Microcirculatory Vessels Research Articles

Clinical and Pathoanatomical Characteristics of Hypoplastic Left Heart Syndrome a Clinical Case

Hypoplastic left heart syndrome (HLHS) is a common congenital heart defect, accounting for 4 to 16% of cases, and is one of the most prognostically unfavorable, involving single-­ventricle hemodynamics. HLHS involves hypoplasia of the left ventricle, often in combination with atresia and/or stenosis of the aortic and/or mitral valves, as well as hypoplasia of various parts of the aorta. Intrauterine reduction in blood flow to the left heart structures at early stages of fetal development subsequently leads to severe changes in cardiac structures, often not amenable to surgical correction. Objective. To study the clinical and morphological features of HLHS using a clinical case as an example. Materials and Methods. The patient was a child from the first pregnancy, which was complicated by anemia and first-­trimester preeclampsia. The defect was diagnosed prenatally at 23/24 weeks. At 39 weeks, labor was induced, and the infant was hospitalized in the intensive care unit with symptoms of cardiac and respiratory failure. On the 6th day of life, surgical treatment was performed — a Norwood operation, which is a single-­ventricle hemodynamic surgical correction of the defect. Despite intensive care measures, multiple organ failure progressed, leading to the development of disseminated intravascular coagulation (DIC) and subsequent death on the 10th day of life. Results. Autopsy, including staining of the myocardium with Masson’s trichrome and hematoxylin and eosin, showed pronounced plethora of the microcirculatory vessels with formation of erythrocyte thrombi. Pronounced perivascular edema and fraying of muscle fibers were detected. Fibroelastosis of the endocardium of the left ventricle was observed, characterized by pronounced sclerotic thickening due to significant proliferation of fibrous tissue, with the spread of connective tissue strands into the myocardium. Endocardial fibroelastosis is a component of HLHS and one of the determining factors in the outcome of cardiac surgery. Conclusion. The results of this work indicate the complexity of HLHS as a developmental defect, the severity of its clinical manifestations, and the limited prospects of currently existing surgical treatment methods. These patients may require heart transplantation in the first days of life. A potential alternative to early postnatal heart transplantation may be the development of methods for intrauterine correction of the existing developmental defect.

In vivo pilot study into superficial microcirculatory characteristics of colorectal adenomas using novel high-resolution magnifying endoscopy with blue laser imaging

BACKGROUND No studies have yet been conducted on changes in microcirculatory hemodynamics of colorectal adenomas in vivo under endoscopy. The microcirculation of the colorectal adenoma could be observed in vivo by a novel high-resolution magnification endoscopy with blue laser imaging (BLI), thus providing a new insight into the microcirculation of early colon tumors. AIM To observe the superficial microcirculation of colorectal adenomas using the novel magnifying colonoscope with BLI and quantitatively analyzed the changes in hemodynamic parameters. METHODS From October 2019 to January 2020, 11 patients were screened for colon adenomas with the novel high-resolution magnification endoscope with BLI. Video images were recorded and processed with Adobe Premiere, Adobe Photoshop and Image-pro Plus software. Four microcirculation parameters: Microcirculation vessel density (MVD), mean vessel width (MVW) with width standard deviation (WSD), and blood flow velocity (BFV), were calculated for adenomas and the surrounding normal mucosa. RESULTS A total of 16 adenomas were identified. Compared with the normal surrounding mucosa, the superficial vessel density in the adenomas was decreased (MVD: 0.95 ± 0.18 vs 1.17 ± 0.28 μm/μm2, P < 0.05). MVW (5.11 ± 1.19 vs 4.16 ± 0.76 μm, P < 0.05) and WSD (11.94 ± 3.44 vs 9.04 ± 3.74, P < 0.05) were both increased. BFV slowed in the adenomas (709.74 ± 213.28 vs 1256.51 ± 383.31 μm/s, P < 0.05). CONCLUSION The novel high-resolution magnification endoscope with BLI can be used for in vivo study of adenoma superficial microcirculation. Superficial vessel density was decreased, more irregular, with slower blood flow.

Effect of a water-soluble form of dihydroquercetin on age-dependent LPS-induced gliovascular remodeling of the substantia nigra in rats

Background. The molecular and cellular mechanisms of action of the water-soluble form of dihydroquercetin (DHA-VF) on neurogliovascular units, age-related disturbances of intrasystem connections that may underlie neuroinflammatory and neurodegenerative diseases, remain unclear.The aim: to study structural changes of microcirculatory vessels and functional responses of micro- and astroglial cells in the substantia nigra of young and old rats in response to intranigral injection of lipopolysaccharide (LPS) and subsequent oral administration of DHA-VF.Methods. Young (250-320 g) and old (390-450 g) Wistar rats were injected with 2 μL LPS solution at a concentration of 0.01 μL/mL (experimental group; n=24) or 2 μL sterile physiological solution (control group; n=12) into the substantia nigra area of the brain using a stereotaxic device. Half of the animals in the experimental groups (6 animals of each age group) were given 2 ml of a solution containing DHA-VF at a concentration of 3 mg/mL daily by gavage with a probe. After 8 weeks, the animals were decapitated and cryostat sections were obtained for histochemical (FITC-labeled tomato lectin) and immunohistochemical (antibodies against GFAP and CD-11β) staining of vascular endothelium and glial cells, respectively. Results. 8 weeks after LPS administration to old rats in the CS, a significant excess of areas occupied by cell bodies and processes of microglial and astroglial cells as well as the number of vessels at standard sites was found compared to both young animals exposed to similar exposure and old control animals. Oral administration of DHA-VF to rats significantly reduced LPS-induced glial activation in young and old animals. In addition, administration of DHA-VF to old animals reduced the intensity of LPS-induced microvascular remodeling of the CS. Conclusions. LPS administration in the rat CS induces neuroinflammation and vascular angiogenesis, which are maximally expressed in old animals. Administration of DHA-VF for 8 weeks significantly reduced these LPS-induced changes.

