Van Gieson Research Articles

Serpina3c deficiency aggravates vascular dysfunction and VSMC dysregulation through regulating adamts4 pathway activity in obese mice

Abstract Background Obesity leads to arterial stiffness (AS), which is an independent risk factor for cardiovascular disease. Vascular smooth muscle cell (VSMC) dysfunction is a critical step in the development of AS. Adamts4 (a disintegrin and metalloproteinase with thrombospondin motifs 4) promoting the ECM remodeling can regulate VSMC differentiation. Serpina3c is a serine protease inhibitor that plays a key role in VSMC proliferation and metabolic diseases. Purpose The present study aimed to investigate the function of Serpina3c in high-fat diet (HFD)-induced AS and regulation of VSMC phenotypic switching and possible mechanisms. Methods 8-week-old male Serpina3c-/- and C57BL/6 mice were fed normal diet (ND) or HFD for 12 weeks. The adipocyte-specific Serpina3c overexpression (AAV8-Serpina3c) were constructed and injected in situ into visceral or perivascular adipose tissue (AT) of Serpina3c-/- mice fed a 12-week HFD to test in vivo effect of AT-derived Serpina3c on arterial stiffness. In vivo vascular stiffness was analyzed by obtaining pulse wave velocity (PWV) measurements. Aortic rings were dissected to assess the ex-vivo effects of Serpina3c deprivation on vascular contractile and diastolic function. Elastin van Gieson (EVG) and Hematoxylin-eosin (H&E) staining were implemented to observe the pathological morphology of aortas. IP-Western analysis was performed for identifying Serpina3c-adamts4 interactions. Results The obese WT mice exhibited significantly increased PWV values, compared with ND-fed mice, which were further increased in HFD+Serpina3c-/- mice (Fig. 1A). Phenylephrine-induced vascular constriction was impaired in HFD fed mice and further deteriorated with Serpina3c deficiency (Fig. 1B). No differences were noted in the endothelial-dependent acetylcholine-mediated relaxation in HFD-fed mice. Notably, Serpina3c-/- mice exhibited poorer endothelium-independent sodium nitroprusside induced relaxation (Fig. 1C and D). The HFD+Serpina3c-/- mice showed an increased breakage of elastin fibres and medial thickness compared with the aortas of HFD+WT mice (Fig. 1E and F). Serpina3c deprivation aggravated collagen deposition and matrix metalloproteinases (MMPs, MMP2 and MMP9) accumulation. VSMC contractile gene α smooth muscle actin were upregulated and synthetic phenotype marker osteopontin were downregulated in HFD+WT mice. The effect was more pronounced in aortas with Serpina3c deficiency (Fig. 1G and H). Overexpression of serpina3c in both visceral and perivascular AT or adamts4 deprivation reversed the above phenotypes. Computer simulations and IP-Western results indicate that Serpina3c and adamts4 bind to and maintain the latter low-expression in response to obesity (Fig. 2). Conclusion Serpina3c interacts with adamts4 to maintain contractile phenotype of VSMC and suppress AS development in the context of nutrient excess. Serpina3c is a novel target for the prevention and treatment of obesity associated artery disease.Serpina3c maintains vascular functionSerpina3c interacts with adamts4

The clinical significance of elastic lamina invasion in patients with pStage II colorectal cancer: a notable prognostic indicator

BackgroundSome colorectal cancers (CRCs) are clinically diagnosed as cT4a with serosal invasion (SI). However, the cT4a is most often underdiagnosed pathologically as pT3 without SI by hematoxylin–eosin (H&E) staining alone. Using Elastica van Gieson (EVG) staining, some pT3 tumors invade the elastic lamina (EL), which extends just below the serosal layer. Recently, EL invasion (ELI) has been described as a poor prognostic factor for disease-free survival (DFS) and overall survival (OS) in patients with pStage II CRC. However, its clinicopathological significance remains unclear due to the limited number of studies and poor understanding of ELI.ObjectiveThis study investigated the association between the ELI and patient prognosis.MethodsAfter 1982, pathological diagnosis was routinely performed using H&E and EVG staining methods, and long-term follow up was performed until 2016. All clinicopathological features including ELI were prospectively registered into our computer and 569 patients with pStage II CRC were collected from the database. Based on the ELI status, pT3 was divided into three pathological categories: pT3ELI − was defined as pT3a, pT3ELI + as pT3b and unidentified EL (pT3EL −) as pT3u.ResultsUsing H&E staining alone, gross cT4a was most often pathologically underdiagnosed as pT3 (93.8%) and very rarely as pT4a, resulting in a large diagnostic discrepancy. Using EVG staining, 60.7% of the cT4a tumors were diagnosed as pT3b. The 10-year DFS and OS rates were similar for pT3a and pT3u patients. However, the 10-year DFS and OS rates of pT3b patients were significantly lower than those of pT3a patients (75.6% vs. 95.6%, p < 0.0001 and 58.4% vs. 70.6%, p = 0.0024, respectively) but did not differ from those of pT4a patients (70.6%, p = 0.5799 and 52.0%, p = 0.1116, respectively). Multivariate analysis revealed that the ELI was the strongest independent risk factor for recurrence and CRC-specific death (p < 0.0001).ConclusionsA better understanding of the ELI allows us to reconsider the diagnostic discrepancy of serosal invasion, i.e., pT3b should be considered pT4a. The ELI-based subclassification of pT3 is expected to be incorporated into the TNM staging system in the future. The ELI is a notable prognostic indicator in patients with pStage II CRC.

