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Aim: The authors aimed to quantitatively assess the variation with age of three of the main components of the aortic wall, namely elastic fibers (FE) and collagen fibers [FC(COL)], and smooth muscle cells (SMCs). Materials and Methods: Four aortic cross sections (base, cross, thoracic, abdominal) were collected during autopsy from 90 cases of all ages, processed using the classical histopathological (HP) technique (formalin fixation and paraffin embedding) and stained with Orcein and Goldner’s trichrome. The obtained histological slides were transformed into virtual slides. Quantitative measurements of the three components were made on identical regions of interest (ROIs) selected from two-paired slides stained with the above-mentioned techniques using custom-made software, developed in MATLAB (MathWorks, USA). Results and Discussions: FE revealed an obvious decreasing general trend with age, present at all four levels of investigation. Smooth muscle fibers (FM) density showed almost no variation with age regardless of the level at which the measurement was made. FC(COL) density had an obvious increasing trend with age, expressed in all four aortic regions FE densities and FM densities were higher in men than in women, while FC(COL) densities were higher in women than in men in three of the aortic regions excepting, in all cases, the cross region. Conclusions: There were differences between men and women concerning the composition of aortic wall cellular and extracellular compartments. FE and FC(COL) dominated the age-related remodeling process of the aortic wall. The process evolved in the same way in all regions of the aorta. Studies need to be continued to define more clearly this complex process of vascular wall remodeling with aging.

The primo vascular system (PVS) is observed in different parts of the body under different physiological and disease conditions. Previously, the PVS was not observed in the vagina. The vaginal samples of this study were collected from the female genitalia of healthy New Zealand white rabbits from the animal house, Faculty of Medicine, Assiut University. The vaginal samples were fixed in Bouin's solution. The sections were stained with hematoxylin and eosin and Crossmon's trichrome. Additionally, the sections were immunohistochemically stained with neuron-specific enolase (NSE) and vascular endothelial growth factor (VEGF). A primo node was observed on the lymph vessel of the vagina and has several characteristics that resemble those of the previously discovered primo nodes. The primo node in this study was surrounded by mesothelial cells that provide positive immunoreactivity to NSE and VEGF. Sinuses of different sizes, floating cells, telocyte-like cell, and primo microcells were observed as the main constituents of the primo node. Additionally, migratory cells were detected, which passed from the primo node to the enclosing lymph vessel.

Objective Dientamoeba fragilis (D. fragilis) is a flagellated protozoan with an amoeba-like morphology, located in the gastrointestinal tract. The hypothesis was that the parasite was transported by Enterobius vermicularis (E. vermicularis) eggs. This study aimed to determine the association of D. fragilis and E. vermicularis with the genotypes of the identified strain of D. fragilis. Results of trichrome staining were compared with those of polymerase chain reaction (PCR), which is widely used in the diagnosis of D. fragilis. Methods A total of 391 samples were obtained. The stool and cellophane slide samples were sent together to the Parasitology Department Laboratory, Faculty of Medicine, Aydin Adnan Menderes University, between 1 October 2017 and 1 October 2018. Stool samples of all patients with E. vermicularis (n=74) and without E. vermicularis (n=74) infection were used. All samples were examined for the presence of D. fragilis by trichrome staining and PCR. The 18S ribosomal RNA region of D. fragilis isolates was sequenced. Demographic characteristics and clinical findings of the patients were evaluated. Results D. fragilis was detected in 42 (28.37%) of 148 samples; 28 (66.6%) of them were detected in patients with E. vermicularis infection. The coexistence of two parasites was significant (p 0.05). Conclusion D. fragilis is frequently associated with E. vermicularis, so the presence of D. fragilis should be also considered in affected patients. The use of high-sensitivity molecular methods such as PCR is important in preventing false results. Amac Dientamoeba fragilis (D. fragilis), amip benzeri morfolojiye sahip, gastrointestinal yerlesimli, kamcili bir protozoondur. Parazitin Enterobius vermicularis (E. vermicularis) yumurtalariyla tasindigi hipotezi kabul gormektedir. Calismamizda D. fragilis ve E. vermicularis birlikteligini incelemek, bulunan D. fragilis’lerin genotiplerini belirlemek ve D. fragilis tanisinda yaygin olarak kullanilan trikrom boyama ile polimeraz zincir reaksiyon (PZR) yontemlerini karsilastirmak amaclanmistir. Yontemler Calismamizda Aydin Adnan Menderes Universitesi Tip Fakultesi, Parazitoloji Laboratuvari’na 1 Ekim 2017-1 Ekim 2018 tarihleri arasinda diski ve selofan lam ornegi birlikte gonderilmis toplam 391 olgu ornegi incelenmistir. Selofanli lam orneklerinde E. vermicularis saptanan tum gonullu olgularin (74 olgu) diski ornegi ile E. vermicularis negatif 74 olgunun diski ornegi calisilmistir. Tum diskilar trikrom boyama ve PZR yontemleri ile D. fragilis varligi acisindan incelenmistir. Saptanan D. fragilis izolatlarinin 18S ribozomal RNA bolgesi sekanslanmistir. Olgularin demografik ozellikleri ve klinigi degerlendirilmistir. Bulgular Toplam 148 olgunun 42’sinde (%28,37) D. fragilis saptanmistir. D. fragilis pozitif olan 42 olgunun %66,6’sini E. vermicularis pozitif olgular olusturmus ve iki parazitin birlikteligi anlamli bulunmustur (p 0,05). Sonuc Arastirmamizda D. fragilis’in siklikla E. vermicularis ile birliktelik gosterdigi ve bu olgularda D. fragilis varligina ayrica dikkat edilmesi gerektigi vurgulanmistir. Yanlis sonuclari engellemede, yuksek duyarliliga sahip PZR gibi yontemlerin onemi bir kez daha gorulmustur.

The current study investigated the morphological, histochemical, and immunohistochemical characteristics of alarm cells and their precursors in ruby-red-fin shark (rainbow shark), Epalzeorhynchos frenatum (Teleostei: Cyprinidae). Precursor alarm cells were shown to be small, cuboidal, pyramidal, or round in shape, with eosinophilic cytoplasm, resting on the basement membrane of the epidermis. The cells later elongated to become columnar in shape. Subsequently, they enlarged and became large oval-shaped cells. They then underwent shrinkage and vacuolation. The superficial alarm cells were collapsed. Alarm cells were found to have an affinity for different histochemical stains, including bromophenol blue, iron hematoxylin, Sudan black, Mallory triple trichrome, Crossman's trichrome, Safranin O, and Weigert's stains, as well as lipase and alkaline phosphatase. Endocrine properties of the alarm cells were identified by silver staining and synaptophysin immunostaining. Alarm cells exhibited stemness activities and had a strong immunoaffinity for CD117. They had a proteolytic function, as identified by lysosome-specific staining with acridine orange and strong immunoaffinity for matrix metalloproteinase (MMP-9). They also exhibited proliferatively, reflected by immunological staining by proliferating cell nuclear antigen. In conclusion, alarm cells are unique epidermal cells with multiple functions. They play immunological, and endocrine, roles. They also retain stemness and proliferative properties.