Influence of excessive salt consumption on the functional state of microvascular vessels of the skin of rats with renal dysfunction

Background. Cardiovascular complications are common in chronic kidney disease (CKD). The relationship between the cardiovascular system and the kidneys is complex and diverse, but the question of the mechanisms of the influence of excess NaCl consumption on the functional state of blood vessels during renal dysfunction remains relevant.The aim: to evaluate the effect of a high-salt diet on the functional state of microcirculatory vessels (MCR) of the skin in rats at an early stage of renal dysfunction.Material and Methods. The study was conducted on Wistar rats. The first SO group included animals subjected to sham surgery (SO) and receiving a standard diet (0.34% NaCl); in the second, HS group – animals that also underwent a sham operation, but received a high-salt diet (4% NaCl); in the third, NE group – rats subjected to ¾ nephrectomy (NE) and receiving a standard diet; the fourth, HS+ NE group included rats subjected to ¾ NE and fed a high-salt diet (4% NaCl). Blood pressure (BP) was measured in the tail using the cuff method (Sistola, Russia), MCR was assessed using laser Doppler flowmetry (LDF). The reactivity of skin microvessels was assessed by changes in the MCR index before and after iontophoresis of acetylcholine (ACh) and sodium nitroprusside (NP). Based on the assessment of the spectral characteristics of fluctuations in the MCR index obtained using Wavelet analysis, the value of endothelial, neurogenic and myogenic microvascular tone was calculated.Results. 4 months after ¾ NE, rats showed a decrease in the excretory function of the kidneys. In rats on a highsalt diet, an increase in blood urea levels was noted. The blood pressure in rats of the SO+HS group did not significantly differ from the control animals of the SO group (131 ± 8 and 125 ± 4 mmHg, respectively, NS), NE led to a significant increase in blood pressure (135 ± 5 mmHg in rats of the NE group, and 145 ± 7 mmHg – in the NE+HS group). The average MCR was higher in rats receiving a high-salt diet (SO+HS and NE+HS groups), compared with animals of the corresponding group receiving a standard diet. The combination of NE and high-salt load has an inhibitory effect on the amplitude of fluctuations in the intensity of perfusion in all studied ranges: endothelial, neurogenic and myogenic, while the calculated value of skin microvascular tone in three ranges was the highest in NE+HS group rats. In all experimental groups, the reactivity to AH was reduced compared with the SO group, and after NP iontophoresis, the average perfusion rate significantly decreased only in groups of animals receiving a high-salt diet: SO+HS and NE+HS.Conclusion. A high-salt diet in rats with an initial stage of renal dysfunction contributes to an increase in blood pressure and a decrease in natriuresis compared with animals with NE who received a standard diet. With excessive consumption of table salt in rats after NE, the intensity of cutaneous MCR increases, while reducing the variability of the MCR index. A high-salt diet in rats with NE modifies the characteristics of the spectral components of fluctuations in the MCR index, indicating an increase in tonic effects on skin microvessels in the endothelial, neurogenic and myogenic ranges. With hypernatrial load, animals with ¾ NE have a decrease in the reactivity of cutaneous microvessels to AH and NP, which is mediated by both a decrease in endothelial NO production and a decrease in MMC sensitivity to NO.

Pathomorphological changes in laboratory animals exposed to lethal doses of disinfec-tants

The use of disinfectants is a crucial aspect of preventive and health improvement measures for infectious diseases in farm and domestic animals. Regulatory documents require the determination of toxicity to macroorganisms, including the establishment of lethal doses and toxicity groups, during the development and registration of antimicrobial agents. This study aimed to investigate the histological changes in the internal organs of laboratory animals when determining lethal doses of innovative disinfectants. The experiments used domestic disinfectants containing glutaraldehyde as the active ingredient. Histological studies were conducted on the internal organs (kidney, liver, stomach, intestines, and spleen) of 75 laboratory animals using an Axioskop 40/40FL microscope (Carl Zeiss, Germany) with video microscopic photography. The methods used were under current standards. Hemodynamic disorders were observed in the renal tissue under the influence of lethal doses of aldehyde disinfectants. These disorders were characterized by capillary dilation and blood filling. Glomerular capillary dilation and overflow with blood cells were also detected. Additionally, stasis was observed in the lumen of the microcirculatory vessels throughout the entire length. The examination of histological sections from animal liver samples revealed a significant expansion of Dissé spaces, variations in the size of hepatocyte nuclei, beam decomposition and fragmentation, small acute perivascular hemorrhages, leukostasis in sinusoids, and hemodynamic disorders. The structure of the organ's beams was also disturbed, and a significant number of venous vessels were dilated and excessively filled with blood cells. Minor changes were detected in the stomach, including desquamation of the epithelial cells of the glands and their exfoliation into the gastric lumen, as well as circulatory disorders. Epithelial desquamation, blood vessel dilation, and signs of connective tissue edema were observed in the intestine. The kidneys exhibited signs of acute venous hemorrhage and stasis in vessels of various calibers, with the development of small acute parenchymal hemorrhages and localized lymphoid cell death in the white pulp. The prospect of further research is to investigate the histomorphological changes in the internal organs of laboratory animals when exposed to modern complex disinfectants with different active ingredients.