Microscopic Anatomy of Orbital Septum.

To study the microscopic anatomy of the orbital septum and evaluate its relationships with the adjoined structures. Histological evaluation of 11 upper eyelids (5 right, 6 left) from 10 Japanese cadavers (age range: 36-94-year-old, average: 71.1) was performed. The specimens were fixed in 10% formalin and stained with Elastica Van Gieson. Specimens were microscopically analyzed using different magnifications (20×, 40×, and 100×). The orbital septum was the single fibrous layer in all the specimens, anteriorly separated from the retro-orbicularis fat pad capsule and posteriorly from the preaponeurotic fat pad capsule. At the junction of the orbital septum with the levator aponeurosis, the orbital septum proceeded towards the tarsal plate, and the preaponeurotic fat pad capsule reflected posteriorly. The orbital septum is constituted of a single layer of fibrous tissue anteriorly separated from the retro-orbicularis fat pad capsule and posteriorly from the preaponeurotic fat pad capsule by the corresponding virtual spaces.

Histological and morphometrical evaluation of the urethral wall after bioresorbable stent implantation in male New Zealand White Rabbits: A preliminary study.

The aim of the study was the histological and morphometrical evaluation of the urethral wall at three time points after bioresorbable stent implantation in male New Zealand White Rabbits. The research was performed on 26 male New Zealand White rabbits aged 3-4 months and weighing 2.1-3.0 kg. Two models of bioresorbable sodium alginate-based stents were developed and implanted into the urethral lumen for one (T1), three (T3), and six weeks (T6). Sections of 5 µm thickness were cut from the urethra at intervals of 2 mm. The sliced sections were stained with hematoxylin-eosin (H&E), Van Gieson's (VG), Von Kossa, and Movat-Russell modified pentachrome (MOVAT) staining methods. The study provided valuable information for future models of urethral stents. The first model of the stent failed to fit the requirements due to inadequate mechanical properties. It curled up on itself losing the ability to adhere to the animals' urethra and was bioresorbed three weeks after implantation. The more rigid no. 2 stent was effective in widening the urethral lumen but did not biodegrade during the experiment. A comprehensive assessment of the second model's properties of biosorption and biointegration requires an extended observation of at least 12 months for an in depth morphological analysis. Stent migration is not likely to be caused solely by the mechanical properties of the urethra or urinary flow but mainly by muscle contraction of the organ wall.

Organization, density, and content of collagen in the body wall of sea cucumbers Acaudina rosettis and Phyllophorus sp.

Marine collagen has been known to have lower immunogenicity compared to bovine collagen. The primary component in the body wall of sea cucumbers is collagen. This study aimed to investigate the density, bundle organization, and yield of collagen in the two sea cucumber species, Acaudina rosettis and Phyllophorus sp. collected from the Madura Strait, Indonesia. The body wall parts, including the dorsal anterior, dorsal median, dorsal posterior, ventral anterior, ventral median, and ventral posterior, from five samples of each species, were fixed and decalcified. They were histologically processed using the paraffin method before staining them with Van Gieson to visualize collagen in the cross-sections of body wall tissue. Collagen density was determined using the ImageJ software, while collagen bundle organization was examined negatively by utilizing the image editing application. Collagen density data were analyzed using repeated-measures ANOVA (α = 0.05). Collagen content in the body wall of each species was examined using acid solubilization-based collagen extraction, which did not significantly differ between different parts of the body wall in each species (p = 0.161 for A. rosettis, p = 0.095 for Phyllophorus sp.). However, Phyllophorus sp. exhibited a higher collagen density and collagen bundles, which were larger and more organized compared to A. rosettis. The collagen content was higher in A. rosettis (10.07%) compared to Phyllophorus sp. (8.07%). Further study can be performed to explore the potential of collagen from these sea cucumber species for commercial use.

Histological characterization of rat vocal fold across different postnatal periods.

