Sacubitril/valsartan reverses cardiac structure and function in experimental model of hypertension-induced hypertrophic cardiomyopathy.

This study evaluated the effect of sacubtril/valsartan on cardiac remodeling, molecular and cellular adaptations in experimental (rat) model of hypertension-induced hypertrophic cardiomyopathy. Thirty Wistar Kyoto rats, 10 healthy (control) and 20 rats with confirmed hypertension-induced hypertrophic cardiomyopathy (HpCM), were used for this study. The HpCM group was further subdivided into untreated and sacubitril/valsartan-treated groups. Myocardial structure and function were assessed using echocardiography, Langendorff's isolated heart experiment, blood sampling and qualitative polymerase chain reaction. Echocardiographic examinations revealed protective effects of sacubitril/valsartan by improving left ventricular internal diameter in systole and diastole and fractional shortening. Additionally, sacubitril/valsartan treatment decreased systolic and diastolic blood pressures in comparison with untreated hypertensive rats. Moreover, sacubitril/valsartan treatment reduced oxidative stress and apoptosis (reduced expression of Bax and Cas9 genes) compared to untreated rats. There was a regular histomorphology of cardiomyocytes, interstitium, and blood vessels in treated rats compared to untreated HpCM rats which expressed hypertrophic cardiomyocytes, with polymorphic nuclei, prominent nucleoli and moderately dilated interstitium. In experimental model of hypertension-induced hypertrophic cardiomyopathy, sacubitril/valsartan treatment led to improved cardiac structure, haemodynamic performance, and reduced oxidative stress and apoptosis. Sacubitril/valsartan thus presents as a potential therapeutic strategy resulted in hypertension-induced hypertrophic cardiomyopathy.

Function of s100 protein in coronary atherosclerosis / Función de la proteína s100 en aterosclerosis coronaria

SUMMARY: Atherosclerosis is a complex disease whose pathogenesis includes endothelial activation, accumulation of lipids in the subendothelium, formation of foam cells, fat bands and formation of atherosclerotic plaque. These complex mechanisms involve different cell populations in the intimate sub-endothelium, and the S-100 protein family plays a role in a number of extracellular and intracellular processes during the development of atherosclerotic lesions. The aim of this study was to determine the phenotypic characteristics of smooth muscle cells and the consequent expression of S100 protein in atherosclerotic altered coronary arteries in advanced stages of atherosclerosis. 19 samples of right atherosclerotic coronary arteries in stages of fibro atheroma (type V lesion) and complicated lesions (type VI lesion) have been analyzed. According to the standard protocol, the following primary antibodies have been used in the immunohistochemical analysis: a-smooth muscle actin (α-SMA), vimentin and S-100 protein. All analyzed samples have been in advanced stages of atherosclerosis, fibro atheroma (stage V lesions) and complicated lesions (type VI lesions). Most of them have had the structure of a complicated lesion with atheroma or fibro atheroma as a basis, subsequently complicated by disruption (subtype VI a), hemorrhage (subtype VI b) or thrombosis (subtype VI c), as well as by the presence of several complications on the same sample. Marked hypocellularity is present in the subendothelium of plaques. Cell population at plaque margins is characterized by immunoreactivity to α-SMA, vimentin, and S100 protein. Some of these cells accumulate lipids and look like foam cells. In the cell population at the margins of the plaques, smooth muscle cells of the synthetic phenotype are present, some of which accumulate lipids and demonstrate S100 immunoreactivity. Summarizing numerous literature data and our results, we could assume that smooth muscle cells, due to their synthetic and proliferative activity in the earlier stages of pathogenesis, as well as the consequent expression of S100 protein, could accumulate lipids in the earlier stages of atherosclerosis which, in advanced stages analyzed in this study, result in immunoreactivity of foam cells of smooth muscle origin to S100 protein.