A Water-soluble Form of Dihydroquercetin Reduces LPS-induced Astrogliosis, Vascular Remodeling, and mRNA VEGF-A Levels in the Substantia Nigra of Aged Rats

Background: The age-dependent sporadic form of PD is characterized by the degeneration of dopaminergic (DA) neurons in the Substantia Nigra (SN), gliosis, and vascular changes. Vascular changes may contribute to the onset of the disease and exacerbate the neurodegenerative process, as some vascular changes occur before the onset of neuronal loss. To demonstrate the anti-neuroinflammatory efficacy of a new compound, a water-soluble form of dihydroquercetin (DHQ-WF), we studied the structural changes of microcirculatory vasculature, astroglial GFAP, and vascular endothelial growth factor –A (VEGF-A) mRNA expression in the SN of young and old rats after unilateral nigral treatment by lipopolysaccharide (LPS) and oral administration of DHQ-WF. Materials and methods: The experiments were performed on 18 young (8 weeks - 10 weeks old; 250 g - 320 g) and 18 old (18 months - 19 months old; 390 g - 450 g) male Vistar rats. Young and adult rats from the experimental groups were stereotactically injected with 2 μL LPS solution (LPS from Escherichia coli; 0,01 μL/mL) into one side of the SN. Control young and old rats were similarly injected with 2 μL sterile saline. Half of the animals in both the control and experimental groups (6 animals in each group) received a 2 ml solution containing DHQ-WF at a concentration of 3 mg/ml orally every day. After 8 weeks, brains were harvested and serial cryostat sections were prepared for histochemical (FITC-labeled tomato lectin), immunohistochemical (anti-GFAP Antibody, Cy3 Conjugate) staining, and real-time PCR (mRNA VEGF-A). Results: Eight weeks after LPS injection into the SN, a significant excess of areas occupied by cell bodies and processes of astroglial cells, the density of microcirculatory vessels, and mRNA VEGF-A expression was observed in old animals compared to control old animals and young LPS-treated rats. Oral administration of DHQ-WF to LPS-treated rats resulted in a significant reduction of these parameters in old animals. Conclusion: Injection of LPS into rat SN induces neuroinflammation and vascular angiogenesis, maximally expressed in old animals. Administration of DHQ-WF for 8 weeks significantly reduces these LPS-induced changes. DHQ-WF may be an effective treatment for reducing the effects of neuroinflammation in the aging brain.

Histology of fetal lungs at different gestational age

The lecture is devoted to the morphological characteristics of the maturation of lung tissue structures in the fetal period. Fetal histology of the lungs presents the intrauterine development of lung tissue in four successive stages: pseudoglandular, canalicular, saccular and alveolar, each has specific morphological criteria. The following morphological features are predetermined: the development of alveolar epithelium, the ratio of mesenchyme towards the area in alveolar spaces, the degree of proliferation and location of vessels of the microcirculatory bed towards prealveolar partitions. During the fetal period the alveolar columnar epithelium is flattened and differentiates into alveolocytes type I and II, the area of the mesenchyme gradually decreases and by the birth of a full-term newborn kid it is present mainly in the thickness between the alveolar septa, microcirculation vessels, initially laying deep in the thickness of the mesenchymal tissue, gradually proliferate, approach the pre-alveolar epithelium, channeling it with the formation of alveolar capillary membranes. Air exchange in the lung tissue is mainly provided with two factors: the presence of second-order alveolocytes capable of producing surfactant, and a sufficient formation of alveoli as well. This work summarizes the basics of fetal lung histology with the demonstration of histological preparations of the lungs at different stages of intrauterine development.

МИКРОЦИРКУЛЯЦИЯ (ЧАСТЬ 1 – ГЛАВНЫЕ ДЕЙСТВУЮЩИЕ «ЛИЦА» И ИХ РОЛЬ В СИСТЕМЕ КРОВООБРАЩЕНИЯ)

The article presents modern views on microcirculation, which is the terminal compartment of the cardiovascular system, built into the tissue structure and determining the target function of blood circulation – the exchange between blood and tissues of substrates, energy and information, which ensures the maintenance of life support processes at the cellular level. The paper highlights the hemodynamic features that determine the main functions of microcirculatory vessels as the most important part of the blood circulation. The structure and morphofunctional features of all levels of microcirculation including the smallest vessels of the terminal bed, which preserve the morphological structure of arteries and veins, are described in detail. The main mechanisms of regulation of arteriolar blood flow are identified: neurohumoral, myogenic, and metabolic. A detailed description of the endothelium is given. The role of the barrier function of the endothelium in maintaining the balance between osmotic and hydrostatic pressure and in the formation of the lymphatic system for drainage of excess interstitial fluid has been noted. The results of studies on microvascular endothelial cells, in which genes encoding basement membrane proteins are especially active, are presented. Attention is focused on the morphological and functional properties of the endothelial cell glycocalyx as a new clinical paradigm. The glycocalyx is present in all capillaries, arteries and veins and is a structure of distant interaction of endothelial cells with their environment, changing under pathological conditions (adhesion, thrombus formation, apoptosis).