To evaluate the vocal fold histological characteristics during different postnatal periods in rats, especially older rats. Sprague-Dawley rats aged 4 days, 4 and 12 weeks, and 12 and 24 months were used for the experiment. Five larynges were obtained for each age and cut into 5-μm consecutive sections. The expression of Ki-67 was assessed using immunohistochemistry to examine cell proliferation. Elastic van Gieson staining was used to detect the collagen and elastin concentrations. The cell type was determined using multicolor immunofluorescence. Ki-67 was not expressed in the macula flava (MF) of 12-week-, 12-month-, and 24-month-old adults. Collagen fibers in the lamina propria (LP) increased with age. The elastic fiber concentrations in the LP decreased significantly at 24 months (p < .01) but remained stable in the MF. All posterior MF cells showed strong glial fibrillary acidic protein and vimentin-positive reactions with weaker expressions of CD68 and α-smooth muscle actin (α-SMA). The myofibroblasts (α-SMA-positive) and macrophages (CD68-positive) in the LP of the 24-month-old rats were significantly the highest (p < .01). The extracellular matrix in the LP increases with age, presenting as an increase in collagen with the loss of elastin, which may be due to myofibroblast proliferation. Moreover, the cellular properties or extracellular matrix components of the mature MF in rats are comparable to those in humans.

Expression of Wnt signaling proteins in rare congenital bladder disorders

Introduction and AimsCongenital bladder anomalies are rare and are a leading cause of end stage renal failure in children. The Wnt signaling pathway, important during embryonic development, has been implicated in the pathogenesis of these conditions through regulation of gene expression, including essential transcription factors. We investigated the expression of four Wnt transcriptional targets, namely, Pygopus 1 (Pygo1), FRA1, TCF7L1 and Connexin 43 (Cx43) in three rare congenital bladder disorders: bladder exstrophy (BE), neurogenic bladder (NGB) and posterior urethral valves (PUV). MethodsBladder tissue samples were collected from patients at the Great Ormond Street Hospital for Sick Children, London, UK, with control (normally-functioning bladder, N=9), BE (N=15), NGB (N=6) and PUV (N=5). Histological analysis was performed using the van Gieson stain to differentiate smooth muscle (SM) and connective tissue (CT) compartments. An unbiased, automated, semi-quantitative immunofluorescence analysis was performed to measure the labelling intensity of four Wnt-related proteins in tissue from these four groups. Results and DiscussionThere was a significant (p<0.05) increase in the expression of Pygo1 in the smooth muscle of all anomalies examined and also in the connective tissue in PUV compared to control. Cx43 also showed overexpression in the smooth muscle across all conditions; however, there was a reduced expression in NGB and an increase in PUV in connective tissue. TCF7L1 showed a significant decrease in both tissue compartments for NGB, whereas FRA1 expression remained unchanged across all anomalies. We also measured colocalization of Wnt-related proteins. TCF7L1 exhibited increased colocalization with Pygo1 and FRA1 in exstrophy compared to control. These results suggest a complex dysregulation of the Wnt pathway in congenital bladder disorders. ConclusionWnt signaling-related proteins show dysregulation in congenital bladder disorders compared to control tissue. Understanding these mechanisms should help towards non-invasive early diagnosis, drug target discovery and development of treatment strategies for these conditions.Summary tableSummary of Wnt-related protein expression and colocalization in bladder exstrophy (BE), neurogenic bladder (NGB) and posterior urethral valves (PUV) disorder compared to normally-functioning control bladder. The table shows the levels of expression as significantly up (↑) down (↓) or no change (-) of Pygopus 1 (Pygo1), Connexin 43 (Cx43), FRA1 and TCF7L1 expression in smooth muscle and connective tissue compartments in BE, NGB and PUV relative to control bladder tissue samples. The table also shows colocalization of six protein pairs of Pygo1 and FRA1; Pygo1 and Cx43; Pygo1 and TCF7L1; FRA1 and Cx43; FRA1 and TCF7L1 and Cx43 and TCF7L1 in bladder disorders compared to control.Summary tableProtein Expression (Smooth muscle)Pygo1Cx43FRA1TCF7L1BE↑↑--NGB↑↑-↓PUV↑↑--Protein Expression (Connective tissue)BE----NGB-↓-↓PUV↑↑--ColocalizationPygo1/FRA1Pygo1/Cx43Pygo1/TCF7L1FRA1/Cx43FRA1/TCF7L1Cx43/TCF7L1Exstrophy--↑-↑-NGB------PUV------

Statin suppresses the development of excessive intimal proliferation in a Kawasaki disease mouse model.

Kawasaki disease (KD) causes vascular injury and lifelong remodeling. Excessive intimal proliferation has been observed, resulting in coronary artery lesions (CALs). However, the mechanisms underlying vascular remodeling in CAL and statin treatment have not been comprehensively elucidated. This study aimed to investigate the effects of statins on vascular remodeling using a KD mouse model. Candida albicans water-soluble substance (CAWS) was intraperitoneally injected in 5-week-old male apolipoprotein-E-deficient mice. They were categorized as follows (n = 4): control, CAWS, CAWS+statin, and late-statin groups. The mice were euthanized at 6 or 10 weeks after injection. Statins (atorvastatin) were initiated after CAWS injection, except for the late-statin group, for which statins were internally administered 6 weeks after injection. Elastica van Gieson staining and immunostaining were performed for evaluation. Statins substantially suppressed the marked neointimal hyperplasia induced by CAWS. Additionally, CAWS induced TGFβ receptor II and MAC-2 expression around the coronary arteries, which was suppressed by the statins. KD-like vasculitis might promote the formation of aneurysm by destroying elastic laminae and inducing vascular stenosis by neointimal proliferation. The anti-inflammatory effects of statins might inhibit neointimal proliferation. Therefore, statin therapy might be effective in adult patients with KD with CAL by inhibiting vascular remodeling.

