Multi-Omics Profiling of Human Endothelial Cells from the Coronary Artery and Internal Thoracic Artery Reveals Molecular but Not Functional Heterogeneity.

Major adverse cardiovascular events occurring upon coronary artery bypass graft surgery are typically accompanied by endothelial dysfunction. Total arterial revascularisation, which employs both left and right internal thoracic arteries instead of the saphenous vein to create a bypass, is associated with better mid- and long-term outcomes. We suggested that molecular profiles of human coronary artery endothelial cells (HCAECs) and human internal mammary artery endothelial cells (HITAECs) are coherent in terms of transcriptomic and proteomic signatures, which were then investigated by RNA sequencing and ultra-high performance liquid chromatography-mass spectrometry, respectively. Both HCAECs and HITAECs overexpressed molecules responsible for the synthesis of extracellular matrix (ECM) components, basement membrane assembly, cell-ECM adhesion, organisation of intercellular junctions, and secretion of extracellular vesicles. HCAECs were characterised by higher enrichment with molecular signatures of basement membrane construction, collagen biosynthesis and folding, and formation of intercellular junctions, whilst HITAECs were notable for augmented pro-inflammatory signaling, intensive synthesis of proteins and nitrogen compounds, and enhanced ribosome biogenesis. Despite HCAECs and HITAECs showing a certain degree of molecular heterogeneity, no specific markers at the protein level have been identified. Coherence of differentially expressed molecular categories in HCAECs and HITAECs suggests synergistic interactions between these ECs in a bypass surgery scenario.

Endothelial Cell Markers Are Inferior to Vascular Smooth Muscle Cells Markers in Staining Vasa Vasorum and Are Non-Specific for Distinct Endothelial Cell Lineages in Clinical Samples.

Current techniques for the detection of vasa vasorum (VV) in vascular pathology include staining for endothelial cell (EC) markers such as CD31 or VE-cadherin. However, this approach does not permit an objective assessment of vascular geometry upon vasospasm and the clinical relevance of endothelial specification markers found in developmental biology studies remains unclear. Here, we performed a combined immunostaining of rat abdominal aorta (rAA) and human saphenous vein (hSV) for various EC or vascular smooth muscle cell (VSMC) markers and found that the latter (e.g., alpha smooth muscle actin (α-SMA) or smooth muscle myosin heavy chain (SM-MHC)) ensure a several-fold higher signal-to-noise ratio irrespective of the primary antibody origin, fluorophore, or VV type (arterioles, venules, or capillaries). Further, α-SMA or SM-MHC staining allowed unbiased evaluation of the VV area under vasospasm. Screening of the molecular markers of endothelial heterogeneity (mechanosensitive transcription factors KLF2 and KLF4, arterial transcription factors HES1, HEY1, and ERG, venous transcription factor NR2F2, and venous/lymphatic markers PROX1, LYVE1, VEGFR3, and NRP2) have not revealed specific markers of any lineage in hSV (although KLF2 and PROX1 were restricted to venous endothelium in rAA), suggesting the need in high-throughput searches for the clinically relevant signatures of arterial, venous, lymphatic, or capillary differentiation.

Landmark native breed of the Orenburg goats: progress in its breeding and genetics and future prospects.

This paper reviews information about a unique and iconic breed of the Orenburg Oblast, the homeland and the only place where the best herds of Orenburg down-hair goats in Russia are concentrated. Three types of these small ruminant animals are widespread on the territory of the region: Orenburg purebred gray goats, Orenburg purebred white goats, as well as crossbred white goats of F1 White Don × White Orenburg. Currently, at the farms of the Orenburg region, animals are selected according to their phenotype, with selected traits being color, weight and length of down hair. In recent years, the Orenburg goat breed has become an object of genetic research using various marker systems including immunogenetic, microsatellite, mtDNA and SNP markers. Overall, these studies evidence about the uniqueness of the allele pool in the landmark native breed of the Orenburg goats, which is a complex dynamic genetic system, prioritizing its further in-depth genome research and breeding applications.

Excessive Adventitial and Perivascular Vascularisation Correlates with Vascular Inflammation and Intimal Hyperplasia.