RESUMEN: La aterosclerosis es una enfermedad compleja cuya patogenia incluye activación endotelial, acumulación de lípidos en el subendotelio, formación de células espumosas, bandas grasas y formación de placa aterosclerótica. Estos complejos mecanismos involucran diferentes poblaciones celulares en el subendotelio íntimo, y la familia de proteínas S-100 juega un papel en varios procesos extracelulares e intracelulares durante el desarrollo de lesiones ateroscleróticas. El objetivo de este estudio fue determinar las características fenotípicas de las células de músculo liso y la consecuente expresión de la proteína S100 en arterias coronarias alteradas ateroscleróticas en estadios avanzados de aterosclerosis. Se analizaron 19 muestras de arterias coronarias ateroscleróticas derechas en estadios de fibroateroma (lesión tipo V) y lesiones complicadas (lesión tipo VI). Según el protocolo estándar, en el análisis inmunohistoquímico se utilizaron los siguientes anticuerpos primarios: α-actina de músculo liso (α-SMA), vimentina y proteína S-100. Todas las muestras analizadas han estado en estadios avanzados de aterosclerosis, fibroateroma (lesiones estadio V) y lesiones complicadas (lesiones tipo VI). La mayoría de ellos han tenido la estructura de una lesión complicada con ateroma o fibroateroma como base, complicada posteriormente por disrupción (subtipo VI a), hemorragia (subtipo VI b) o trombosis (subtipo VI c), así como por la presencia de varias complicaciones en la misma muestra. La hipocelularidad marcada estaba presente en el subendotelio de las placas. La población celular en los márgenes de la placa se caracterizaba por inmunorreactividad a α-SMA, vimentina y proteína S100. Algunas de estas células acumulan lípidos y parecen células espumosas. En la población celular en los márgenes de las placas, estaban presentes las células de músculo liso de fenotipo sintético, algunas de las cuales acumulaban lípidos y mostraban inmunorreactividad S100. Resumiendo numerosos datos de la literatura y nuestros resultados, podríamos suponer que las células del músculo liso, debido a su actividad sintética y proliferativa en las primeras etapas de la patogénesis, así como la consecuente expresión de la proteína S100, podrían acumular lípidos en las primeras etapas de la aterosclerosis que, en estadios avanzados analizados en este estudio, dan como resultado inmunorreactividad de células espumosas de origen muscular liso a la proteína S100.

The effect of final irrigation with MTAD, QMix, and EDTA on smear layer removal and mineral content of root canal dentin.

To compare the smear layer removal ability and mineral content of root canal dentine after initial irrigation with NaOCl and final irrigation with MTAD, QMix, and 17% EDTA. Forty extracted human maxillary incisors before root canal preparation and irrigation with NaOCl were randomly divided into four groups (n = 10) according to the type of final irrigants used: MTAD, QMix, 17% EDTA, and control (sterile distilled water). Scanning electron microscopy (SEM) was used to assess the presence of smear layer. SEM energy-dispersive X-ray spectroscopy was used to quantify dentin mineral composition in MTAD, QMix, 17% EDTA group, and in no-treatment samples (no-treatment group; n = 10). Among the various chelating agents, there were no significant differences in the smear layer removal in the middle and coronal thirds (p > .05). In the apical third, QMix removed significantly more smear layer than 17% EDTA (p < .05), but similarly to MTAD (p > .05). Final irrigation with MTAD resulted in a significant increase in the carbon (C) value compared to EDTA (p < .001). There was no significant difference in the mineral composition between the MTAD and the QMix group, although the values of the mineral elements were significantly altered in the MTAD group. QMix had smear layer removal capability similar to MTAD but better than EDTA in the apical third. MTAD yielded the most pronounced effect on mineral component of root dentin; however, differences were significant only for C level compared to 17% EDTA.

Histochemical, immunohistochemical and ultrastructural analysis of aortic wall in neonatal coarctation.