Cycloastragenol Suppresses Hepatic Stellate Cells Activation to Alleviate Liver Fibrosis Through the PI3K/Akt/mTOR Signaling Pathway and Autophagy

Liver fibrosis is a pathological process of various liver diseases, and a large number of studies have shown that liver fibrosis is reversible. Natural products are one of the important sources of drugs for treating liver fibrosis. The present study evaluated the protective effect of cycloastragenol (CAG) on liver fibrosis and the underlying mechanism. The mouse model of liver fibrosis was established by intraperitoneal injection of carbon tetrachloride (CCl4) and olive oil. Subsequently, CAG was administrated orally at different doses(2.5 mg/kg/d, 5 mg/kg/d, 10mg/kg/d), and the degree of liver fibrosis was assessed using hematoxylin-eosin, Van Gieson, and Sirius Red staining. Alanine aminotransferase, aspartate aminotransferase, hyaluronic acid, and hydroxyproline levels were detected using biochemical analyses. The mRNA and protein expression levels of α-SMA, LC3, P62, PI3K, Akt, and mTOR in the liver tissue were assessed by reverse-transcription polymerase chain reaction and western blot, respectively. The expression of fibrogenesis-related marker α-SMA in the liver tissue was assessed using immunofluorescence. As expected, CAG alleviated liver fibrosis and suppressed hepatic stellate cell activation. And then, CAG inhibited autophagy and activated the PI3K/Akt/mTOR signaling pathway. Thus, CAG attenuated CCI4-induced liver fibrosis in mice by inhibiting autophagy and by activating the PI3K/Akt/mTOR signaling pathway.

CEREBRAL MICROVASCULATURE IN AN EXCLUSIVELY FAT DIET MODEL

The microcirculation system plays a major role in the process of food intake and assimilation by the body. It ensures the distribution of oxygen and nutrients among neurons, taking into account their functional activity. The capillaries of the villi in the choroid plexus of cerebral ventricles remain the main source of cerebrospinal fluid production, which determines most physiological functions of the body. The aim of the study is to identify the peculiarities of remodeling of the microvasculature and vascular plexus of the third cerebral ventricle in rats kept exclusively on a fat diet. Materials and Methods. The work was performed on 20 white mongrel male rats (200–250 g.), divided into control and experimental groups. Animals of the control group were on a regular diet. Rats of the experimental group were fed exclusively with fatty food (sheep tail fat). On the 15th and 30th days, the animals were withdrawn from the experiment. A study of biochemical blood parameters (cholesterol, glucose, and protein) was carried out. After decapitation, the brain was fixed in formalin, brain sections were stained with hematoxylin and eosin (Van Gieson stain). The authors conducted light microscopy and morphometry on an Olympus B×40 microscope (Japan). Results. The animals showed a significant increase in the levels of cholesterol, glucose and albumin in the blood serum under an exclusively fat diet. By the 30th day of the experiment, the smooth muscles of the cerebral arteries undergo paresis, proteolysis, vacuolar dystrophy, hypoplasia with a sharp expansion of the vessel lumen. Signs of myoelastofibrosis are observed in the adventitia. Vein walls are thinned, the lumen is dilated, intravascular thrombi are observed. In the choroid plexus of the 3rd cerebral ventricle, a deficit of plasma flow through the sinusoidal capillaries with compensatory ependymocyte hyperfunction is noted. Conclusion. An exclusively fat diet leads to remodeling of the cerebral microvasculature, including the capillaries of the villi of the choroid plexus of the 3rd ventricle. All changes are compensatory and adaptive in nature. However, by the 30th day of the experiment, some of them become irreversible.

The morphological structure of prostate gland under the condition of experimental prostatopathy and after using of cholecalciferol in the different schemes of hypofertility correction