Патоморфологічні та морфометричні особливості гострого апендициту в дітей з цукровим діабетом І типу

Metabolic disorders caused by chronic hyperglycemia in the gastrointestinal tract of patients with type I diabetes mellitus (T1DM) not only affect natural course of this disease but also modify clinical course of concomitant abdominal purulent-inflammatory diseases in children due to changes of the internal intestinal wall. Purpose - to analyze pathomorphological and morphometric features of acute appendicitis in children with T1DM. Materials and methods. We carried out a pathomorphological and morphometric case-control study of the surgical specimens (appendices and fragments of the peritoneum). Two groups were created: the Group I (n=11) - patients with acute appendicitis, peritonitis and T1DM; the Group II (n=24) - patients with acute appendicitis, peritonitis but without T1DM. Statistical analysis was performed using software package STATISTICA v.10.0 (StatSoft, USA). Results. There were signs of diabetic angiopathy in the Group I and a significant number of inflammatory cells, represented by a large number of segmented nuclear leukocytes - 431±18.2 in 1 mm2, plasma cells - 146±11.13 in 1 mm2, lymphohistiocytic elements - 196±23,32 in 1 mm2. The density of the inflammatory cell infiltrate was 773±36.2 cells in 1 mm2. Morphometric features of the Group II are as follows, number of segmented nuclear leukocytes - 228±15,7 in 1 mm2, plasma cells - 112±10,41 in 1 mm2, lymphohistiocytic elements - 132±21,2 in 1 mm2. The density of the inflammatory cell infiltrate was 773±36.2 cells in 1 mm2. Conclusions. There is an increase in the density of the inflammatory cell infiltrate, the number of neutrophils, plasma cells, and lymphohistiocytic elements in samples of appendices and peritoneum in children with acute appendicitis and T1DM in comparison with the Group II (p˂0.001). Furthermore, those specimens feature presence of hyalinosis, thickening of the vascular wall and partial obliteration of small caliber vessels. In the Group II (patients without T1DM), the parietal and visceral peritoneum had a large number of inflammatory elements and presence of macrophage elements upon normal histological structure of the microcirculatory vessels. The research was carried out in accordance with the principles of the Helsinki Declaration. The study protocol was approved by the Local Ethics Committee of participating institution. The informed consent of the patient was obtained for conducting the studies. No conflict of interest was declared by the authors.

COVID-19 in Children: Molecular Profile and Pathological Features.

Although the World Health Organization has declared the end of the COVID-19 pandemic, doctors continue to register new cases of the disease among both adults and children. Unfortunately, the course of COVID-19 in children can have a severe form, with death being a potential outcome. The absence of published works discussing the pathological morphology of COVID-19 in children prevents the objective analysis of the disease's pathogenesis, including among the adult population. In this vein, the objective of our study is to identify the morphological features of the lungs' involvement and evaluate virus-host interactions in the case of COVID-19 in patients at a pediatric medical practice. We present the results of the study of the lungs of three children who died due to COVID-19, highlighting the predominant involvement of their respiratory organs at different stages of the disease (5, 21, and 50 days). This article presents data obtained from histopathological and immunohistochemical investigations, taking into account the results of clinical and laboratory indicators and intravital and postmortem SARS-CoV-2 PCR investigations. The common finding of all of the examined COVID-19 cases is the involvement of the endothelium in microcirculation vessels, which are considered to be a primary target of various pathogenic influencing factors. We also discuss both the significance of apoptosis as a result of virus-host interactions and the most likely cause of endothelium cell destruction. The results of this study could be useful for the development of endothelium-protective therapy to prevent the progression of disseminated intravascular coagulation syndrome.

STAGES OF MORPHOLOGICAL CHANGES OF TARGET ORGANS IN EXPERIMENTALLY SIMULATED ACUTE PERITONITIS

One of the urgent problems in urgent abdominal surgery remains acute disseminated peritonitis (AP), the mortality rate of which remains high due to the development of multiple organ failure syndrome, despite the use of modern methods of diagnosis and treatment.The purpose of the study. To substantiate the possibility of using laparoscopic methods of treatment depending on the severity and stage of morphological changes of the target organs and the duration of peritonitis simulated in the experiment.Material and methods. The experiment used 150 sexually mature non-linear female rats, weighing 180.0-220.0 grams, which were kept on the standard ration of the vivarium of the Central Research Laboratory with maintenance of food and drinking regimes recommended by the norms of keeping laboratory animals. The animals were divided into 5 groups – a control group (30 rats) and 4 experimental groups (120 rats), which underwent HP simulation by intraperitoneal injection of 1 ml of 30 % autocal. The animals of the research group were removed from the experiment under sodium thiopental anesthesia by bloodletting on the 1st, 3rd, 5th, and 7th days. Results of the study. Morphohistochemical study of the target organs on the 1st day of the experiment: singleorgan changes were observed (examination of the peritoneum revealed its pronounced swelling). A slight vascular reaction of the basal membrane of the mucous membrane was established in the small intestine. On the 3rd day, the changes remained monooraneous with increases in infl ammatory phenomena. On the 5th day, the spread of the systemic infl ammatory response to the lungs and kidneys was confi rmed. Areas with foci of dystrophic- necrotic changes in the epithelium of the tubules were visualized in individual excretory tubules in the kidneys.In the lungs: the number of erythrocytes and fi brin increased in the cellular component. On the 7th day of the experiment, the heart and liver were involved in the process. In the heart, dystonia and paresis of microcirculatory vessels, erythrocyte stasis, and the formation of cell aggregates in the lumen of vessels were observed. In the liver, it is observed: congestion of the central veins, manifestations of venous stasis.Conclusion. Based on the stages of morphological changes of the studied organs at diff erent times, the simulation of acute peritonitis allowed us to theoretically justify a diff erentiated approach to the selection of the type of surgical interventions.