The neonatal type of coarctation is characterized by the presence of the ductal sling and coarctational shelf placed proximally in relation to the ductal orifice. Those morphological features are not described in detail yet from immunohistochemical and transmission electron microscopy (TEM) aspects, so the aim of this study was to investigate the smooth muscle cells (SMCs) phenotype in aortic intimal thickening, presence of inflammatory cells and contents of intimal and medial, and adventitial connective tissue. We examined samples of coarctation segments excised at surgery after end-to-end anastomosis from 30 patients, ages from 14 days to three months, histochemicaly, immunocytochemically and by TEM. In all samples, it is noticed focal intimal thickening on the posterior aortic wall, with accumulation of SMCs, which show immunoreactivity on alpha-smooth muscle actin (α-SMA) and vimentin (but not on desmin) and also expressed proliferating cell nuclear antigen (PCNA) and S-100 protein. At TEM analysis, those SMCs show a fibroblast-like morphology, so their functions could be to proliferate and secrete extracellular matrix (ECM) components (a synthetic phenotype). In all studied samples of the coarctation, on the posterior wall, the immunocytochemical and TEM examination revealed the presence of SMCs of the synthetic phenotype. Results also showed an increase of the cell number in intima of this part of aortic wall, followed by proliferated SMCs in inner media and absence of inflammatory cells. This finding suggests that proliferation of the SMCs, their synthetic activity and increase of the cell number could lead to formation of the intimal thickening on the posterior wall.

The mineral content of the hard dental tissue of mesiodens.

OBJECTIVE: Mesiodens is the most common form of supernumerary tooth mainly located between the maxillary central incisors. Its etiology is not completely understood but both genetic and environmental factors are assumed. The degree of mineralization and inorganic element content in hard tooth tissues is poorly understood as well as is the durability and suitability for allo- and auto-transplantation. Therefore aim of this study was to examine the content of inorganic elements. MATERIALS AND METHODS: This study included 26 mesiodens teeth and 26 normal central incisor teeth as controls. All specimens were prepared for SEM/EDS analysis which was aimed at specific sites on the enamel, dentine and cementum in order to evaluate the weight percentage and ratio of important inorganic elements. RESULTS: and Conclusion. The results showed that there was a difference in the weight percentage of selected inorganic elements (calcium, phosphorus, oxygen, carbon, magnesium and sodium) in all three types of dental hard tissues but the differences were mostly expressed in the cementum tissue. The statistical analysis showed that the differences were marginally significant especially for calcium and phosphorus values and ratio in the enamel and dentine. The carbon and magnesium content in all three hard tissues showed the most differences, but overall, the hard tissues mineral content of the mesiodens did not differs significantly from healthy teeth.

Murine Cytomegalovirus Infection Induces Susceptibility to EAE in Resistant BALB/c Mice.

In contrast to C57BL/6 mice, BALB/c mice are relatively resistant to the induction of experimental autoimmune encephalomyelitis (EAE) after challenge with MOG35-55 peptide. Here, we provide the first evidence that infection with murine cytomegalovirus (MCMV) in adulthood abrogates this resistance. Infected BALB/c mice developed clinical and histological signs similar to those seen in susceptible C57BL/6 mice. In addition to CD4+ cells, large proportion of cells in the infiltrate of diseased BALB/c mice was CD8+, similar with findings in multiple sclerosis. CD8+ cells that responded to ex vivo restimulation with MOG35-55 were not specific for viral epitopes pp89 and m164. MCMV infection favors proinflammatory type of dendritic cells (CD86+CD40+CD11c+) in the peripheral lymph organs, M1 type of microglia in central nervous system, and increases development of Th1/Th17 encephalitogenic cells. This study indicates that MCMV may enhance autoimmune neuropathology and abrogate inherent resistance to EAE in mouse strain by enhancing proinflammatory phenotype of antigen-presenting cells, Th1/Th17, and CD8 response to MOG35-55.

Computational and experimental model of electroporation for human aorta.