The His bundle is a part of the specialized electrical conduction system that provides a connection between the atrial and ventricular myocardial compartments in both normal and abnormal hearts. The aim of this study was to perform a morphometric analysis of His bundle characteristics of in humans, dogs, horses and pigs and compare them in these studied species. Histological sections of 5 μm thickness were obtained and stained with hematoxylin–eosin and Masson's trichrome; the desmin and periodic acid–Schiff methods were also used for precise identification of cells. The His bundle was found to be longer in horses (2.85 ± 1.02 mm) and pigs (1.77 ± 0.9 mm) than in dogs (1.53 ± 0.8 mm) or humans, in which it was shortest (1.06 ± 0.6 mm). The area and diameters in His bundle cells, were significantly larger in pigs and horses than in humans (p

Background Microsporidia are a large family of obligate intracellular protozoa; these medically important species are recognized as opportunistic agents in intestinal complications in HIV+/AIDS patients. Methods The current cross-sectional study was designed and conducted from October 2018 to June 2019 to determine intestinal microsporidia in HIV+/AIDS patients by trichrome/Zeihl-Neelsen staining and SYBR Green-based real-time PCR. Results Out of 80 HIV+/AIDS patients, 23.75% (n=19) and 12.5% (n=10) were identified by molecular and microscopic methods, respectively. The predominant species in patients was Encephalitozoon (94%), which was found by quantitative real-time PCR and its high resolution melting tool. Conclusion As far as we know, this is the first report from the Alborz region. The prevalence of intestinal microsporidiosis in this area in HIV+/AIDS patients was higher than both the global and national average. In addition to the need for further studies to prove protozoan pathogenicity in the aforementioned group, preventive measures should be considered.

Background: Diabetes Mellitus (DM) is a worldwide common disease and it is considered a serious public health disorder .The reproductive function is highly sensitive to changes in metabolic status, with defective reproductive capacity. Aim of Work: To demonstrate the histological changes in the prostate of rats with STZ induced diabetes type and to examine to what extent insulin could reverse such changes. Materials and methods: Twenty six male albino rats were divided into three groups. Group I: control; Group II: Diabetic was exposed to induction of diabetes by a single intraperitoneal injection of Streptozotocin(STZ) 60 mg/kg in 0.1 M citrate buffer; Group III: Diabetic then received insulin (3IU/day S.C( (for 4 weeks). The rats of each group were sacrificed after 4 weeks. The prostates were processed for electron microscopic sections and other sections stained with H&E, Masson's trichrome, immunohistochemical staining of α smooth muscle actin, Vimentin and PCNA and statistical analysis were applied. Results: Diabetes resulted in deterioration of normal structure of the prostatic acini and some prostatic acini showed cell proliferation with increase Masson's trichrome staining and increase immunoreactivity for α smooth muscle actin, Vimentin and PCNA. Electron microscopic examination reavealed dilatation of the cisternae of the Golgi complex, endoplasmic reticulum, and some epithelial cells showed proliferation and irregular contour of their nuclei. Insulin administration mostly reversed these changes. Conclusion: insulin administration greatly improved the histological architecture of the prostate gland.

Colpodella spp. are free-living flagellates closely related to the apicomplexans. Human infections by Colpodella sp. have been reported. A biflagellated trophozoite and cyst stage comprise the known life cycle stages of Colpodella sp. However, the process of encystation and excystation within the life cycle is unclear. Life cycle stages initiating human infections are unknown. We performed a detailed investigation of the life cycle of Colpodella sp. (ATCC 50594) in culture using Sam-Yellowe's trichrome stains and differential interference contrast (DIC) for light microscopy and fluorescence microscopy of Congo red-stained cells and investigated ultrastructure using transmission electron microscopy (TEM). We report previously undocumented stages of Colpodella sp. Asymmetric and asynchronous division was detected inside cysts by trichrome staining and by TEM. Odd-numbered juveniles and cysts containing more than four juvenile trophozoites were identified. Live imaging of active cultures captured the excystation and egress of juvenile trophozoites and confirmed the presence of multinucleate cysts. The ultrastructure of the multinucleate cyst is reminiscent of apicomplexan schizonts. Insights gained from the life cycle stages observed in culture allowed the construction of the life cycle of Colpodella sp. Knowledge of the life cycle will aid biochemical and molecular characterization of Colpodella sp. and help identify stages in human infections.

Background Intimal calcification typically develops in advanced atherosclerosis, and microcalcification may promote plaque progression and instability. Conversely, intraplaque hemorrhage and erythrocyte extravasation may stimulate osteoblastic differentiation and intralesional calcium phosphate deposition. The presence of erythrocytes and their main cellular components (membranes, hemoglobin, and iron) and colocalization with calcification has never been systematically studied. Methods and Results We examined three types of diseased vascular tissue specimens, namely, degenerative aortic valve stenosis (n = 46), atherosclerotic carotid artery plaques (n = 9), and abdominal aortic aneurysms (n = 14). Biomaterial was obtained from symptomatic patients undergoing elective aortic valve replacement, carotid artery endatherectomy, or aortic aneurysm repair, respectively. Serial sections were stained using Masson–Goldner trichrome, Alizarin red S, and Perl's iron stain to visualize erythrocytes, extracelluar matrix and osteoid, calcium phosphate deposition, or the presence of iron and hemosiderin, respectively. Immunohistochemistry was employed to detect erythrocyte membranes (CD235a), hemoglobin or the hemoglobin scavenger receptor (CD163), endothelial cells (CD31), myofibroblasts (SMA), mesenchymal cells (osteopontin), or osteoblasts (periostin). Our analyses revealed a varying degree of intraplaque hemorrhage and that the majority of extravasated erythrocytes were lysed. Osteoid and calcifications also were frequently present, and erythrocyte membranes were significantly more prevalent in areas with calcification. Areas with extravasated erythrocytes frequently contained CD163-positive cells, although calcification also occurred in areas without CD163 immunosignals. Conclusion Our findings underline the presence of extravasated erythrocytes and their membranes in different types of vascular lesions, and their association with areas of calcification suggests an active role of erythrocytes in vascular disease processes.

The olfactory organs of two African lungfishes,Protopterus amphibiusandP. dolloi, were investigated using a stereo microscope and a compound light microscope and were described anatomically, histologically, and histochemically. Like other lungfishes, these species present the following general features: i) elongated olfactory chamber (OC), ii) anterior nostril at the ventral tip of the upper lip, iii) posterior nostril on the palate of the oral cavity, iv) lamellae with multiple cell types such as olfactory receptor neurons, supporting cells, basal cells, lymphatic cells, and mucous cells (MC), and vi) vomero-like epithelial crypt (VEC) made of glandular epithelium (GE) and crypt sensory epithelium. Some of these features exhibit differences between species: MCs are abundant in both the lamellar and inner walls of the OC inP. amphibiusbut occur only in lamellae inP. dolloi. On the other hand, some between feature differences are consistent across species: the GE of bothP. amphibiusandP. dolloiis strongly positive for Alcian blue (pH 2.5)-periodic acid Schiff (deep violet coloration), and positive with hematoxylin and eosin and with Masson’s trichrome (reddish-brown staining), unlike the MCs of the two species which stain dark red with both Alcian blue (pH 2.5)-periodic acid Schiff and Masson’s trichrome but respond faintly to hematoxylin and eosin. The differing abundance of MCs in the two lungfishes might reflect different degrees in aerial exposure of the olfactory organ, while the neutral and acid mucopolysaccharide-containing VEC, as indicated by staining properties of the MCs, is evolutionary evidence thatP. amphibiusandP. dolloiare the closest living relatives to tetrapods, at least in the order Dipnoi.