Albeit multiple studies demonstrated that vasa vasorum (VV) have a crucial importance in vascular pathology, the informative markers and metrics of vascular inflammation defining the development of intimal hyperplasia (IH) have been vaguely studied. Here, we employed two rat models (balloon injury of the abdominal aorta and the same intervention optionally complemented with intravenous injections of calciprotein particles) and a clinical scenario (arterial and venous conduits for coronary artery bypass graft (CABG) surgery) to investigate the pathophysiological interconnections among VV, myeloperoxidase-positive (MPO+) clusters, and IH. We found that the amounts of VV and MPO+ clusters were strongly correlated; further, MPO+ clusters density was significantly associated with balloon-induced IH and increased at calciprotein particle-provoked endothelial dysfunction. Likewise, number and density of VV correlated with IH in bypass grafts for CABG surgery at the pre-intervention stage and were higher in venous conduits which more frequently suffered from IH as compared with arterial grafts. Collectively, our results underline the pathophysiological importance of excessive VV upon the vascular injury or at the exposure to cardiovascular risk factors, highlight MPO+ clusters as an informative marker of adventitial and perivascular inflammation, and propose another mechanistic explanation of a higher long-term patency of arterial grafts upon the CABG surgery.

Calciprotein Particles Link Disturbed Mineral Homeostasis with Cardiovascular Disease by Causing Endothelial Dysfunction and Vascular Inflammation.

An association between high serum calcium/phosphate and cardiovascular events or death is well-established. However, a mechanistic explanation of this correlation is lacking. Here, we examined the role of calciprotein particles (CPPs), nanoscale bodies forming in the human blood upon its supersaturation with calcium and phosphate, in cardiovascular disease. The serum of patients with coronary artery disease or cerebrovascular disease displayed an increased propensity to form CPPs in combination with elevated ionised calcium as well as reduced albumin levels, altogether indicative of reduced Ca2+-binding capacity. Intravenous administration of CPPs to normolipidemic and normotensive Wistar rats provoked intimal hyperplasia and adventitial/perivascular inflammation in both balloon-injured and intact aortas in the absence of other cardiovascular risk factors. Upon the addition to primary human arterial endothelial cells, CPPs induced lysosome-dependent cell death, promoted the release of pro-inflammatory cytokines, stimulated leukocyte adhesion, and triggered endothelial-to-mesenchymal transition. We concluded that CPPs, which are formed in the blood as a result of altered mineral homeostasis, cause endothelial dysfunction and vascular inflammation, thereby contributing to the development of cardiovascular disease.

EMbedding and Backscattered Scanning Electron Microscopy: A Detailed Protocol for the Whole-Specimen, High-Resolution Analysis of Cardiovascular Tissues.

Currently, an ultrastructural analysis of cardiovascular tissues is significantly complicated. Routine histopathological examinations and immunohistochemical staining suffer from a relatively low resolution of light microscopy, whereas the fluorescence imaging of plaques and bioprosthetic heart valves yields considerable background noise from the convoluted extracellular matrix that often results in a low signal-to-noise ratio. Besides, the sectioning of calcified or stent-expanded blood vessels or mineralised heart valves leads to a critical loss of their integrity, demanding other methods to be developed. Here, we designed a conceptually novel approach that combines conventional formalin fixation, sequential incubation in heavy metal solutions (osmium tetroxide, uranyl acetate or lanthanides, and lead citrate), and the embedding of the whole specimen into epoxy resin to retain its integrity while accessing the region of interest by grinding and polishing. Upon carbon sputtering, the sample is visualised by means of backscattered scanning electron microscopy. The technique fully preserves calcified and stent-expanded tissues, permits a detailed analysis of vascular and valvular composition and architecture, enables discrimination between multiple cell types (including endothelial cells, vascular smooth muscle cells, fibroblasts, adipocytes, mast cells, foam cells, foreign-body giant cells, canonical macrophages, neutrophils, and lymphocytes) and microvascular identities (arterioles, venules, and capillaries), and gives a technical possibility for quantitating the number, area, and density of the blood vessels. Hence, we suggest that our approach is capable of providing a pathophysiological insight into cardiovascular disease development. The protocol does not require specific expertise and can be employed in virtually any laboratory that has a scanning electron microscope.

Co-Culture of Primary Human Coronary Artery and Internal Thoracic Artery Endothelial Cells Results in Mutually Beneficial Paracrine Interactions.