Morphologic characteristic of rat myocardium in comorbid pathology

Diabetic cardiomyopathy is a serious complication of diabetes mellitus (DM). Aim. Therefore, we aimed to study myocardial changes in adult rats with streptozotocin (STZ)-induced DM exposed to chronic immobilization stress (CIS). Materials and methods. A total of 26 adult albino male rats weighing 180–200 g were examined. All the animals were divided into three groups: Group I included 10 rats with STZ-induced DM exposed to CIS; Group II comprised 10 rats with STZ-induced DM; Group III included 6 intact animals. The samples were collected on the 14th and 56th days of the experiment. Histological, histochemical, electron microscopy, and biochemical methods were used. Results. On the 14th day of the experiment, in Group I and Group II, increased blood flow was observed in the capillaries, venules, and veins, while an arteriolar spasm in the microcirculation was found. In addition, cardiomyocyte surface area in different myocardial regions reduced due to low glycogen content as confirmed by histochemical and ultrastructural studies. On the 56th day of the experiment, in Group I and Group II, hyperemia occurred due to red blood cell aggregation and microthrombi. The surface area of all microcirculatory vessels increased as compared to that of intact animals, as evidenced by an increase in their wall surface area leading to an increase in their wall-to-lumen ratio. Such morphometric changes in the microcirculatory vessels were indicative of decreased vascular permeability and impaired myocardial blood flow. At the histological level, in Group I and Group II, focal cardiomyocyte lysis, moderate to diffuse stromal edema, lymphohistiocytic infiltration were seen. Such changes pointed to sterile inflammation, probably due to myocardial infraction secondary to diabetic microangiopathy. In cardiomyocytes, karyolysis, vacuolar degeneration, apical ballooning, subsarcolemmic edema, fibrosis and lysis of myofibrils, colliquative necrosis were observed. Conclusions. STZ-induced DM and stress resulted in pronounced destructive changes in the myocardium of rats, including interstitial edema, focal cardiosclerosis, myolysis. Such changes occurred on the background developing diabetic microangiopathy. The most pronounced myocardial changes were recorded in animals with a comorbidity.

Malondialdehyde Serum Levels in Patients with Systemic Sclerosis Relate to Dyslipidemia and Low Ventricular Ejection Fraction.

Systemic sclerosis (SSc) is a chronic disease characterized by vasculopathy with the involvement of dysfunctional microcirculatory vessels. Features of the disease include progressive fibrosis of the skin and internal organs and systemic inflammation characterized by the presence of circulating autoantibodies and proinflammatory cytokines. Furthermore, macrovascular disease and atherosclerosis are more common in patients with SSc than in the general population. Oxidative stress plays a crucial role in the development of several processes, including endothelial dysfunction, cancer, inflammation, and atherogenesis. Malondialdehyde (MDA) is a well-established marker of oxidative stress. In this work, we have analyzed the relationship between serum MDA levels and clinical, laboratory, and vascular characteristics in a well-characterized cohort of 53 patients with SSc. A multivariable analysis was performed to study the relationship between circulating MDA and disease characteristics in patients with SSc. Cardiovascular assessment was also performed, including ultrasonography of the carotid and aorta, and echocardiography. MDA showed a significant and positive relationship with the serum levels of lipid profile molecules such as total cholesterol (β coefficient = 0.006 (95% CI: 0.0004 to 0.01), nmol/mL, p = 0.037) and LDL cholesterol (β coefficient = 0.008 (95% CI: 0.001 to 0.01) nmol/mL, p = 0.017). On the contrary, most manifestations of the disease, including skin, lung, and joint involvement, as well as the presence of digital ulcers, were not related to MDA. However, high MDA levels were significantly and independently associated with lower ventricular ejection fraction after adjustment for covariates (β coefficient = -0.04 (95% CI: -0.06 to -0.02), nmol/mL, p = 0.001). In conclusion, serum MDA levels were related to higher levels of total and LDL cholesterol and a lower left ventricular ejection fraction in patients with SSc. MDA could serve as a potential biomarker of dyslipidemia and heart failure in SSc.