Venous invasion (VI) is a powerful prognostic factor in colorectal cancer (CRC) that is widely underreported. The ability of elastin stains to improve VI detection is now recognized in several international CRC pathology protocols. However, concerns related to the cost and time required to perform and evaluate these stains in addition to routine hematoxylin and eosin (HE) stains remains a barrier to their wider use. We therefore sought to determine whether an elastin trichrome (ET) stain could be used as a "stand-alone" stain in CRC resections, by comparing the sensitivity, accuracy, and reproducibility of detection of CAP-mandated prognostic factors using ET and HE stains. Representative HE- and ET-stained slides from 50 CRC resections, including a representative mix of stages and prognostic factors, were used to generate 2 study sets. Each case was represented by HE slides in 1 study set and by corresponding ET slides from the same blocks in the other study set. Ten observers (3 academic gastrointestinal [GI] pathologists, 4 community pathologists, 3 fellows) evaluated each study set for CAP-mandated prognostic factors. ET outperformed HE in the assessment of VI with respect to detection rates (50% vs. 28.6%; P0.0001), accuracy (82% vs. 59%, P0.0001), and reproducibility (k=0.554 vs. 0.394). No significant differences between ET and HE were observed for other features evaluated. In a poststudy survey, most observers considered the ease and speed of assessment at least equivalent for ET and HE for most prognostic factors, and felt that ET would be feasible as a stand-alone stain in practice. If validated by others, our findings support the use of ET, rather than HE, as the primary stain for the evaluation of CRC resections.

Objective: To evaluate the biological effects of coral scaffold with or without adipose derived stem cells (ADSCs) on healing of bony defects in albino rats. Methods: Fifty four male albino rats weighing approximately 250-300 gm were selected and randomly divided into 3 groups. Each rat received one bone defect on the left side of mandible near the diastema. The defects were either left empty (group A) n=18, treated with coral only (group B) n=18 or received a combination of coral and adipose derived stem cells (group C) n=18 which were harvested from the rats peri renal region. Bone healing was evaluated using heamatoxylin and eosin and trichrome stain, then the results were subjected to digital image analysis followed by one- way ANOVA statistical analysis. Results: Bony defects received the combination of coral scaffold and stem cells show better healing in respect to the quantity and the quality of the newly formed bone. Conclusion: ADSCs loaded onto coral granule scaffold have ameliorating effect on the repair of the mandibular defects in rat model.

The evaluation of the morphology and organization of collagen fibers is critical in understanding wound healing and tissue remodeling after a thermal injury of the skin. However, histological analysis conducted by pathologists is often labor-intensive and limited to qualitative evaluations and scoring within a narrow field of view. In this study, we propose a convolutional neural network (CNN) model to classify Masson’s trichrome (MT)-stained histology images of burn-induced scar tissue and to characterize the microstructures of normal tissue and scar tissue in a quantitative manner. The scar tissue is created on in vivo rodent models and prepared for MT-stained histology slides after wound healing. A CNN model is developed, trained, and tested with various sizes of the histology images for classification and characterization. The proposed model classifies both normal tissue (i.e., without burn, as the control) and scar tissue at various scales with over 97% accuracy. The features acquired from the proposed CNN model visually characterizes the density and directional variance of the collagen fibers distributed in the dermal layers from whole histology images. The proposed deep learning technique can provide an objective and reliable method to rapidly assess and quantify wound repair and remodeling.

Background: Periodontitis affects approximately 20%–50% of the global population and is caused by gram-negative bacteria, such as Porphyromonas gingivalis (P. gingivalis). Host modulation therapy (HMT) is part of a periodontal therapy that is used as an adjunct to conventional periodontal treatment to reduce tissue damage. Lemuru fish oil containing EPA and DHA can reduce the formation of MMPs and will further increase the number of fibroblasts there by stimulating collagen formation. Purpose: To determine the effect of lemuru fish oil gel on the collagen density and width of the periodontal tissue induced by P. gingivalis and the correlation between these parameters. Methods: Thirty male Wistar rats were divided into five groups. Induction of P. gingivalis was carried out first, then lemuru fish oil gel was applied to the gingival sulcus for 14 days, according to collagen scores in histological preparations using Masson's trichrome (MT). The width of the periodontal ligament was measured with an image raster program in µm. The d ata were analysed using statistics to test hypotheses using SPSS version 24. Results: Significant differences in the results of the c ollagen density were observed between groups K- and K+ and groups K+ and P2. Meanwhile, no significant difference was observed between groups K- and P2, P3, P2 and P3 and K+ and P1 . The mean values of the periodontal ligament widths were K- (299.61 ± 51.82µm), K+ (425.85 ± 61.54µm), P1 (346.93 ± 33.53µm), P2 (370.15 ± 49.42µm) and P3 (379.6 ± 49.26). Conclusion: Lemuru fish oil can affect the width of the ligament and the collagen density with an optimal concentration of 20%. The correlation between the collagen density and the periodontal ligament width was negative and not significant.

This study assessed intestinal protozoa with stress on Blastocystis among patients with CKD andthose on hemodialysis (HD) in Nephrology Unit, Medical Research Institute, Alexandria.The patients were divided into three groups. GI: 50 patients on HD for at least one year, GII: 50 CKDpatients not on HD, and GIII: 50 healthy controls. Fresh stool samples were collected in labeled plasticboxes and examined as direct smears after staining with trichrome, Modified Ziehl-Neelsen (MZN)stains and modified trichrome (MT) stains. Besides, stool samples were cultured for Blastocystis hominisand Copro-antigen (ICT) was used to detect E. histolytica /dispar, G. lamblia and C. parvum.The results showed that the parasitic infections were detected in 79% of the patients; 40% in HD and39% in CKD patients. The positive samples were (47%) detected by direct wet mount followed byiodine smear (43%) and then trichrome stain (42%). Microsporidiosis was detected in 55 (55%) HDand CKD patients using the MT stain. ICT agreed with microscopic examination in diagnosis ofE.histolytica/ dispar and G. lamblia and showed perfect agreement with MZN in C. parvum diagnosis.

The aim of this study was evaluating the effects of vacuum on microstructure and biocompatibility of bovine decellularized pericardium. So the bovine pericardia were decellularized and then the vacuum was applied for two periods of time; 90 and 180 min. DNA, glucose amino glycan, collagen and elastin content assay, scanning electron microscopy (SEM) examination, hematoxylin and eosin (H&E) and Masson's trichrome stainings performed to evaluate microstructure of tissues. MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) test, subcutaneous implantation, and tensile test were used to assay biocompatibility and mechanical properties of decellularized tissues. The results showed that applying vacuum reduced residual DNA significantly. Vacuum after 180 min reduced more residual DNA. There were no significant differences in the content of glucose amino glycan (GAG), collagen, and elastin between the vacuumed and control groups. SEM examination was revealed that vacuum for 180 min increased pore size and porosity more than 90 min and control groups. H&E and Masson's trichrome stainings revealed extracellular matrix preservation after decellularization in all groups. Cell viability was increased in vacuumed samples significantly after 72 h in vaccumed samples. H&E staining and tensile test after implantation of tissues were showed less inflammation in the vacuum applied tissues and increased durability. The vacuum increased DNA removal, pore size, porosity, and biocompatibility in vitro and in vivo and durability of bovine decellularized pericardium in vivo. Considering the important role of time, more studies should be performed to optimize time, intensity, and method of application of vacuum in decellularization of different tissues as well as bovine pericardium.