Although saphenous veins (SVs) are commonly used as conduits for coronary artery bypass grafting (CABG), internal thoracic artery (ITA) grafts have significantly higher long-term patency. As SVs and ITA endothelial cells (ECs) have a considerable level of heterogeneity, we suggested that synergistic paracrine interactions between CA and ITA ECs (HCAECs and HITAECs, respectively) may explain the increased resistance of ITA grafts and adjacent CAs to atherosclerosis and restenosis. In this study, we measured the gene and protein expression of the molecules responsible for endothelial homeostasis, pro-inflammatory response, and endothelial-to-mesenchymal transition in HCAECs co-cultured with either HITAECs or SV ECs (HSaVECs) for an ascending duration. Upon the co-culture, HCAECs and HITAECs showed augmented expression of endothelial nitric oxide synthase (eNOS) and reduced expression of endothelial-to-mesenchymal transition transcription factors Snail and Slug when compared to the HCAEC-HSaVEC model. HCAECs co-cultured with HITAECs demonstrated an upregulation of HES1, a master regulator of arterial specification, of which the expression was also exclusively induced in HSaVECs co-cultured with HCAECs, suggestive of their arterialisation. In addition, co-culture of HCAECs and HITAECs promoted the release of pro-angiogenic molecules. To conclude, co-culture of HCAECs and HITAECs results in reciprocal and beneficial paracrine interactions that might contribute to the better performance of ITA grafts upon CABG.

The total content of toxic elements in horsehair given the level of essential elements.

Elemental status of 214 mares aged 3-7 years from 11 breeds was studied: Arabian purebred (n = 20), Bashkir (n = 20), Kabarda (n = 20), Vyatka (n = 20), Tuva (n = 19), Yakutsk (n = 30), Mezenskaya (n = 20), Thoroughbred (n = 20), Akhal-Teke (n = 20), Russian trotter (n = 15), Soviet Heavy Draft (n = 10) bred in 13 regions of Russia. The research objective is to study the content of chemical elements in hair from the horse's mane, depending on the sum of toxic elements in animal hair expressed in moles. The elemental composition of the hair was defined by atomic emission and mass spectrometry (AES and MS). Elemental composition of biosubstrates was studied by 25 indicators (Al, As, B, Ca, Cd, Co, Cr, Cu, Fe, I, K, Li, Mg, Mn, Na, Ni, P, Pb, Se, Si, Sn, Hg, Sr, V, Zn). In the studies, an estimate of the total toxic load of the horse's body (∑tox) was given as the sum of mmoles of Al, Cd, Pb, Sn, Hg, and Sr in horsehair. Based on ∑tox percentile calculations, animals were divided into three groups up to 25 percentile (n = 54) with concentrations up to 1.09 mmol/kg, within the 25 and 75 percentile limits (n = 105) and over 75 percentile (n = 55) with a concentration above 6.08 mmol/kg. As follows from the obtained results, the ∑tox indicator in the mane's hair is closely connected with the total hair mineralization. For the studied range of ∑tox values, the relationship of this indicator with 13 essential and conditionally essential chemical elements is described. Moreover, as ∑tox increases, it indicates an increase in the concentration of eleven (Ca, P, Co, Cr, Fe, I, Mn, Li, Ni, V, As) and a decrease of two elements in hair (B, Si); for six elements (K, Mg, Na, Cu, Zn, Sn), such a connection was not revealed. At ∑tox values higher than 75 percentile, a critical increase in the exchange pools of two or more toxic elements in the body was observed in 85% of cases. Intensive exchange of selenium and iodine is observed; it is expressed by an increase in the number of animals with the content of these elements in hair beyond the "physiological standard," estimated as the range of 25-75 percentile.

The content of toxic elements in hair of dairy cows as an indicator of productivity and elemental status of animals.