THE COMPLEX EFFECT OF VITAMIN D AND ALPHA TOCOPHEROL ACETATE ON THE LUNG MICROCIRCULATION IN THE EXPERIMENTAL INDUCTION OF SYSTEMIC SCLEROSIS

Abstract. The aim of the study was to investigate the effects of vitamin D3 (vitamin D) and alpha-tocopherol acetate (vitamin E) on the microcirculatory vessels of the lungs in an experimental model of systemic sclerosis (SSc).
 Methods. We divided the experimental animals into four separate groups: an intact group (IG) consisting of 15 animals, a control group (CG) of 20 animals, experimental group 1 (EG1) and experimental group 2 (EG2) consisting of of 25 animals each. Randomization of groups was carefully performed using the weight variance minimization method. To induce SSc in laboratory animals of EG1, subcutaneous injections of 0.5 ml of 5% sodium hypochlorite (NaClO) were administered 3 times a week for 6 weeks in a row. The CG of rats received an isotonic solution following the same scheme, while the IG was not subjected to any injections. In addition to NaClO, the EG2 received intramuscular injections of vit E (10 mg/100 g) and vit D (1000 IU/100 g) for three weeks during the second half of the experiment.
 After 8 weeks, the animals were euthanized, and lung tissue samples were obtained for histological and electron microscopic analysis. The histological preparations were examined using light microscopy and photographed using a Leica DME light microscope, a DCM 900 digital microscope camera, and a Nicon Coolpix P5100 camera. Meanwhile, the electron microscopy was performed to visualize the ultrastructural characteristics of the lung tissue by use of "PEM-125K" (Selmi, Ukraine) with subsequent photography at magnifications from 2000 to 20000 times.
 All animal procedures were carried out in strict compliance with bioethical principles. The Ethics Commission of the IFNMU granted approval for the study under document number 117/20 on November 19, 2020.
 Results. The histological analysis of the lung specimens of CG animals showed a normal morphological structure. In the EG1, the lungs exhibited abnormalities in the structure of the hemocapillaries, including narrowing of the vessel lumen due to thickening of the vascular wall, edema, and infiltration of polymorphonuclear cells. However, in the EG2, there was an improvement in the histological features, with a reduction in edematous processes and a decrease in the intensity of polymorphonuclear cell infiltration.
 The electron microscopic examination of the lung samples from EG1 animals revealed the presence of platelet adhesion and aggregation, as well as aggregates of erythrocytes within the hemocapillary lumen. Swelling of endothelial cells and narrowing of the hemocapillary lumen were also observed, along with uneven thickening of the basement membrane. In contrast, the combination of vitamins D and E in EG2 animals resulted in improved rheological properties of blood and ultrastructural characteristics of the hemomicrocirculatory elements. The lumen of the hemocapillaries contained single erythrocytes and inactive platelets. The endothelial cells’ membraines showed minimal alterations, with clear contours and no micropinocytotic vesicles or cytoplasmic growths. The nuclei of the endothelial cells had a fine-grained matrix with evenly distributed chromatin granules.
 Conclusions. This study proves that the combined use of vitamins D and E has a positive effect on the state of the microcirculatory vessels in the lungs of experimentally induced systemic sclerosis.

Роль клеток стромы в ангиогенезе кожных ран: обзор литературы

INTRODUCTION: The study of skin wound regeneration has not lost its relevance, especially among surgeons. Angiogenesis is a key factor in wound healing. It is known that the optimal blood supply provides the tissue with nutrients, growth factors, immune cells necessary for the complete repair of skin defect. Insufficient ability to restore functional blood supply to the wound area can lead to chronic non-healing wounds. Therefore, the identification and validation of various factors influencing angiogenesis in the wound bed is of great interest for the development of potential therapeutic agents. The aim of this review is to highlight the current understanding of the mechanisms and morphological manifestations of skin wound angiogenesis, as well as the functional significance of stromal cells involved in this process. The contribution of macrophages, pericytes, mast cells, telocytes to the mechanisms of formation of microcirculatory vessels is shown. The search for literary sources was carried out in the electronic systems eLIBRARY.RU and PubMed by the main keywords of the topic: histology, skin wound regeneration, angiogenesis, stromal cells, macrophages, mast cells, pericytes, telocytes. The publications were analyzed over the last 12 years. The criteria for including a research in the review were the simultaneous mention of several keywords in the article. CONCLUSION: Analysis of the literature demonstrates the relevance of the study of angiogenesis. Modern morphological methods permitted to describe complex intercellular interactions providing the growth of the vascular network. Despite the large number of studies in this area, morphological data are constantly being supplemented to expand the potentials of histological and immunohistochemical studies. Cellular interactions during wound healing are being actively studied and confirmed to be used in the study of the mechanisms of action of drugs and in the development of new methods for treating wounds of various origin. This is of great importance, for example, in search for new reconstructive technologies, for creation of blood vessel grafts, and for development of endothelial prostheses based on non-degradable synthetic materials.