This study was conducted to evaluate the bone regeneration effects of collagen/strontium-doped Bioglass (BG-Sr) scaffolds. The present study comprised four parts including designing and fabricating scaffolds, characterization tests such as biodegradation, water uptake, porosity, FTIR (Fourier transform infrared spectroscopy), and SEM (Scanning electron microscope), in vitro tests such as MTT ((3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide)), alkaline phosphatase, calcium assay, Alizarin Red staining, and real-time PCR (Polymerase chain reaction), and ultimately in vivo and histopathological tests such as evaluating bone regeneration, mineralization with Hematoxylin-Eosin, Masson's trichrome and Alizarin Red staining, and histomorphometric studies 4, 8 and 12 weeks after implanting the scaffolds in the skull and femur of 36 rabbits. Significant reductions were observed in degradability, water absorption, and porosity in the BG-Sr samples. In vitro results showed that the synthetic scaffolds were less toxic, stimulated, and induced cell growth. The ALP (Alkaline phosphatase) activity was higher on day 14 than on day 7, and the highest calcium content was observed in the BG-Sr samples on day 14. In line with the SEM and MTT results, the DAPI (Diamidino-2-phenylindole) results confirmed cell proliferation. Moreover, osteocalcin and ALP gene expression were high in the BG-Sr group. The results of the histopathological tests showed high levels of bone formation in the BG-Sr scaffolds. As a scaffold in regenerative medicine, the BG-Sr composite can be an effective product for bone regeneration.

Purpose: The aim of this study was to elucidate whether ovarian tissue is able to withstand a double freezing-thawing procedure. Methods: Human ovarian cortical biopsies from 4 thawed whole ovaries were divided into 4 experimental subgroups: (a) frozen-thawed non-grafted group, (b) frozen-thawed xenografted group, (c) refrozen-rethawed non-grafted group, and (d) refrozen-rethawed xenografted group. Xenografting was performed using 8 severe combined immunodeficient mice for a total duration of 21 days. The following analyses were conducted: classic hematoxylin and eosin staining, Ki67 immunolabeling, transmission electron microscopy, Masson's green trichrome, and double CD34 immunostaining. Results: Morphologically normal preantral follicles were detected in all groups. We observed a dramatic decline of more than 65% in early preantral follicle survival rates after grafting of both frozen-thawed (p < 0.0001) and refrozen-rethawed (p < 0.0001) ovarian tissue. However, mean follicle densities remained comparable between the frozen-thawed and refrozen-rethawed non-grafted groups, as well as both grafted groups. Equivalent proportions of proliferating early preantral follicles were identified in frozen-thawed and refrozen-rethawed samples, whether the tissue was grafted or not. Furthermore, we did not observe any significant difference in atretic follicle rates between any of the four groups, and the ultrastructural quality of follicles appeared unaffected by the refreezing procedure. Similar proportions of fibrosis were noted in the frozen-thawed and refrozen-rethawed groups, irrespective of grafting. Finally, no significant differences were witnessed in terms of vascularization. Conclusion: We were able to demonstrate, for the first time, that refrozen-rethawed ovarian tissue has the same functional characteristics as frozen-thawed ovarian tissue.

Context: Oral Mucositis (OM) is a very painful inflammatory condition, complicating patient’s life. Hence it was appealing to find natural remedies, dietary modifications or both to manage this condition successfully. Aim: to evaluate the efficiency of Propolis (P) and / or Intermittent Fasting (IF) in ameliorating OM. Materials & Methods: 40 male Albino rats weighing 200-250 grams, divided into: Negative Control Group (-ve control Gp): 8 rats kept healthy, Positive Control Group (+ve): 8 rats subjected to OM induction in their buccal mucosa, (P) Gp: 8 rats subjected to OM Induction, given Propolis orally, (IF) Gp: 8 rats subjected to OM induction & IF, (P&IF) Gp: 8 rats subjected to OM induction, maintained on both P &IF simultaneously. After three weeks, buccal mucosae were processed and stained by H&E, Masson’s Trichrome, and Anti-PCNA, examined by light microscope. Results: +ve control Gp showed significant increase in vacuolated epithelial cells, apparent decrease in immature collagen fibers in CT, and significant reduction in PCNA-positive cells. In the P, IF, and P&IF Gps, there were marked improvement in epithelium and CT. The (P&IF) Gp manifested maximum decrease in vacuolated epithelial cells, increase of immature collagen fibers, and maximum increase in PCNA positive cells. Discussion & conclusion: OM caused tissue destruction, which was evident in the +ve control Gp. Propolis intake greatly improved the histological appearance (confirming anti-inflammatory role of Propolis) as well as IF (confirming immune-boosting action). However, using both P & IF resulted in maximum improvement due to combined actions of both.

Two young opossums were necropsied and diagnosed with rickets. This study aims to describe the clinical-pathological aspects of rickets in Didelphis albiventris. Macroscopically, the opossums presented kyphosis and scoliosis, lateral deviation of the limbs in varus, locomotion difficulty, and enlargement with softening of costochondral junctions (rickety rosary). Samples of bones and joints were processed for hematoxylin and eosin staining and Masson’s trichrome. Microscopically, we observed thickening of the epiphyseal plate, characterized by irregular and multifocal proliferation of serialized and hypertrophic cartilage zones, which formed circular groups of large, dysplastic chondrocytes towards the spongy zone, often surrounded by non-mineralized osteoid tissue. In the cortical bone, there were pale eosinophilic zones around the Havers channels consistent with non-mineralized osteoid. The staining of Masson’s trichrome evidenced the accumulation of osteoid tissue in cortical and trabecular bones. It is possible that a mixed cause of absorption deficiency of vitamin D3 associated with an unbalanced Ca:P diet based on lactose-free milk and fruits may have triggered the disease. RESUMO: Dois gambás jovens foram necropsiados e diagnosticados com raquitismo. O objetivo do trabalho é descrever os aspectos clínico-patológicos de raquitismo em Didelphis albiventris. Macroscopicamente os gambás apresentaram cifose e escoliose, desvio lateral dos membros em varus, dificuldade de locomoção e alargamento com amolecimento das junções costocondrais (rosário raquítico). Amostras dos ossos e articulações foram processadas para coloração de hematoxilina e eosina e Tricrômico de Masson. Microscopicamente havia espessamento da placa epifisária, caracterizada pela proliferação irregular e multifocal das zonas de cartilagem seriada e hipertrófica, que formavam grupos circulares de condrócitos grandes, displásicos em direção a zona esponjosa frequentemente cercados por tecido osteoide não mineralizado. No osso cortical haviam zonas eosinofílicas pálidas ao redor dos canais de Havers consistentes com osteoide não mineralizado. A coloração de Tricrômico de Masson evidenciou o acúmulo de tecido osteoide no nosso cortical e trabecular. Acredita-se que uma causa mista de déficit de absorção de vitamina D3 associada a uma dieta desbalanceada em Ca:P a base de leite sem lactose e frutas tenha desencadeado a doença.