The impact of negative factors, stress, and modern living conditions damages men's health and leads to infertility. Prostatitis is often a cause of hypofertility. It is now shown that vitamin D may play a role in regulating the functioning of reproductive system organs. The aim of the study was to determine the effect of cholecalciferol on the histological structure of the prostate gland in rats with experimental prostatitis and after its application alone or in combination with a prostate protector. Experimental prostatitis was induced by cold intraoperative damage to the prostate gland. To correct prostatitis, vitamin D3 (cholecalciferol) was administered orally at a dose of 4000 IU. The prostate protector (Prostatilen, Pr) and its pharmaceutical composition, as well as vitamin D3, were administered rectally. Rats with modeled prostatitis were divided into groups: EP (cold experimental prostatitis without treatment); EP + seed oil (on the background of experimental prostatitis, animals received a solvent – apricot kernel oil); EP + vit D3 (per os) (on the background of experimental prostatitis, animals received vitamin D3); EP + Pr (rec) (on the background of experimental prostatitis, males were administered Prostatilen per rectum); EP + vit. D3 (per os) + Pr (rec) (on the background of experimental prostatitis, animals received vitamin D3 (per os) and Prostatilen gel (per rectum)); EP + (vit. D3 + Pr) (rec) (on the background of experimental prostatitis, rats were administered Prostatilen gel with vitamin D3 per rectum). Intact animals (Intact group) and sham-operated rats (Control group) were used as controls. Paraffin sections of the prostate gland were stained with hematoxylin, eosin, and Van Gieson's method. In addition to the review microscopy of the ventral lobes of the prostate gland and the isthmus between them, the power of histochemical reactions was measured, the severity of inflammation and fibrosis was assessed, the number of terminal sections of the prostatic glands with a visually unchanged state, with lumen expansion, and with wall destruction was counted, the longitudinal diameter of the acini and the height of the epithelial cells of the prostatic glands were measured. Statistical analysis of the results was performed using the standard software package "Statistica 6.0" with the use of Student's t-test and nonparametric analog of one-way analysis of variance – Kruskal-Wallis test, as well as Mann-Whitney test. It was found that rats with experimental prostatitis exhibit pronounced changes in the morphological structure of the prostate gland. The prostate-protective effect of vitamin D per os at a dose of 4000 IU was established, which reduced the manifestations of atrophic and destructive processes, signs of tissue inflammation, and coarsening of the prostate gland stroma. Signs of fibrosis development in the prostate gland in males of this group were reduced, and the number of destructive changes and the longitudinal diameter of the terminal sections of the prostatic glands of prostate ventral lobe in rats with experimental prostatitis were decreased. Thus, the addition of cholecalciferol to the basic therapy for infertility has a more pronounced corrective effect on the morphological structure of the prostate than the separate use of cholecalciferol and the prostate protector. Combining basic therapy with vitamin D enhances the prostate-protective properties of the latter and is promising for restoring reproductive function overall.

Changes in the liver of Djungarian hamsters under conditions of a three-month supply of water-soluble silicon of various concentrations

BACKGROUND: Silicon enters the human body through drinking water, air and food. Silicon nanoparticles used in the cosmetic, pharmaceutical and food industries are known to have biological activity. Taking into account the widespread prevalence of silicon compounds, the issue of the safety of its use is becoming more urgent. AIM: To study the effect of water-soluble silicon on the morphological structure of the liver of Djungarian hamsters for three months. MATERIALS AND METHODS: The experiment was carried out on Djungarian hamsters kept in normal vivarium conditions under natural light. The animals were divided into three groups: control, which received bottled drinking water; the first experimental group, which received the same water, but with the addition of sodium metasilicate nine-hydrate at a concentration of 10 mg/l in terms of silicon; the second experimental group, which also received the same water, but with the concentration of sodium metasilicate nine-hydrate doubled (up to 20 mg/l). After three months, the animals were removed from the experiment. Sections were processed by general histological (hematoxylin and eosin, Van Gieson method, toluidine blue), histochemical (monoamine oxidase-positive cells) methods. RESULTS: In the liver of hamsters from the experimental groups, changes in the micromorphological structure were revealed, such as an increase in the nuclear area, nuclear-cytoplasmic ratio of hepatocytes, and the diameter of sinusoidal capillaries. Moreover, more pronounced changes were observed in the liver of hamsters of the second experimental group, such as polymorphic cell infiltration of the portal tracts, an increase in the number of eosinophils, deformation of hepatocyte nuclei and the appearance of apoptotic bodies. A decrease in the area of mast cells and an increase in their number, as well as the number of monoamine oxidase-positive cells in the liver of hamsters of both experimental groups were recorded. CONCLUSIONS: An increase in the concentration of silicon supplied with drinking water in both cases is reflected in the micromorphological structure of the liver of hamsters. Moreover these changes are more pronounced in the liver of hamsters of the second experimental group.

The Potential of Transforming Growth Factor-Beta1 (TGFβ1) in Fibrotic Tissue of Leiomyoma Specimens

