Impedance Sensor (ECIS) test in vitro and Miles assay in vivo. These results suggest N2-01 as a promising agent for treating a wide range of conditions associated with uncontrolled inflammation and endothelial dysfunction

A short G1 phase is an intrinsic determinant of naïve embryonic stem cell pluripotency

AbstractA short G1 phase is a characteristic feature of mouse embryonic stem cells (ESCs). To determine if there is a causal relationship between G1 phase restriction and pluripotency, we made use of the Fluorescence Ubiquitination Cell Cycle Indicator (FUCCI) reporter system to FACS-sort ESCs in the different cell cycle phases. Hence, the G1 phase cells appeared to be more susceptible to differentiation, particularly when ESCs self-renewed in the naïve state of pluripotency. Transitions from ground to naïve, then from naïve to primed states of pluripotency were associated with increased durations of the G1 phase, and cyclin E-mediated alteration of the G1/S transition altered the balance between self-renewal and differentiation. LIF withdrawal resulted in a lengthening of the G1 phase in naïve ESCs, which occurred prior to the appearance of early lineage-specific markers, and could be reversed upon LIF supplementation. We concluded that the short G1 phase observed in murine ESCs was a determinant of naïve pluripotency and was partially under the control of LIF signaling

Участие транскрипционного фактора ZBTB16 в процессах физиологического образования костной ткани и при патологической кальцификации аортального клапана

Degenerative calcific aortic valve stenosis is the most common type of heart valve disease in the Western world. Patients with severe stenosis are associated with 50 percent chance of mortality within two years in the absence of intervention. Surgical interventions are the only treatment method for severe calcific aortic valve stenosis to date. Pharmacological approaches have so far failed to affect the course of the disease. Thus, there is an urgent need to develop novel treatment strategies that could slow down the progression of the stenosis. ZBTB16 is a zinc finger protein with N-term BTB/POZ domain (protein-protein interaction motif) and 9 zinc finger domains (DNA binding motif) in C-term. There is growing evidence proving the participation of ZBTB16 in skeletal development. ZBTB16 has been shown to play a role in the specification of limb and axial skeleton patterning. Moreover, the expression of ZBTB16 is increased in patients with ectopic bone formation. Nowadays, the evidence supports that the mechanisms that play key roles in the formation of bone tissue are similar to the processes occurring during the development of ectopic ossification of the aortic valve. Thus, it can be assumed that ZBTB16 is heavily involved in osteogenic transformation in the aortic valve. Understanding similarities and differences in the mechanisms that mediate osteogenic differentiation of stem cells during bone formation and pathological ossification of tissues can help to find the ways to control the osteogenic differentiation in the human body. The aim of this review is to summarize data on the role of ZBTB16 and its products in the regulation of differentiation and proliferation of cells involved in osteogenesis and in the development of ectopic calcification of the aortic valve. The study of the dynamic changes of ZBTB16 expression in aortic valve calcification is a new and relevant study field.Дегенеративный кальцинированный стеноз аортального клапана – наиболее распространенная в мире патология клапанов сердца. При развитии кальцинированного стеноза аортального клапана прогноз двухлетней выживаемости без хирургического вмешательства составляет 50%. Единственным на сегодняшний день методом лечения тяжелой кальцификации клапана выступает хирургическое вмешательство. Фармакологические подходы до сих пор не смогли изменить течение заболевания, поэтому разработка новых стратегий лечения, замедляющих развитие стеноза аортального клапана, представляет актуальную клиническую потребность. Белок ZBTB16 является транскрипционным фактором с N-концевым BTB/POZ-доменом для белок-белкового взаимодействия и девятью C-концевыми доменами типа цинковый палец для связывания ДНК. В литературе представлены данные об участии ZBTB16 в развитии скелета. Показано, что ZBTB16 играет роль в спецификации паттернов аксиального скелета и конечностей. Кроме того, экспрессия ZBTB16 повышена в клетках пациентов, страдающих эктопическим формированием костной ткани. На сегодняшний день мы имеем множество подтверждений тому, что ключевые механизмы формирования костной ткани в норме, сходны с процессами при эктопической оссификации тканей аортального клапана. Таким образом, можно сделать предположение о значительном участии ZBTB16 и в остеогенной трансформации клеток клапана аорты. Понимание сходств и различий механизмов, опосредующих остеогенную дифференцировку клеток во время физиологического формирования кости и патологической оссификации тканей, может дать предпосылки для возможности управления процессами остеогенной дифференцировки в организме человека. Целью данного обзора стало обобщение сведений о роли ZBTB16 и его продуктов в регулировании дифференцировки и пролиферации клеток, участвующих в процессах физиологического остеогенеза и при эктопической кальцификации тканей, в том числе аортального клапана. Изучение динамической вариабельности экспрессии ZBTB16 в клетках аортального клапана при кальцификации ткани является новым и актуальным направлением