Background: Microsporidia are a group of obligate intracellular organisms that can infect all animals including man causing disease in both immunocompetent individuals and immunosuppressed patients. The most common microsporidia causing gastrointestinal infection worldwide are Enterocytozoonbieneusi and Encephalitozoonintestinalis. Objectives: The present study was designed with the aim of estimating the rate of microsporidia in diarrheic children and the evaluation of different staining techniques and nested polymerase chain reaction (PCR) for the clinical diagnosis of intestinal microsporidiosis. Subjects and Methods: One hundred and fifty diarrheic stool samples were collected from Abou El-Rich children hospital laboratory. Samples were examined by “Ryan-Blue” modified trichrome stain (MTS), modified ZiehlNeelsen (MZN), acid fast trichrome (AFT) stain, and nested polymerase chain reaction (nPCR). Results:Microsporidium spp. was detected in 12 (8%) of samples using MTS. Compared with the other detection techniques the respective sensitivities and specificities of MZN stain was 100% and 98.57%; nPCR was 80% and 100%; and AFT was 85.71% and 97.87%. Conclusion: Using specific staining techniques as MZN and AFT for diagnosis of intestinal microsporidiosis in diarrheal stool samples are nearly as efficacious as PCR, but even better in some cases with the added advantage of being an inexpensive diagnostic method compared to PCR. Another benefit is the detection of other oocyst forming parasites such as Cryptosporidium, which makes staining techniques very suitable for developing countries

Aim. This experimental work presents a comparative assessment of the effect of hyaluronic acid and crystalline calcium hydroxyapatite on derma in separate and combined application setting.Materials and methods. The study used rats (30 animals) for subdermal injection of 0.05 ml medium. Group 1 was administered calcium hydroxyapatite (Radiesse (Merz, Germany)), group 2 received hyaluronic acid (Restylane (SabQ, Sweden)) and Group 3 — both preparations combined in ratio 1:1 (Mix). Results were evaluated 4 months after the filler injection. Sections were stained with haematoxylin–eosin, van Gieson’s and Masson’s trichrome techniques. Collagen types I and III were detected with polyclonal antibodies (Abcam, England). Fibroblasts were positively identified with vimentin (LabVision), macrophages — with CD68 (LabVision) tagging. Effect of extracellular matrix remodelling was studied with α-SMA actin (Abcam, England).Results. We demonstrate that separate filling of hyaluronic acid and calcium hydroxyapatite produces different response mechanisms, while their combined administration does not lead to a marked voluming of the dermal extracellular matrix.Conclusion. Combined administration of the two fillers under study may facilitate a prolonged effect of their combined action exceeding 4 months, due to the absence of fibrosis, complications and side effects.

The goldfish is a model organism showing great potential for research, particularly in comparative endocrinology concerning the neuroendocrine signalling and regulation of vertebrate reproduction. Furthermore, this teleost is increasingly stressed as a relevant alternative to more common fish model organisms, namely zebrafish. However, quality descriptions and illustrations of the complete goldfish gonadal histology are surprisingly scarce, but needed, to support research using this fish. Therefore, the main aim of this work is to describe in detail and adequately illustrate the goldfish oogenesis, from oogonia to late maturation, by applying routine stains (haematoxylin-eosin) and special procedures (periodic acid-Schiff and Goldner's trichrome). We hypothesized that the combined strategies would enable not only to observe the most general features but also to perceive some poorly described details of oocytes better. We describe the details of the following maturation stages: oogonia proliferation, chromatin-nucleolus, primary growth (one nucleolus step, multiple nucleoli step, perinucleolar step, cortical alveoli step) and secondary growth (early secondary growth step, late secondary growth step). Additionally, we report aspects of early and late follicular atresia. The study allowed comparisons with other species and showed that the Goldner's trichrome has the best discriminative power and should be the preferred stain, despite more time-consuming.

To date, we could find no study concerning the relationship between mechanoreceptors in the joint capsule of the first metatarsophalangeal joint and hallux valgus deformity. We aimed to investigate the presence of mechanoreceptors in samples obtained from the first metatarsophalangeal joint capsules of patients with hallux valgus deformity to improve our understanding of the clinical and histopathological features of the disease. Samples were taken from the first metatarsophalangeal joint capsules of 13 fresh-frozen cadavers with normal anatomy (controls) and 29 patients undergoing surgery for hallux valgus (cases). For light microscopy, excised specimens were fixed in 10% formaldehyde and processed for routine histopathological investigation. All samples were dehydrated in a series of ethanol, cleared in xylene, and embedded in paraffin. Orientation of collagen fibers was determined on Masson's trichrome-stained sections, and mechanoreceptors were evaluated on S-100-immunostained sections. In the sections stained with Masson's trichrome, the orientation of collagen fibers was regular in the control group. However, coarse and disoriented collagen bundles were observed in the hallux valgus cases (P ≤ .05). S-100 immunostaining was positive in the sections of both the cases and controls. Finally, free nerve endings were more abundant in the samples obtained from the capsules of hallux valgus cases than from the control group (P ≤ .05). An increase in the number of free nerve endings within the capsules of the first metatarsophalangeal joints in feet with hallux valgus deformity might have a role in the development of clinically relevant joint pain and instability.

Background: Varicose veins (VVs) are a major chronic venous disease, yet the underlying mechanisms are unclear. This disorder is characterized by progressive architectural remodeling of the vessel wall. Recent studies have focused on the role of matrix metalloproteinases (MMPs) in the pathogenesis of VVs. Aim of Study: The present study was assigned to demon-strate the morphologic and morphometric changes in the wall of VVs and to correlate these changes with the expression of MMP-1 and MMP-9. Material and Methods: A total number of 18 human long saphenous vein samples (8 normal and 10 VVs) were included in the study. The specimens were processed and examined histologically using hematoxylin and eosin, Masson's trichrome and orcein, together with immunohistochemistry for alpha smooth muscle actin (Eo-SMA). The mRNA expression of MMP-1 and MMP-9 was analyzed using real-time polymerase chain reaction (PCR). Quantitative morphometric measure-ments were carried out by the image analyzer. Results: The wall of VVs showed damaged endothelial surface, interrupted internal elastic lamina and numerous vasa vasora. There was significant increase in intima and media thickness compared to normal veins. Significant increase in the mean collagen area percent and decrease in elastin area percent was also observed. The mean area percent of Eo-SMA immunostaining and count of smooth muscle cells (SMCs) showed significant increase, indicating SMCs hypertrophy and hyperplasia, while the optical density of Eo-SMA immu-nostaining decreased. These changes were associated with significant increase in mRNA expression of MMP-1 and MMP-9. Conclusion: Varicose veins showed marked disorganiza-tion in all components of the vessel wall (collagen, elastin and SMCs). The associated increase in mRNA expression of MMPs might be one of the molecular mechanisms underlying vascular remodeling.

Here, we reported a spontaneous case of membranoproliferative glomerulonephritis observed in a young ICR mouse. A 5-week-old female mouse was euthanized owing to abdominal swelling and increased body weight. At necropsy, generalized subcutaneous edema, and clear, colorless, non-viscous ascites were observed. Histologically, the kidneys showed diffuse, bilateral glomerular lesions. The lesions were characterized by thickening and double contour of the basement membrane and an increase in mesangial cells and matrix, resulting in the narrowing of the capillary lumen. Additionally, eosinophilic hyaloid material accumulated in the subendothelial areas and Bowman's space. The material was positive for periodic acid-Schiff, complement component C3, or immunoglobulin G, stained red by Masson's trichrome, and stained blue by phosphotungstic acid-hematoxylin stain and was considered to be plasma due to glomerular leakage. The glomerular lesion was diagnosed as membranoproliferative glomerulonephritis, and an uncertain endothelial injury was suspected as the cause.