Background: Fibrosis can define as the development of excessive fibrous tissue in an organ or tissue. In the situation of the uterus, leiomyoma or scar tissue may involve fibrosis state. The main feature of fibrosis in myoma is the existence of disordered smooth muscle cells and collagen that can be recognized via histological examinations in addition to extracellular component deposition. Leiomyoma is the most public uterine disorder that is mainly consists of smooth muscle cells, myo-fibroblast, and an obvious content of extracellular matrix. The myofibroblast in myoma lesion produced an elevated content of ECM. This paper displays the fibrotic circumstantial in leiomyoma and the mechanisms related to deposition of ECM components. Methods: this study involved (50) patients of 18-80 years old women diagnosed with benign uterine fibroid. Tumor samples were attained by consent from women undergoing hysterectomy or myomectomy at the Al Karama Teaching Hospitals in Kut, Wasit Province, Iraq between October 2022 to March 2023. samples from normal and fibroid were subjected to routein histological processing and subsequently stained with routein stains for general description, Masson Trichrome stain, Van Gieson's stain to visualize collagen content and histochemical stain (Alcian Blue-Periodic Acid Schiff) to evaluate the profile of mucins secreted via tumor cells.Normal myometrium and myoma tissue expression of TGF β1 growth factor was compared using immunohistochemical staining. Qupath and Image J programs were used to analysis the content of fibrous tissue within normal and myoma tissue. Results: Microscopical results revealed that myoma have a proliferation of overlap fusiform cells organized in bundles alienated by variable amounts of connective tissue fibers, enclosed peripherally by a pseudo capsule. Special stained sections analysis revealed deposition of extracellular martix components specially collagen fibers that vary within each nodular fibroid (intrafascicular, interfascicular) enclosed myocytes that reflect the state of fibrosis that appears as blue – green colors enclosed in Masson's trichrome stain, while in Van Gieson's stain appears as red color. Proliferative tumor cells were revealed via Alcian Blue-PAS stain in dark blue color. results showed distribution of perivascular fibers in myoma as compared with normal adjacent myometrium, which appears in red color in Van Gieson's stain. Immunostaining recorded the significantly higher intensity of TGF-β in the leiomyoma tissue than in the normal myometrium (P &lt; 0.05). The effects of the TGF-β1 inhibitor significantly differed between normal myometrium and leiomyoma tissue, with a greater decrease in cell survival in the leiomyoma tissue (P &lt; 0.05). Conclusion: highly fibrous tissue contents showed within myoma and perivascular than adjacent myometrium, these fibrous tissues were detected via special colours in both Masson Trichrome and Van Gieson's stains. The fibrous tissue within leiomyoma showed higher intensity to TGF than normal myometrium.

P163 EMPAGLIFLOZIN AND ITS CARDIOPROTECTIVE EFFECTS IN THE HEART DURING VOLUME OVERLOAD IN AN EXPERIMENTAL MODEL INDUCED BY AORTOCAVAL FISTULA

Mitral regurgitation causes volume overload of the heart, and due to the deregulation of extracellular matrix proteins, the electrical communication between cardiomyocytes provided by connexin channels is disturbed. The aim of this study was to investigate the effect of cardiac volume overload (VO) on its functional parameters, structural remodeling, and connexin43 (Cx43), a protein ensuring electrical synchronization of the heart, and the cardioprotective effect of empagliflozin. Cardiac VO was induced by surgical aortocaval fistula (ACF) in adult, 6-month-old male Wistar rats. Mitral regurgitation and VO developed within four weeks. Subsequently, empagliflozin was administered daily for five weeks at a dose of 10 mg/kg of body weight. Echocardiographic examination showed increased cardiac output and increased Vmax of the aortic valve, decreased ejection fraction and fractional shortening due to ACF-VO. Echocardiographic parameters showed an increase in the diameter of the left ventricle in both systole and diastole, as well as an increase in the volume of the left ventricle at the end of systole and diastole. Empagliflozin normalized the negative parameters in the ACF-VO group. Myocardial protein Cx43 levels were reduced by ACF-VO but normalized by empagliflozin treatment. In addition, empagliflozin suppressed the ACF-VO-induced increase in interstitial collagen detected by van Gieson staining. These findings were consistent with increased protein expression of MMP-2, which is involved in structural remodeling of the extracellular matrix, and was normalized by empagliflozin. Protein kinase C epsilon levels associated with Cx43 and extracellular matrix modulation were reduced by ACF-VO and normalized by empagliflozin. However, protein levels of pro-apoptotic and pro-hypertrophic protein kinase C delta were increased by ACF-VO and normalized by empagliflozin. Protein expression of transforming growth factor beta as well as fibroblast growth factor 21 was increased by ACF-VO and normalized by empagliflozin. The results indicate a cardioprotective effect of empagliflozin due to suppression of myocardial structural remodeling and preservation of Cx43 in an experimental model simulating volume overload. The research was supported by grants APVV-21-0410,VEGA-2/0006/23,VEGA-2/0133/24,VEGA-2/0002/20

Abstract P453: Dissecting tunica responsibility in arterial stress relaxation: Smooth muscle need not apply