#3482 IN VIVO EVALUATIONS OF THE PROTECTIVE EFFECTS OF NEW FORMULATIONS FOR PERITONEAL DIALYSIS ADOPTING A POWERFUL MULTI-PHOTON MICROSCOPY BASED APPROACH

Abstract Background and Aims Peritoneal dialysis (PD) is a renal replacement therapy that enables metabolic waste products and excess fluids to be eliminated through the peritoneal membrane. Exposure to conventional dialysates at high glucose concentrations is considered critical for the pathogenesis of peritoneal fibrosis, angiogenesis, and epithelial-mesenchymal transition (EMT). These events largely contribute to peritoneal membrane aging, resulting in ultrafiltration and clearance failure. Pre-clinical research in this field suffers from the lack of valid and reproducible in vivo systems and so far has been limited to in vitro systems of mesothelial or endothelial cell lines. In vitro studies on mesothelial cell monolayers have validated the significant protective effects of dialysates with alternative osmo-metabolic agents. With their better biocompatibility these novel formulations could provide a valid substitute for conventional solutions (1,2). The method proposed, based on multi-photon microscopy, aims to study the physiology of the peritoneal membrane during dialysis exchange and to validate the effects of biocompatible dialysates in animal models of fibrosis as observed in the course of dialysis treatment. Method We have implemented a surgical procedure to optimize the stability of a flap of parietal peritoneum to allow direct microscope observation. The innovative technology of multi-photon microscopy enables one to observe the vessels of microcirculation in vivo and provides a 3D rendering of the peritoneal membrane, giving an overview of all the single layers without the use of specific markers (Figure 1). One may also assess specific phenomena induced by the fibrotic process, such as the thickening of the sub-mesothelial interstitium, exploiting the autofluorescence signal from excited collagen fibers caused by the physical phenomenon of Second Harmonic generation. In vivo microscopy evaluation of peritoneal membrane senescence parameters was conducted in rats subjected to one daily intraperitoneal injection (10 mL/day) for 15 days with one of the following PD solutions: a commercially available glucose-based neutral-pH, low-GDP, PD solution (Physioneal 3.86%, Baxter Healthcare, Italy), and a new single-chamber low glucose PD solution containing osmo-metabolic agents such as L-carnitine and xylitol (XyloCore HS, Galenica Senese, Italy). XyloCore HS is a lactate-buffered PD solution with glucose (1.5%), L-carnitine (0.02%) and xylitol (2.0%). The osmotic strength of the two PD solutions tested was comparable. Results Treatment with XyloCore HS was associated with a significant reduction in thickness of the sub-mesothelial interstitium (p = 0.013), the density of collagen fibers (p = 0.012) and the vascular composition (p = 0.006), as well as the number of branch points (p = 0.0335), when compared to rats treated with a commercial glucose-based PD solution, Physioneal. Figure 2 shows the different thicknesses of sub-mesothelial stroma upon treatment with the PD solutions under investigation. Conclusion Previous in vitro studies have shown that XyloCore (1,2), a novel glucose-sparing PD solution currently in Phase III clinical development (ELIXIR - NCT03994471), is able to counteract the glucotoxic effects on the peritoneal membrane induced by conventional dialysates. Our pre-clinical in vivo methodology not only confirms our previous in vitro findings, but also suggests that long-term protective effects may be achieved with XyloCore. Further studies are in progress in a diabetic animal model to extend the favorable peritoneal in vivo findings of XyloCore treatment to a systemic level, possibly by improving glycemic control.

MICROSTRUCTURAL ORGANIZATION OF THE SUBMANDULAR GLAND UNDER CONDITIONS OF 5-WEEK OPIOID INFLUENCE

The use of opioids is an effective method of treatment for severe and acute pain; however, when they used inappropriately, in addition to their powerful pain-relieving properties, opioids cause significant side effects. The initial changes in the human body caused by the use of narcotic substances (opiates) are detected in the oral cavity, in particular, in the salivary glands. The earliest manifestations of pathological changes in the oral cavity are caries, stomatitis, gingivitis, periodontitis and xerostomia. Based on the results of our research and the study of the effect of opioids on other organs, we can conclude that long-term use of opioids has a significant negative effect on the structure of the submandibular gland and homeostasis of the oral cavity as a whole. The presence of a number of unsolved questions regarding the problems of structural reconstruction of the salivary glands during the use of narcotic agents, as well as the possibility of selecting the most effective methods of treatment and prevention of salivary gland diseases determined the choice of this research topic.
 Materials and methods. The research was performed on 15 mature, white, male rats with body weight 200–220 g, aged 4.5–6 months, which were kept under standard conditions at the vivarium of the Danylo Halytsky Lviv National Medical University. Animals were divided into two groups: experimental and control. In the experimental group, animals (10 rats) were daily administered an opioid analgesic (nalbuphine) intramuscularly according to the following schedule: 1st week – 8 mg/kg, 2nd week – 15 mg/kg, 3rd week – 20 mg/kg, 4th week – 25 mg/kg, V week – 30 mg/kg. Controls were 5 rats that were injected with 0.9% sodium chloride solution in a volume of 1 ml for 5 weeks. We used the histological method to conduct the research.
 Results. Damage to the organs of the oral cavity and the submandibular gland, in particular, is an element of comorbid pathology caused by long-term use of opioids. During the 5-week administration of the opioid, we observed irreversible destructive changes in the structural organization of the submandibular gland, which was characterized by the dilation of the vessels of the microcirculation, dystrophic and necrotic changes of the final serous and mucous secretory acini, necrosis of serocytes. Such pathological processes in the structural organization of the submandibular gland can cause an imbalance in the quantitative and qualitative composition of saliva and, as a result, a violation of the homeostasis of the oral cavity and the upper parts of the gastrointestinal tract.
 Conclusions. The results of the research demonstrate the negative impact of long-term opioid exposure on the structural organization of the submandibular gland and are similar to changes in other organs and body systems during long-term opioid exposure. The damage to microcirculation vessels is the starting and primary mechanisms in this process with the subsequent increase of irreversible destructive changes of the submandibular gland. The data of the scientific work have a wide practical and theoretical significance for morphologists, pathomorphologists and dentists in the aspect of developing new methods of diagnosis, prevention and treatment of salivary gland pathology in patients with opiomania and make it possible to establish safe terms of use of «nalbuphine».