We analyzed the peripheral blood lymphocyte subsets of cancer patients infected with intestinal parasites, with an aim to find out the relationship between the levels of different types of lymphocytes with the prognosis of patients. 201 cancer patients aged 18 and over were included. Stool samples of the patients were examined using native-lugol, trichrome, modified trichrome (Weber's Trichrome stain), and modified Ziehl-Neelsen staining methods. Microsporidia and Cryptosporidium parvum were investigated at the genus and species levels using PCR. Lymphocyte subsets were determined by flow cytometry in blood samples. One or more parasite species were detected in 115 (56.7%) patients. The most common parasite species were Microsporidia, Blastocystis and Entamoeba coli, respectively. The frequency of parasites was high in patients with low lymphocyte percentage, CD3(+) T cell and CD3(+) CD4(+) T (Th) cell levels in blood samples studied by flow cytometry. Microsporidia infection was significantly higher in patients with low lymphocyte percentage and Th cell levels. Similarly, C. parvum infection was found to be significantly higher in patients with low T lymphocyte percentage and Th cell level. Finally, Blastocystis infection was significantly higher in patients with low lymphocyte percentage and CD4/CD8 ratio higher than 1. The decrease in lymphocyte percentage, CD3(+) T cell and Th cell count, and low CD4/CD8 ratio in cancer patients increase the frequency of intestinal parasitic infections. Based on these results, lymphocyte subsets may help identify cancer patients at high risk of opportunistic parasites. We suggest that opportunistic parasitic infections affecting the clinical course of the disease should be considered by clinicians during the follow-up and treatment of patients.

Decellularization is one of the promising techniques in tissue engineering used to create a biological scaffold for subsequent repopulation with the patient’s own cells. This study aims to compare two different decellularization protocols to optimize the process of auricle decellularization by assessing and characterizing the decellularization effects on human auricular cartilage. Herein, 12 pairs (8 females, 4 males) of freshly frozen adult human cadaveric auricles were de-epithelialized and defatted leaving only the cartilaginous framework. An auricle from each pair was randomly assigned to either protocol A (latrunculin B-based decellularization) or protocol B (trypsin-based decellularization). Gross examination of the generated scaffolds demonstrated preservation of the auricles’ contours and a change in color from pinkish-white to yellowish-white. Hematoxylin and eosin staining demonstrated empty cartilaginous lacunae in both study groups, which confirms the depletion of cells. However, there was greater preservation of the extracellular matrix in auricles decellularized with protocol A as compared to protocol B. Comparing protocol A to protocol B, Masson’s trichrome and Safranin-O stains also demonstrated noticeable preservation of collagen and proteoglycans, respectively. Additionally, scanning electron micrographs demonstrated preservation of the cartilaginous microtopography in both study groups. Biomechanical testing demonstrated a substantial decrease in Young’s modulus after decellularization using protocol B (1.3 MPa), albeit not significant (P-value > 0.05) when compared to Young’s modulus prior to decellularization (2.6 MPa) or after decellularization with protocol A (2.7 MPa). A DNA quantification assay demonstrated a significant drop (P-value < 0.05) in the DNA content after decellularization with protocol A (111.0 ng/mg) and protocol B (127.6 ng/mg) in comparison to before decellularization (865.3 ng/mg). Overall, this study demonstrated effective decellularization of human auricular cartilage, and it is concluded that protocol A provided greater preservation of the extracellular matrix and biomechanical characteristics. These findings warrant proceeding with the assessment of inflammation and cell migration in a decellularized scaffold using an animal model.

O tecoma é uma neoplasia ovariana rara, sendo, portanto, escassas suas descrições na medicina veterinária. Uma de suas formas de apresentação é o tumor estromal esclerosante, representando cerca de 2-6% dos tumores estromais com origem no ovário. Este subtipo de tecoma ainda não foi descrito em cadelas. Este trabalho objetiva relatar a ocorrência de um tecoma de grandes dimensões, classificado como tumor estromal esclerosante. A cadela era da raça labrador, 10 anos de idade, e o diagnóstico foi estabelecido após exame necroscópico. O histórico clínico constava aumento de volume abdominal, diminuição da evacuação e presença de coágulos de sangue na urina. A avaliação histopatológica da massa demonstrou um agregado de células fusiformes, por vezes ovais, dispostas em feixes, com núcleo central, citoplasma claro de aspecto vacuolizado, indicando a presença de lipídios. O diagnóstico foi confirmado pela técnica histoquímica do Tricromio de Masson, permitindo observar a presença de fibras colágenas dispostas entre as células, concluindo o diagnóstico de tecoma do subtipo estromal esclerosante.

Colpodella species are free-living close relatives of apicomplexans that were recently reported to cause red blood cell infection in an immunocompromised human host and in a tick-borne human infection resulting in neurological symptoms. Unambiguous identification of the life cycle stages of Colpodella sp. using routine stains for light microscopy will aid rapid diagnosis in infections. Similarly, cells in culture and environmental samples can be rapidly identified by staining. Staining protocols are currently unavailable for cell detection by light microscopy. In this study, we investigated the feasibility of performing routine staining techniques for light microscopy for differentiating Colpodella sp. (ATCC 50594) and Bodo caudatus cysts in Hay medium cultures. We tested different basic and acidic dyes alone and in combination and also utilized a commercial trichrome staining protocol. The nonspecific fluorescent dye Calcofluor white was also evaluated. Staining times, dye concentrations, use of tap or distilled water rinses, use of a mordant and inclusion, or omission of decolorizers after staining were evaluated. We compared the intensity of color, clarity of morphological features, and cytoplasmic structures detected after staining. We report a new trichrome staining technique that allowed clear identification and differentiation of cyst stages of Colpodella sp. and B. caudatus. Immature Colpodella sp. cysts were identified as having an irregular, dual-colored (demilune), dark blue-purple and white appearance. Mature Colpodella sp. cysts stained dark red-blue and were identified in four-way mitotic division, while cysts of B. caudatus in diprotist or monoprotist (ATCC 30905) cultures were detected as spherical and red-pink in appearance.

The expressions of matrix metalloproteinases 2 and 9 have been described during the development, as an example in heart and tooth but not in the small intestine yet. In this context, this study aimed to evaluate the expressions of MMP-2 and MMP-9 in the small intestine of Wistar rats during intrauterine (IU) and postnatal (PN) life. Expressions were determined on the 15th and 18th days of IU life and the 3rd, 10th, 17th, 25th, and 32nd days of PN life. Intestinal samples obtained from six animals were submitted to zymography, immunohistochemistry, and staining with Masson's trichrome. The results showed that MMP-2 and MMP-9 were not expressed during IU life; however, after birth, MMP-9 was immunolocalized in the goblet and mast cells. In conclusion, our results showed that MMP-2 and MMP-9 were not expressed in absorptive epithelial cells during the IU period of the small intestine but after birth, MMP-9 was expressed in the goblet cells, and mast cells present in the lamina propria.

The aim of the current investigation was to identify the cellular changes of the cell populations positioned within the connective tissue of the harderian glands in Turkeys at different ages extended from one day hatched offspring till the age of seven months. To conduct such research, 50 specimens of harderian glands from the above-dated ages of turkey were collected. Post euthanasia and dissection of the birds, specimens were cut and fixed in 10% neutral buffered formalin and Bouin’s solution. The subsequent routine histological technique was made on these specimens such as dehydration, clearing, paraffin embedding, block preparation, and sectioning. Tissue sections were obtained and stained with different stains such as hematoxylin-eosin, Masson’s trichrome, periodic acid-Schiff (PAS) and Alcian blue (AB). Microscopic findings showed connective tissue of the harderian gland was populated by three types of cells that were heterophils, lymphocytes and plasma cells. The incidence of each of these cells was changed with advance of the age. During the growth of the birds, a gradual shift was taking place in the domination of these three cell types. The lymphoid nodules were detected only in 40 days old poults, whilst in adult birds the harderian glands were contained the largest number of mature plasma cells. Some plasma cells enclosed distinctly Russell bodies with different sizes and shapes. In conclusion, 4 weeks aged turkeys developed lymphoid tissue in the harderian gland which is a critical age for vaccination in this avian species.