Inflammation and Mechanical Stress Stimulate Osteogenic Differentiation of Human Aortic Valve Interstitial Cells

Background: Aortic valve calcification is an active proliferative process, where interstitial cells of the valve transform into either myofibroblasts or osteoblast-like cells causing valve deformation, thickening of cusps and finally stenosis. This process may be triggered by several factors including inflammation, mechanical stress or interaction of cells with certain components of extracellular matrix. The matrix is different on the two sides of the valve leaflets. We hypothesize that inflammation and mechanical stress stimulate osteogenic differentiation of human aortic valve interstitial cells (VICs) and this may depend on the side of the leaflet.Methods: Interstitial cells isolated from healthy and calcified human aortic valves were cultured on collagen or elastin coated plates with flexible bottoms, simulating the matrix on the aortic and ventricular side of the valve leaflets, respectively. The cells were subjected to 10% stretch at 1 Hz (FlexCell bioreactor) or treated with 0.1 μg/ml lipopolysaccharide, or both during 24 h. Gene expression of myofibroblast- and osteoblast-specific genes was analyzed by qPCR. VICs cultured in presence of osteogenic medium together with lipopolysaccharide, 10% stretch or both for 14 days were stained for calcification using Alizarin Red.Results: Treatment with lipopolysaccharide increased expression of osteogenic gene bone morphogenetic protein 2 (BMP2) (5-fold increase from control; p = 0.02) and decreased expression of mRNA of myofibroblastic markers: α-smooth muscle actin (ACTA2) (50% reduction from control; p = 0.0006) and calponin (CNN1) (80% reduction from control; p = 0.0001) when cells from calcified valves were cultured on collagen, but not on elastin. Mechanical stretch of VICs cultured on collagen augmented the effect of lipopolysaccharide. Expression of periostin (POSTN) was inhibited in cells from calcified donors after treatment with lipopolysaccharide on collagen (70% reduction from control, p = 0.001), but not on elastin. Lipopolysaccharide and stretch both enhanced the pro-calcific effect of osteogenic medium, further increasing the effect when combined for cells cultured on collagen, but not on elastin.Conclusion: Inflammation and mechanical stress trigger expression of osteogenic genes in VICs in a side-specific manner, while inhibiting the myofibroblastic pathway. Stretch and lipopolysaccharide synergistically increase calcification

Generation of two iPSC lines (FAMRCi007-A and FAMRCi007-B) from patient with Emery-Dreifuss muscular dystrophy and heart rhythm abnormalities carrying genetic variant LMNA p.Arg249Gln.

Human iPSC lines were generated from peripheral blood mononuclear cells of patient carrying LMNA mutation associated with Emery–Dreifuss muscular dystrophy accompanied by atrioventricular block and paroxysmal atrial fibrillation. Reprogramming factors OCT4, KLF4, SOX2, CMYC were delivered using Sendai virus transduction. iPSCs were characterized in order to prove the pluripotency markers expression, normal karyotype, ability to differentiate into three embryonic germ layers. Generated iPSC lines would be useful model to investigate disease development associated with genetic variants in LMNA gene