Harnessing the synergy of perfusable muscle flap matrix and adipose-derived stem cells for prevascularization and macrophage polarization to reconstruct volumetric muscle loss.

Muscle flaps must have a strong vascular network to support a large tissue volume and ensure successful engraftment. We developed porcine stomach musculofascial flap matrix (PDSF) comprising extracellular matrix (ECM) and intact vasculature. PDSF had a dominant vascular pedicle, microcirculatory vessels, a nerve network, well-retained 3-dimensional (3D) nanofibrous ECM structures, and no allo- or xenoantigenicity. In-depth proteomic analysis demonstrated that PDSF was composed of core matrisome proteins (e.g., collagens, glycoproteins, proteoglycans, and ECM regulators) that, as shown by Gene Ontology term enrichment analysis, are functionally related to musculofascial biological processes. Moreover, PDSF-human adipose-derived stem cell (hASC) synergy not only induced monocytes towards IL-10-producing M2 macrophage polarization through the enhancement of hASCs' paracrine effect but also promoted the proliferation and interconnection of both human skeletal muscle myoblasts (HSMMs) and human umbilical vein endothelial cells (HUVECs) in static triculture conditions. Furthermore, PDSF was successfully prevascularized through a dynamic perfusion coculture of hASCs and HUVECs, which integrated with PDSF and induced the maturation of vascular networks in vitro. In a xenotransplantation model, PDSF demonstrated myoconductive and immunomodulatory properties associated with the predominance of M2 macrophages and regulatory T cells. In a volumetric muscle loss (VML) model, prevascularized PDSF augmented neovascularization and constructive remodeling, which was characterized by the predominant infiltration of M2 macrophages and significant musculofascial tissue formation. These results indicate that hASCs' integration with PDSF enhances the cells' dual function in immunomodulation and angiogenesis. Owing in part to this PDSF-hASC synergy, our platform shows promise for vascularized muscle flap engineering for VML reconstruction.

Morphometric assessment of the effects of monosodium glutamate on the carotid sinus wall: an experimental study

The increase in global burden of stroke is hard to overestimate. Every year it continues to be a leading cause of mortality worldwide. Extracranial pathology of the carotid arteries is a major underlying reason of stroke. Given the role of alimentary factors in the development of atherosclerosis of the carotid arteries, possible influence of food additives on the carotid sinus structure is of special interest. Monosodium glutamate is one of the most common food additives that is allowed for consumption in many countries due to it being considered relatively safe. Recent scientific research however points towards the possible adverse effects of monosodium glutamate on the living organism. The aim of this study was to analyze qualitative and quantitative parameters of carotid sinus structural organization of white male albino rats under normal conditions, in the setting of oral monosodium glutamate consumption and after its withdrawal. Carotid sinuses of 30 white male albino rats that had been daily consuming 10 mg/kg of monosodium glutamate for 4 or 8 weeks with subsequent withdrawal for 2 weeks were subjected to qualitative and quantitative analysis at different time-points of the experiment. The data obtained was compared with the results of morphologic and morphometric study of carotid sinuses of 9 rats of the control group. For morphometric analysis, the following parameters were considered: intima thickness, media thickness, adventitia thickness, diameters of the arterioles, venules and capillaries of the carotid sinus vasa vasorum. Summarizing the morphometric analysis results, it is worth noting that, for the duration of 8 weeks of the experiment, steady increase in the thickness of all layers of the vascular wall, but especially intima and adventitia, was observed in the study group. At the same time, the diameter of the arterioles located in the carotid sinus adventitia was decreasing, while venules and capillaries demonstrated invariable increase of their lumen diameter. After 2 weeks of monosodium glutamate withdrawal, the thickness of intima and adventitia continued to increase, while media thickness had somewhat decreased, probably due to fibrosis and dystrophy. The tendency towards narrowing of the arterioles’ lumen and widening of the venules and capillaries persisted for at least 2 weeks despite discontinuation of monosodium glutamate. Therefore, systematic consumption of monosodium glutamate may lead to impairment of carotid sinus structural organization, particularly endothelial proliferation, fibrotic and dystrophic changes of media, adventitia thickening as well as microcirculatory vessels damage, that continue to persist despite the food additive withdrawal.

Invasive Detection of Coronary Microvascular Dysfunction: How It Began, and Where We Are Now.

The landscape of interventional cardiology is ever evolving. Contemporary practice has shifted from a stenosis-centred approach to the total characterisation of both the epicardial and microcirculatory vessels. Microcirculatory dysfunction plays an important role in the pathophysiology of acute and chronic coronary syndromes, and characterisation of the microcirculation has important clinical consequences. Accordingly, the invasive diagnosis of microcirculatory dysfunction is becoming a key feature of the interventional cardiologist's toolkit. This review focuses on the methodology underpinning the invasive diagnosis of microvascular dysfunction and highlights the indices that have arisen from these methodologies.