The present study was conducted to study the endocrine portion of the pancreas represented by Islets of Langerhans in the adult males and females guinea pigs. Eight animals of each sex were collected to conduct these aims. Histological sections prepared from the specimens which were collected from three identified lobes of the pancreas and subsequently fixed by 10% neutral buffered formalin. Post conducting the routine histological procedure on specimens, three stains were used such as hematoxylin and eosin, Masson’s trichrome and Gomori's Aldehyde trichrome for identification of cellular composition of the pancreatic Islet's of Langerhans. The endocrine islets showed predominantly s, and for a lesser extent α cells. The δ cells were rarely detected. The percentage of s cells in the islets of male pancreas was higher than those in females. On contrary, the percentage of α cells was higher in the islets of females than those in the males. Such findings were proportional to hormones analysis of insulin and glucagon which were corresponding products to these cells. In conclusion, Percentage of s cells in the islets of male pancreas was higher than that in females. On the contrary, the percentage of α cells was higher in the islets of females than that in the males. Such observations were parallel positively with hormones analysis of insulin and glucagon which were corresponding products to these cells.

Potentially pathogenic free-living amoebae (FLA) are unique protozoa that present a healthcare challenge. The appropriate choice of staining technique is crucial to improving the efficiency of microscopic diagnosis of FLA isolated from environmental as well as clinical samples. This study aimed to evaluate different staining techniques for morphologicalidentification of Acanthamoeba and Naegleria isolated from tap water samples in different localities in Assiut City, Egypt. A multi-attribute evaluation was performed to compare these techniques considering staining quality, ease of performance, cost and time of each procedure. Seven staining techniques were used including Lugol’s iodine and lactophenol cotton blue as temporary stains, with Giemsa, trichrome, iron hematoxylin, acid-fast and Gram stains as permanent stains. Both lactophenol cotton blue and Lugol's iodine showed good structural details of different stages of FLA and good contrast at a very short time and reasonable cost, followed by Giemsa. Both trichrome and iron hematoxylin stains gave comparable staining quality but at a longer time with more complex staining procedures and higher cost. Both modified acid-fast and Gram stains didn't provide sufficient staining quality to identify morphological details of FLA. According to the multi-attribute evaluation, both Lugol’s iodine and lactophenol cotton blue achieved the highest rank (92.85%) followed by Giemsa and Gram stains (64.28 %), modified acid-fast stain (57.14%), while iron hematoxylin and Masson’strichrome had the last rank (46.42%).

Acanthamoeba genus is a medically important free living amoeba causing serious humans infections. Amoebic keratitis (AK) is a sight threatening infection of cornea caused by Acanthamoeba pathogenic genotypes, which prevalence remarkably increased in developed countries. The study compared different methods for diagnosing AK and identified Acanthamoeba genotypes by molecular examination in contact lens wearers (CLWs). Patients were 79 clinical corneal swaps (CS) and 15 samples from contact lens storage cases (CLSC). Clinical CSs were divided into four groups; GI: 20 patients suffering from chronic corneal ulcers, GII: 15 patients with traumatic ulcers, GIII: 24symptomatic CLWs and GIV: 20asymptomatic control individuals. CLSC were provided from apparently healthy asymptomatic CLWs (15). Swabs and solution samples were underwent microscopic and staining examination, cultivation on non-nutrient agar (NNA) plates andPCR molecular analysis. Sequencing and genotyping of PCR- positive samples were performed. The results showed that Acanthamoeba parasites were detected in 3.8% of CS and 6.7% of CLSC samples. The highest significantly positive results were by culture (3.8%) followed by Giemsa and trichrome stains (2.5%) and lastly direct microscopy (1.3%) of CS samples. Only one positive sample (6.7%) was detected in CLSC by all methods, but without statistical significance.Sensitivity of PCR compared to culture was 25%. Acanthamoeba parasites in CS were from subgroup II with 12.5% detection rate in CLWs, but the positive case from CLSC was from subgroup I with 6.7% detection rate. This study confirmed different risk factors in association with AK in CLWs. Genotype determination for Acanthamoeba positive case by PCR revealed homology with Acanthamoeba genotype T9 isolate ICS20.

Background: curcumin has many pharmacological effects including antioxidant, anti-inflammation, eliminating free radicals, anti-tumor, lipid regulation and anti-coagulation. Objectives: to compare the histological effects of curcumin and atorvastatin in experimental induced atherosclerosis in the rabbit’s aorta and heart. Subjects and Methodology: thoracic aorta sections were prepared and stained by Hx&E, orcein, Masson's trichrome and immuonohistochemical staning using CD34. Cardiac sections were prepared and stained by Hx&E, Masson's trichrome stain and immuonohistochemical staning using troponin T. Morphometrical study included, the thickness of both tunica intima and media of thoracic aorta , area % of the elastic fibers contents of both tunica intima and media in thorathic aorta, and the area %of the immune reaction of CD34 in thoracic aorta . Results: the atherosclerotic group showed a significant increase in the thickness of both tunicae intima& media when compared with the control group. There was a significant decrease in elastic fibers content and significant increase in the mean area % of the immune reaction of CD34 versus control. In atorvastatin treated group: there was a significant increase in comparison to control group, while there was a significant decrease in comparison to atherosclerotic group. There was significant decrease in elastic fibers content and significant increase in the mean area % of the immune reaction of CD34 in comparison to control group. In curcumin treated group: the thickness of both tunica intima and media showed a significant increase in comparison to control group. Conclusion: curcumin has potential role in treatment of atherosclerosis and myocardial infarction.

Objective: To compare the effects of curcumin and nanocurcumin oral supplementation on the muscle healing rate of an animal model of surgical muscle laceration. Methods: Thirty-two male adult rats were randomly divided into sham, control, curcumin, and nanocurcumin groups. Partial transection of the gastrocnemius muscle was made in the right limb of the control and treatment groups. The sham and control groups received normal saline, curcumin group received 500 mg/kg of curcumin and nanocurcumin group received 100 mg curcumin-loaded nanomicelles orally every day. They euthanized two weeks later and the specimens were stained by hematoxylin-eosin (H&E) and Masson’s trichrome methods. Aspartate transaminase (AST) and creatine phosphokinase (CPK) were measured in blood samples. Results: The percentage of collagen fibers in the nanocurcumin group was significantly lesser than the control and curcumin groups (p

Nematodes of the genus Libyostrongylus parasitize ostriches, causing high mortality rates. These nematodes are found in the proventriculus and ventriculus of ostriches, but little is known about their distribution and the possible anatomopathological changes they cause in the various regions of these organs. This paper describes the distribution and quantification of Libyostrongylus and pathological changes found in regions of the proventriculus and ventriculus of ostriches with high and low levels of both natural and experimental infection. Ostriches were necropsied and tissue samples from the distinct regions of both organs were analysed based on nematode counts and histopathology after staining with haematoxylin and eosin, Masson's trichrome or Alcian blue/PAS. The cranial and glandular regions of the proventriculus were the most parasitized. The ventriculus contained more nematodes in the caudal region. No macro- or microscopic pathological changes were observed in either of these organs of experimentally-infected birds. However, naturally-infected birds with high levels of infection presented proventriculus with macroscopic lesions and heterophilic infiltrates surrounding nematodes. In the glandular region of this organ, nematodes were located in the adenomeres of the secretory ducts, causing altered architecture and erosions and ulcerative lesions with damaged epithelium. Nematode eggs were found in the koilin layer of the middle and caudal regions of the ventriculus only of these birds. The pH of the regions assessed by Alcian blue/PAS staining changed from acidic in the proventriculus to more alkaline in the caudal region of the ventriculus. These data add knowledge to the biology of Libyostrongylus. RESEARCH HIGHLIGHTS The most parasitized areas were the cranial and glandular regions of the proventriculus. Naturally-infected birds with high levels of infection presented macro lesions in the proventriculus and damaged epithelium. Nematode eggs were found in the ventriculus. The proventriculus had an acidic pH, which turned alkaline towards the ventriculus.