PURPOSE: In this study the computational and experimental electroporation model with human aorta tissue is made in order to examine the reduction of smooth muscle cells. METHODS: The segments in native state of the aorta are treated by electroporation method through a series of electrical impulses from 50 V/cm to 2500 V/cm. For each patient we analyzed one sample with and one sample without electroporation as a control. In the computational study, electrical field distribution is solved by the Laplace equation. The Pennes Bioheat equation without metabolism and blood perfusion heating is used to solve heat transfer problems. Different conductivity values are used in order to fit the experimental results. RESULTS: Experimental histology has shown us that there are a smaller number of vascular smooth muscle cells (VSMC) nuclei at the tunica media, while the elastic fibre morphology is maintained 24 h after electroporation. In the computational model, heat generation coupled with electrical field is included. The fitting procedure is applied for conductivity values in order to make material properties of the aorta tissue. The fitting procedure gives tissue conductivity of 0.44 [S/m] for applied electrical field of 2500 V/cm. CONCLUSIONS: Future studies are necessary for investigation of a new device for in-vivo ablation with electroporation of plaque stenosis. It will open up a new avenue for stenosis treatment without stent implantation.

Computer simulation of cervical tissue response to a hydraulic dilator device.

BACKGROUND: Classical mechanical dilators for cervical dilation are associated with various complications, such as uterine perforation, cervical laceration, infections and intraperitoneal hemorrhage. A new medical device called continuous controllable balloon dilator (CCBD) was constructed to make a significant reduction in all of the side effects of traditional mechanical dilation. METHOD: In this study we investigated numerically the cervical canal tissue response for Hegar and CCBD using our poroelastic finite element model and in-house software development. Boundary conditions for pressure loading on the tissue for both dilators in vivo were measured experimentally. Material properties of the cervical tissue were fitted with experimental in vivo data of pressure and fluid volume or balloon size. RESULTS: Obtained results for effective stresses inside the cervical tissue clearly showed higher stresses for Hegar dilator during dilation in comparison with our CCBD. CONCLUSION: This study opens a new avenue for the implementation of CCBD device instead of mechanical dilators to prevent cervical injury during cervical dilation.

Continuous controllable balloon dilation: a novel approach for cervix dilation.

BACKGROUND: Cervical dilation using mechanical dilators is associated with various complications, such as uterine perforation, cervical laceration, infections and intraperitoneal hemorrhage. To achieve safe and painless cervical dilation, we constructed a new medical device to achieve confident mechanical cervical dilation: a continuous controllable balloon dilator (CCBD). METHODS: Controlled pumping of incompressible fluid into the CCBD increases the pressure and outer diameter of the CCBD, continuously dilating the cervical canal. The reliability of the CCBD was confirmed in vitro (testing for consistency and endurance, with no detected risk for breakage) and in vivo. A multi-center clinical study was conducted,with 120 pregnant women randomly assigned to one of three groups: Group I,control group, no dilation;Group II,mechanical dilation, Hegar dilator (HeD); and Group III,CCBD. The tissue material for histological evaluation was obtained from the endocervical mucosa before and after dilation using the HeD or CCBD. RESULTS: The CCBD dilations were successful and had no complications in all 40 patients of Group III. The cervical tissue was markedly less damaged after CCBD dilation compared with HeD dilation (epithelium damage: 95% (HeD) vs. 45% (CCBD), P <0.001; basal membrane damage: 82.5% (HeD) vs. 27.5% (CCBD), P <0.001; stromal damage: 62.5% (HeD) vs. 37.5% (CCBD), P <0.01). Cervical hemorrhagia was observed in 90% of the patients after HeD dilation versus in 32.5% of the patients after CCBD dilation. CONCLUSIONS: The CCBD should be used as a replacement for mechanical dilators to prevent uterine and cervical injury during cervical dilation. TRIAL REGISTRATION: ISRCTN54007498.

Dystocia as a cause of untimely cesarean section.