The study was conducted on a model of dairy cows of the Holstein breed. At the first stage of research, the elemental composition of cow hair was studied (n = 198). Based on this study, the percentile intervals of chemical elements concentrations in hair were established; values of 25 and 75 percentiles were determined, and they were considered as "physiological standard." At the second stage, the elemental composition of hair from the upper part of withers of highly productive Holstein cows during the period of increasing milk yield was analyzed (n = 47). The elemental composition of biological substrates was studied according to 25 indicators, using the methods of atomic emission and mass spectrometry (AES-ICP and MS-ICP). An assessment of productivity parameters of cows depending on the level of toxic elements in hair revealed a negative statistically significant relationship with the level of lead. Lead content in hair was negatively correlated with the yield of fat (r = - 0.50), protein (r = - 0.37), and dry matter (r = - 0.48) in milk. Based on these data, cows were divided into three groups: group I, with Pb concentration in hair 0.0245-0.0449 mg/g, group II-between 0.0495 and 0.141 mg/kg, and in group III-between 0.145 and 0.247 mg/g. It was established that increasing Pb content decreases daily production of milk fat by 18.8 (P ≤ 0.05) and 25.3% (P ≤ 0.05), protein by 9.7 (P ≤ 0.05) and 10.7% (P ≤ 0.05), and dry matter by 8.0 and 13.0% (P ≤ 0.05) in cows. Average daily milk yield, adjusted for 1% of fat, decreased by 19.2 (P ≤ 0.05) and 25.3% (P ≤ 0.05), respectively. As the concentration of lead in hair increased, the content of toxic elements (Al, As, Cd, Hg, Pb, Sn, Sr) increased from 0.07 to 0.235 mmol/kg in group I, in group II from 0.082 to 0.266 mmol/kg, and in group III-from 0.126 to 0.337 mmol/kg. It was concluded that it is necessary to further study the use of physiological standard indicators of the content of toxic chemical elements in hair of dairy cows to increase productivity and maintain animal health and to create an effective system of individual health monitoring of highly productive cattle.

Assessment of Gender Effects and Reference Values of Mane Hair Trace Element Content in English Thoroughbred Horses (North Caucasus, Russia) Using ICP-DRC-MS.

The objective of the present study was assessment of gender differences in hair trace element content in English Thoroughbred horses (North Caucasus, Russia) using ICP-DRC-MS and calculation of the reference values. Trace element content in mane hair of 190 stallions and 94 mares (3-7 years old) bred in North Caucasus (Russia) was assessed using inductively coupled plasma mass spectrometry. Mane hair Co, Cr, Mn, Li, Si, and Sr levels in mares exceeded those in stallions by 77%, 63%, 64%, 42%, 39%, and 64%, respectively. Hair Fe and Si content was nearly twofold higher in female horses as compared to the males. Only hair Zn content was 5% higher in stallions as compared to mares. In addition, mares were characterized by 63%, 65%, 29%, and 40% higher levels of As, Pb, Sn, and Ni levels in hair as compared to the respective values in stallions. In turn, hair Al and Hg were more than twofold higher in mares than in stallions. The reference intervals of mane hair content (µg/g) for Co (0.006-0.143), Cr (0.028-0.551), Cu (4.17-6.84), Fe (10.11-442.2), I (0.026-3.69), Mn (0.551-12.55), Se (0.108-0.714), Zn (97.43-167), Li (0.011-0.709), Ni (0.060-0.589), Si (0.665-29.12), V (0.006-0.584), Al (1.98-168.5), As (0.006-0.127), Cd (0.002-0.033), B (0.494-16.13), Pb (0.018-0.436), Sn (0.002-0.144), Sr (1.0-9.46), and Hg (0.0018-0.017) in the total cohort of horses were estimated using the American Society for Veterinary Clinical Pathology Quality Assurance and Laboratory Standard Guidelines. The reference intervals were also estimated for stallions and mares bred in North Caucasus (Russia) and may be used for interpretation of the results of hair trace element analysis in horses.

The content of essential and toxic elements in the hair of the mane of the trotter horses depending on their speed.