Perivascular adipose tissue (PVAT) is being recognized as an important arterial tunica. Stress relaxation, a gradual reduction in stress over time while under a constant strain, is considered a property of smooth muscle (ex. intestine, uterus, bladder). While aortic PVAT (aPVAT) contains a microvasculature, it does not contain organized smooth muscle and isolated PVAT stress relaxes to a greater extent than the vessel itself. This raised the idea that other tunicas lacking smooth muscle may also stress relax. Here we hypothesize that appropriately oriented and functional smooth muscle was not necessary for stress relaxation to occur. To test this, we used histochemical staining and isometric contractility of thoracic aorta from the male Dahl salt-sensitive (SS) rat. Some rings of aorta were separated into adventitia (no muscle), media (muscle), and PVAT (no muscle), and responses of these isolates compared to the intact aortic ring (muscle + integration of tunicas). Masson Trichrome (MT) and Verhoeff Van Gieson (VVG) staining validated the dissection of the different tunicas with minimal contamination of cells from other tunicas. Stress relaxation was measured after increasing, cumulative amounts of passive tension (6 grams total) was applied in the presence or absence of Ca 2+ , essential for smooth muscle contraction. Each step was followed by challenge with a maximum concentration of the a 1 adrenergic agonist phenylephrine (PE, 10 -5 M). Stress relaxation occurred in all tunicas at each passive tension, whether or not Ca 2+ was present (figures). In fact, loss of Ca 2+ improved stress relaxation in adventitia and PVAT. Importantly, in the presence of Ca 2+ , PE contraction was observed at each passive tension in tissues containing smooth muscle: media (min, 250 mg PT: 26.67 +/- 8.51 mg; max, 6000 mg PT: 589.59 +/- 114.53 mg) and aorta + PVAT (min, 250 mg PT: 562.61 +/- 238.45 mg; max, 6000 mg PT: 1322.91 +/- 352.73mg). Little to no measurable contraction was recorded in the adventitia and aPVAT. There was little to no measurable contraction in tunicas in the absence of Ca 2+ . Taken together, these data suggest that all tunicas of the rat thoracic aorta stress relax and can do so without the presence of organized smooth muscle. Of all tunicas, PVAT demonstrated the greatest ability to stress relax. Collectively, these findings force reconsideration of individual responsibilities of arterial tunicas, all of which participate in the (dys)function of an artery.

Quercetin Promotes Abdominal Aortic Aneurysm Regression Through the RAGE/PI3K/AKT/mTOR Axis

Objective: Abdominal aortic aneurysms (AAA) are a common critical cardiovascular disease with high morbidity and mortality rates. Quercetin, a flavonoid and a traditional Chinese medicinal ingredient commonly found in vegetables and fruits, has a favorable inhibitory effect on experimental aneurysms without side effects. This study aimed to model elastase AAA and investigate the mechanism of action of quercetin in alleviating AAA. Methods: Quercetin was administered at a dose of 60 mg/kg once daily starting on the day of AAA induction for 5 weeks. The therapeutic effect of quercetin on AAA was demonstrated using biochemical assays, observation of pathological tissue sections, Elastic Van Gieson staining, immunohistochemistry, molecular docking simulations, and Western blot protein validation. Results: Our study showed that quercetin treatment significantly alleviated abdominal aortic vessel wall dilatation, elastin degradation and adhesion to surrounding tissues caused by elastase. Additionally, quercetin significantly inhibited the mRNA and protein expression of the receptor for advanced glycation end products(RAGE)/ phosphatidylinositol 3-kinase (PI3K)/ phosphatidylinositol 3-kinase (AKT) / mammlian target of rapamycin (mTOR) pathway. Conclusion: Quercetin can alleviate abdominal aortic injury caused by elastase via RAGE/PI3K/AKT/mTOR and provide experience in clinical use.

Application of a xenopericardial plate for closing a laparostomy in conditions of a purulent-inflammatory process

The aim of the study is to study the morphological changes in tissue in the laparostomy area when it is closed with a xenopericardial plate under conditions of an active purulent-inflammatory process. Material and methods. Analysis of data from patients at the Penza regional clinical hospital named after. N.N. Burdenko, with diseases of the abdominal organs, complicated by diffuse purulent peritonitis. During treatment, xenopericardial plates with perforations in a checkerboard pattern were used (hole diameter 0.5 cm). Disinfection of the material was carried out with a sterile solution of furacilin. Using the standard hematoxylin and eosin staining protocol, as well as alternative staining methods according to Van Gieson and Weigert, light-optical examination of stained sections was carried out at a magnification of 40–400 times. Results. In 8 out of 9 patients participating in the study, a favorable course of the inflammatory process was observed; on the 21st day after laparotomy, the symptoms of peritonitis were eliminated, the surgical wound was sutured. In one case, progression of local and general signs of peritonitis was observed within two weeks. After replacing the xenopericardial plate, the course of changes in the surgical wound was favorable. On day 21, the laparotomy wound was also sutured. Repeated examination of the biomaterial one year and two months after implantation of the xenopericardial plate into the anterior abdominal wall did not reveal signs of inflammation. Conclusions. The xenopericardium performed well in conditions of purulent inflammation and in most cases was not subject to melting. In the long term, biointegration of the xenopericardium into its own connective tissue occurred.