BACKGROUND/AIM: One of the most frequent indications for cesarean section is dystocia. It is impossible to predict, difficult to identify and coincident with the rapid expiry of the expected time, so it is important to point out some mistakes in expecting vaginal delivery. The aim of this study was to examine the frequency and the length of dystocia-related cesarean delivery, as well as the vitality of the newborn immediately after birth. METHODS: A prospective 3-year study was conducted including a total number of 6470 deliveries regardless of whether they were completed using cesarean section after an unsuccessful attempt of spontaneous vaginal delivery or not. The Apgar score, a proved useful tool for the assessment of the vitality of newborn children in the first minute, was used. RESULTS: On the basis of the established indications, 653 (10.10%) of deliveries were completed using cesarean section. Dystocia was the third most common indication for cesarean section (16.38%). Deliveries in which dystocia was established as a diagnosis lasted much longer (p = 0.030) which resulted in weaker vitality of newborn children (p = 0.000) compared to the deliveries ended by spontaneous vaginal delivery. CONCLUSION: This study shows that deliveries caused by dystocia last much longer and newborn children are of weaker vitality compared to other deliveries caused not by dystocia. Decisions concerning cesarean section must be made in a timely fashion.

ARTreat Project: three-dimensional numerical simulation of plaque formation and development in the arteries.

Atherosclerosis is a progressive disease characterized by the accumulation of lipids and fibrous elements in arteries. It is characterized by dysfunction of endothelium and vasculitis, and accumulation of lipid, cholesterol, and cell elements inside blood vessel wall. In this study, a continuum-based approach for plaque formation and development in 3-D is presented. The blood flow is simulated by the 3-D Navier-Stokes equations, together with the continuity equation while low-density lipoprotein (LDL) transport in lumen of the vessel is coupled with Kedem-Katchalsky equations. The inflammatory process was solved using three additional reaction-diffusion partial differential equations. Transport of labeled LDL was fitted with our experiment on the rabbit animal model. Matching with histological data for LDL localization was achieved. Also, 3-D model of the straight artery with initial mild constriction of 30% plaque for formation and development is presented.

Ovarian endometroid adenocarcinoma in pregnancy.

BACKGROUND: Ovarian cancer is very rare in pregnancy. It is mainly of epithelial origin, low grade and low malignant potential. CASE REPORT: We presented a patient in which ultrasound confirmed the presence of clearly limited tumor in the left ovary when she was eight weeks pregnant. The results of 4D Color Doppler showed a central type of tumor vascularisation with resistance index (IR) less than 0.5. The Consultancy Board in Gynecology of the Institute for Oncology and Radiology of Serbia decided to remove the patient's left adnexa and intensively monitor the pregnancy period. The operation (Adnexectomia lateralis sinistra) was performed at 18th week of gestation. Histopathological analysis showed adenocarcinoma invasivum, endometroid well-differentiated type (histological grade I, nuclear grade 2). In 37th week of gestation, the patient gave birth to a male child by cesarean section. In the next 3 years the patient had no subjective interference, laboratory tests and ultrasonographic findings were normal. CONCLUSION: Ovarian cancer in pregnancy is usually asymptomatic and diagnosed during routine clinical and ultrasound examination. The color Doppler technique have particular importance in the diagnosis of pathological blood supply in tumors and in indication of malignant ovarian mass.

Computer simulation and experimental analysis of LDL transport in the arteries.

Atherosclerosis develops from oxidized low-density lipoprotein molecules (LDL). When oxidized LDL evolves in plaque formations within an artery wall, a series of reactions occur to repair the damage to the artery wall caused by oxidized LDL. Macrophages accumulate inside arterial intima, they started to collect oxidized LDL and form foam cells. Smooth muscle cells accumulate in the atherosclerotic arterial intima, where they proliferate and secrete extracellular matrix to form a fibrous cap. In this study, experimental model of LDL transport on the isolated blood vessel from rabbit on high fat diet after 8 weeks is simulated numerically by using a specific model and histological data. The 3D blood flow is governed by the Navier-Stokes equations, together with the continuity equation. Mass transfer within the blood lumen and through the arterial wall is coupled with the blood flow by the convection-diffusion equation. LDL transport in lumen of the vessel is described by Kedem-Katchalsky equations. The inflammatory process is solved using three additional reaction-diffusion partial differential equations. Matching of histological rabbit data is performed using 3D histological image reconstruction and 3D deformation of elastic body. Computed concentrations of labeled LDL of 5.2 % and macrophages distribution of 4.2% inside the media are found to be in good agreement with experimental results. This simulation study provides a useful tool for understanding and prediction of LDL transport through the arterial wall and evolution of atherosclerotic plaques.