A study on the Russian trotting breeds was conducted to assess the impact of horses' sporting results and the degree of accumulation of chemical elements in the hair. In the first phase of the research, the elemental composition of the mane hair of trotter horses (n = 215) was studied. Based on these studies, percentile intervals for the distribution of concentrations of chemical elements in the hair have been established, and the values of 25 and 75 percentile adopted as a "physiological standard" have been defined. In the second stage of the research into clinically healthy Russian trotting breeds (n = 56), it was estimated that the sporting results were dependent on the elemental status defined by the hair. The elemental composition of the hair was defined by 25 chemical elements using atomic emission and mass spectrometry. It is established that the mane hair is closely related to the sporting results of trotter horses. Thus, in animal wool with the highest sporting achievements, there were reliably less I, Cr, Co, Li, V, Al, Pb, and Cd, and reliably more Si than the low ones. Differences in individual elements exceeded 200%. As sporting performance diminished, the number of elements within the standard increased. For example, for mares with average speed, there were deviations from the physiological standard by 6 elements (P, Fe, Mn, I, Co, Si), with the low one by 13 elements (P, Fe, Cu, Mn, I, Co, Si, K, Cr, Ni, V, Al, Pb). A comparative estimate of the mineralization of the horses' mane measured by the sum of the amount of substances showed that there was a negative correlation between the accumulation of toxic elements and the speed (r = - 0.59). On the basis of the above, a conclusion is reached on the future use of the mane hair to assess the speed qualities of trotter horses.

The Reference Intervals of Hair Trace Element Content in Hereford Cows and Heifers (Bos taurus).

The objective of the present study was to assess hair trace element content in Hereford heifers and cows (Bos taurus) living in South Ural region and calculate the site-specific reference intervals. Hair trace element content in 150 cows and heifers farmed in the Southern Urals of Russia was assessed using inductively coupled plasma mass-spectrometry. Dietary trace element content corresponded to the adequate values as estimated by recommendations of USSR State Agriculture Committee and U.S. National Research Council. Comparative analysis demonstrated that heifers are characterized by significantly higher hair Se (3-fold), Hg (4-fold), and Sn (46%) content, whereas cows had significantly higher levels of hair Co (56%), I (33%), Si (2-fold), V (27%), B (55%), Cd (19%), Pb (47%), and Sr (23%). At the same time, no significant group difference in hair Cr, Cu, Fe, Li, Mn, Zn, As, and Ni was detected between Hereford cows and heifers. The reference intervals and 90% confidence intervals for the lower and upper limits were calculated in agreement with the American Society for Veterinary Clinical Pathology Quality Assurance and Laboratory Standard Guidelines.

Apoptosis-mediated endothelial toxicity but not direct calcification or functional changes in anti-calcification proteins defines pathogenic effects of calcium phosphate bions.

Calcium phosphate bions (CPB) are biomimetic mineralo-organic nanoparticles which represent a physiological mechanism regulating the function, transport and disposal of calcium and phosphorus in the human body. We hypothesised that CPB may be pathogenic entities and even a cause of cardiovascular calcification. Here we revealed that CPB isolated from calcified atherosclerotic plaques and artificially synthesised CPB are morphologically and chemically indistinguishable entities. Their formation is accelerated along with the increase in calcium salts-phosphates/serum concentration ratio. Experiments in vitro and in vivo showed that pathogenic effects of CPB are defined by apoptosis-mediated endothelial toxicity but not by direct tissue calcification or functional changes in anti-calcification proteins. Since the factors underlying the formation of CPB and their pathogenic mechanism closely resemble those responsible for atherosclerosis development, further research in this direction may help us to uncover triggers of this disease.

Preparing for a motor perturbation: early implication of primary motor and somatosensory cortices.

Although preparation of voluntary movement has been extensively studied, very few human neuroimaging studies have examined preparation of an intentional reaction to a motor perturbation. This latter type of preparation is fundamental for adaptive motor capabilities in everyday life because it allows a desired motor output to be maintained despite changes in external forces. Using fMRI, we studied how the sensorimotor cortical network is implicated in preparing to react to a mechanical motor perturbation. While maintaining a given wrist angle against a small force, subjects were instructed to prepare a reaction to a subsequent wrist angle displacement. This reaction consisted of, either resisting the imposed movement, or remaining passive. During the preparation of both reactions we found an early implication of M1 and S1 but no implication at all of the higher order motor area preSMA. This is clearly different from what has been found for voluntary movement preparation. These results show that the sensorimotor network activation during preparation of voluntary motor acts depends on whether one expects a motor perturbation to occur: when external forces can interfere with ongoing motor acts, the primary sensorimotor areas must be ready to react as quickly as possible to perturbations that could prevent the goal of the ongoing motor act from being achieved.