Animal models of pulmonary arterial hypertension associated with atrial septal defect

Pulmonary arterial hypertension (PAH) is a well-known complication of congenital heart disease (CHD). The lack of a satisfactory animal model for PAH associated with CHD (PAH-CHD) has limited progress in understanding the pathogenesis of PAH and the development of therapeutic agents. The development of a rat model for PAH associated with atrial septal defect (ASD) was achieved through atrial septal puncture and thermal ablation. Two and 4 weeks after modeling, hematoxylin and eosin staining showed that the vascular thickness, vascular thickness index, vascular area, and vascular area index in pulmonary arteries with an outer diameter of 50–300 μm in the PAH-ASD 2 and 4 weeks group were higher than those in the sham group (all P < 0.05). Alpha-smooth muscle actin (ɑ-SMA) staining showed that the medial thickness, medial thickness index, medial area, and medial area index in pulmonary arteries with an outer diameter of 50–300 µm at 2 and 4 weeks after modeling were significantly higher than those in the sham group (all P < 0.05). Additionally, mean pulmonary arterial pressure (mPAP) and pulmonary vascular resistance (PVR) in the PAH-ASD 2 and 4 weeks groups were significantly higher than those in the sham group (both P < 0.05). Elastin van Gieson staining showed that the vascular obstruction score in the PAH-ASD 2 and 4 weeks group was significantly higher than that in the sham group (both P < 0.05). The PAH-ASD rats were successfully generated. These findings suggest that our model would be useful for further research into the pathogenesis, prevention, and treatment of PAH-ASD.

ОСОБЕННОСТИ СТРОЕНИЯ СЕМЯПРОВОДОВ У КРОЛИКОВ

The aim of the study is to investigate the anatomical and histological structure of the ductus deferens in representatives of the order Lagomorpha. The results of the micromorphology of the vas deferens in representatives of the order Lagomorpha are presented. Histological preparations were prepared and exami¬ned at the Omsk State Agrarian University (Department of Anatomy, Histology, Physiology and Pathological Anatomy). The objects of the study were material obtained from five sexually mature male Californian rabbits. For histological examination, pieces of organs were selected using the generally accepted method. Sections were stained with hematoxylin and eosin, as well as the Van Gieson method. Morphological, histological, morphometric and statistical methods were used in the study. The ductus deferens in rabbits is a paired tubular organ. The thickness of the mucous membrane of the left ductus deferens varies within 85.77 ± 3.13 μm, of the right ductus deferens was 136.14 ± 3.20 μm. The thickness of the muscular membrane of the left ductus deferens was 287.16 ± 3.48 μm, of the right ductus deferens was 333.79 ± 4.07 μm. The thickness of the adventitia of the left ductus deferens was 246.08 ± 4.75 μm, of the right ductus deferens was 258.56 ± 3.84 μm. During the study, special attention was paid to the adaptation of the muscular layer, which demonstrates a high degree of development and the ability to powerful peristaltic contractions, ensuring the movement of sex cells to the urethra. We can also assume the formation of sphincter systems of the ductus deferens in the indicated anatomical areas.

Morphological and functional spleen development in crossbreed rabbits

The accelerated growth of muscle mass in productive broiler breeds is often associated with delayed organ development of integral body systems, particularly the immune structures. The spleen is the largest secondary immune organ in the mammalian body and is responsible for initiating immune responses to blood-borne antigens. It can only perform this function successfully if all of its tissue components are fully morphologically and functionally mature. The spleen was studied in meat production rabbits (early maturing crossbreed Hyplus) at 1, 10, 20, 30, 60 and 90 days of age. Morphological studies included anatomical dissection, clarification of topography, determination of mass parameters, preparation of smears and histological sections. Spleen histological sections were stained with haematoxylin and eosin, according to Van Gieson, and impregnated with silver nitrate, followed by microscopy. Qualitative and quantitative indicators of spleen cellular and tissue component development were determined using ImageJ software. It was found that in crossbred rabbits the topography of the spleen corresponds to general anatomical principles of localisation, has a fixed place and is an anatomically formed organ. In day-old animals, the histological differentiation of the spleen is limited to the connective tissue stroma and the parenchyma. The organ's parenchyma is formed by reticular tissue, with hematopoietic and lymphoid cells among the cells, without differentiation into white and red pulp. In 10-day-old rabbits, the white pulp is represented by the periarterial lymphatic sheath (PALS) and marginal zones. Single primary lymphoid nodules without germinal centres are seen in the spleen from 20 days of age. By 30 days of age, the white splenic pulp has all the major structural and functional zones, including formed lymphoid nodules with germinal centres and mantle zones. During the second and third months of life, the spleen gradually increases the relative area of all white pulp functional zones, reaching a maximum at 90 days of age. In productive rabbits, the cellular composition of the splenic white pulp is represented by lymphocytes (small, medium, large), reticular cells, macrophages and plasma cells. During postnatal ontogenesis, the number of small lymphocytes increases in all white pulp functional zones, reaching maximum values in 90-day-old animals. On the contrary, the relative number of medium and large lymphocytes decreases. The number of reticulocytes in the periarterial lymphatic sheath and lymphoid nodules zones does not change significantly, and in the marginal zone it decreases substantially by the end of the study. The results of determining the timing of morphological and functional maturation of immunocompetent structures in the spleen of meat rabbits are of great biomedical and economic importance. They will serve as a control for comparing changes in rabbit spleen lymphoid tissues during the development of pathological processes, as well as under the influence of external factors.