[Histochemical and immunohistochemical analysis of ruptured atherosclerotic abdominal aortic aneurysm wall].

BACKGROUND/AIM: The main complication of the atherosclerotic abdominal aortic aneurysm (AAA) is her rupture that begins with lesion in intima and rupture. The purpose of this work was to determine immunocytochemical and morphofunctional characteristics of the cells in aortic wall in ruptured atherosclerotic abdominal aortic aneurysm. METHOD: During the course of this study, 20 samples of atherosclerotic AAA were analyzed, all of them obtained during authopsy. The samples were fixed in 4% formalin and embedded in paraffin. Sections of 5 microm thickness were stained histochemically (of Heidenhain azan stain and Periodic acid Schiff--PAS stain) and immunocytochemically using a DAKO LSAB+/HRP technique to identify alpha-smooth muscle actin (alpha-SMA), vimentin, myosin heavy chains (MHC), desmin, S-100 protein, CD45 and CD68 (DAKO specification). RESULTS: The results of our study showed that ruptured atherosclerotic AAA is characterized by a complete absence of endothelial cells, the disruption of basal membrane and internal elastic lamina, as well as a presence of the remains of hypocellular complicated atherosclerotic lesion in intima. On the plaque margins, as well as in the media, smooth muscle cells (SMCs) are present, which express a alpha-SMA and vimentin (but without MHC or desmin expression), as well as leukocyte infiltration, and a large number of foam cells. Some of the foam cells show a CD68- immunoreactivity, while the others show vimentin- and S-100 protein-immunoreactivity. Media is thinned out with a disorganized elastic lamellas, while adventitia is characterized by inflammatory inflitrate (infection). CONCLUSION: Rupture of aneurysm occurs from the primary intimal disruption, which spreads into thinned out media and adventitia. Rupture is caused by unstable atherom, hypocellularity, loss of contractile characteristics of smooth muscle cells in intima and media, neovascularization of the media, as well as by the activity of the macrophages in the lesion.

[Immunocytochemical characteristics of submucosal uterine myomas].

BACKGROUND/AIM: Myomas of the uterus, the most common benign tumors, have been studied for decades from the aspects of different basic and clinical disciplines. Despite this fact, their pathogenesis is still poorly understood. The aim of this study was to determine immunocytochemical characteristics of smooth muscle cells and connective tissue components of submucosal myomas of the uterus. METHOD: During the course of this study, 25 samples of submucosal myomas of the uterus were analyzed, all of them obtained during the surgery, after abdominal histerctomy by Aldridge. The samples were fixed in 4% formalin and embedded in paraffin. Sections of 5 microm thickness were stained immunocytochemically using the DAKO LSAB+/HRP technique to identify alpha-smooth muscle actin (alpha-SMA), vimentin, desmin, CD34, CD45, CD68 and PCNA (DAKO specification). RESULTS: Our results suggest that submucosal myomas of the uterus are build-up of smooth muscle cells which are immunoreactive to alpha-SMA and desmin, but also to a certain number of smooth muscle cells which are immunoreactive to alpha-SMA and vimentin. Some of vimentin-immunoreactive cells also show an immunoreactivity of PCNA. In the build-up of connective stroma CD34-immunoreactive fibroblasts and neovascular formations are also present. By examining the distribution of CD45 antigen, at all the analyzed samples we observed a weak reaction. CONCLUSION: Submucosal myomas of the uterus are made-up of smooth muscle cells of the highly differentiated contractile phenotype (alpha-SMA- and desmin-immunoreactivity), as well as smooth muscle cell of the synthetic phenotype which proliferate (alpha-SMA-, vimentin- and PCNA-immunoreactivity). In submucosal myoma of the uterus there is a significant presence of connective tissue as a result of synthetic activity of fibroblasts, which clearly differ in their immunocytochemical characteristics from smooth muscle cells of the synthetic phenotype.