While epiglottis is essentially a mammalian structure, studying its microstructure in any placental model will add an important information to the field of comparative anatomy and the related branches of biology. The aim of this study was to describe the structure of the epiglottis in dromedary camels using light and scanning electron microscopy (SEM), with reference to the possible functions. A total of 11 epiglottis cartilages from 11 larynges were used. The study revealed unusual, deeply situated glands just beneath the cartilage plate. They have unusually, wide surface-openings, while their ducts were partly located within the cartilage. This is presumed to be an adaptation to the need for rapid and efficient mucosal surface hydration in the arid conditions. The possible secretion transport mechanisms in these glands were also discussed. Furthermore, the SEM revealed for the first time, the presence of taste buds in camel epiglottis. However, in histological sections, visibility of taste buds was dependent upon the staining techniques. The taste buds were not seen with standard H& E stain, as they blended imperceptibly with the surrounding epithelium. Conversely, Mallory's trichrome showed contrasting colors, and taste buds were visible. In conclusion, camel epiglottis has an unusual structure, which may be correlated to environmental adaptation and important for the general health of upper respiratory tract in this species.

While first described in 1904, the characterisation of the peridural membrane, which is frequently encountered, yet usually unnoticed, during lumbar decompression surgery, remains inconclusive. This relatively little known membrane is continuous with the posterior longitudinal ligament and lines the epidural space. In this study, we are comparing the membrane and ligamentum flavum from patients to analyse the variations of the histological and ultrastructural compositions. We took samples of the membrane and ligamentum flavum from five separate patients who were undergoing lumbar spine decompression surgery for herniated discs which were then analysed with transmission electron microscopy and stained with H&E (morphology), trichrome (collagen content), and Verhoeff-Van Gieson (elastin content). Upon analysis of the peridural membrane, we observed tightly packed collagen fibres, interspaced with elastin fibres and very few fibroblasts. While the ligamentum flavum showed a significantly higher elastin to collagen ratio and looser arrangement of collagen fibres with a larger extracellular matrix. The peridural membrane was similar in appearance and constituent parts to the dura mater. The peridural membrane is a distinctive and important membrane in the spinal canal, and given its high collagen to elastin ratio and it tightly packed nature, we conclude that it forms a protective layer around the spinal cord which may help in minimising the compressive nature of intervertebral disc herniation.

The present study was aimed to compare different decellularization protocols for human endometrial fragments. The freeze–thaw cycles in combination with treatment by Triton X-100 and four concentrations of sodium dodecyl sulfate (SDS; 0.1, 0.5, 1, and 1.5%) with two exposure times (24 and 72 h) were applied for tissues decellularization. After analysis the morphology and DNA content of tissues the group with better morphology and lower DNA content was selected for further assessments. The nucleus by Acridine orange and extracellular matrix (ECM) using Masson's trichrome, Alcian blue, and periodic acid–Schiff staining were studied. The amount of tissues collagen types I and IV, fibronectin, glycosaminoglycans (GAGs), and elastin was analyzed by Raman spectroscopy. The ultrastructure and porosity of decellularized scaffold were studied by scanning electron microscopy (SEM). The MTT assay was applied for assessments of cytotoxicity of scaffold. The treated group with 1% SDS for 72 h showed the morphology similar to native control in having the minimum level of DNA and well preserved ECM. Raman spectroscopy results demonstrated, the amount of collagen types I and IV, GAG, and fibronectin was not significantly different in decellularized scaffold compared with native group but the elastin protein level was significantly decreased (P

With the rapid advancement in multiplex tissue staining, computer hardware, and machine learning, computationally-based tools are becoming indispensable for the evaluation of digital histopathology. Historically, standard histochemical staining methods such as hematoxylin and eosin, periodic acid- Schiff, and trichrome have been the gold standard for microscopic tissue evaluation by pathologists, and therefore brightfield microscopy images derived from such stains are primarily used for developing computational pathology tools. However, these histochemical stains are nonspecific in terms of highlighting structures and cell types. In contrast, immunohistochemical stains use antibodies to specifically detect and quantify proteins, which can be used to specifically highlight structures and cell types of interest. Traditionally, such immunofluorescence-based methods are only able to simultaneously stain a limited number of target proteins/antigens, typically up to three channels. Fluorescence-based multiplex immunohistochemistry (mIHC) is a new technology that enables simultaneous localization and quantification of numerous proteins/antigens, allowing for the possibility to detect a wide range of histologic structures and cell types within tissue. However, this method is limited by cost, specialized equipment, technical expertise, and time. In this study, we implemented a deep learning-based pipeline to synthetically generate in silico mIHC images from brightfield images of tissue slides-stained with routinely used histochemical stains, in particular PAS. Our tool was trained using fluorescence-based mIHC images as the ground-truth. The proposed pipeline offers high contrast detection of structures in brightfield imaged tissue sections stained with standard histochemical stains. We demonstrate the performance of our pipeline by computationally detecting multiple compartments in kidney biopsies, including cell nuclei, collagen/fibrosis, distal tubules, proximal tubules, endothelial cells, and leukocytes, from PAS-stained tissue sections. Our work can be extended for other histologic structures and tissue types and can be used as a basis for future automated annotation of histologic structures and cell types without the added cost of actually generating mIHC slides.

The placenta is an essential organ in the proper development of pregnancy, and it can present a lot of structural and vascular lesions that can affect fetal development. One of the pathologies associated with pregnancy, which can change the placental structure is thrombophilia (TPh), and this can be correlated with an intrauterine growth restriction (IUGR) of the fetus. Maternal clinical aspects (age, weight) can be correlated with fetal ones (weight, gender), but also with the structural and vascular aspect of the placenta. The placental structure associated with TPh and IUGR shows macroscopic changes, such as fibrin deposition, calcifications and placental infarctions, but microscopic lesions are best highlighted by classical staining techniques: Hematoxylin-Eosin (HE), Masson's trichrome (MT) and Periodic Acid-Schiff (PAS)-Hematoxylin, but also by immunohistochemistry technique with the help of anti-cluster of differentiation 34 (CD34) antibody that could make it possible to quantify vascular density depending on the pathology. Microscopic changes were massive infarcts caused by vascular ischemia, intravenous and extravillous fibrin deposits, calcifications, and vascular thrombosis. All these clinical, morphological and morphopathological data are interconnected and may vary in the presence of TPh and IUGR.

Background: Glucose-6-phosphatase-- β (3), one of multiple isoforms of glucose-6-phosphatase, catalyzes the final step in gluconeogenesis. It is known that mutated G6P3 is associated with severe neutropenia in addition to congenital heart defects, but little is known about the histological changes in cardiac tissue as a result of mutated or deleted G6PC34. Objectives: We sought to further characterize the histological alterations caused by deleted G6PC3 and determine the role of collagen deposition, myocyte proliferation and apoptosis in these changes. Methods: Cardiac tissue from G6PC3 knockout mice and WT mice were harvested, imbedded and stained for markers of collagen (Trichrome), proliferation (KI-67), apoptosis (caspase 3) and hematopoietic stem cells (CD34). Slides were digitally uploaded, and Leica stain quantification was calculated. Results: We demonstrated that in G6PC3 knock out adult mice have significant differences in heart morphology including decreased left ventricular collagen, decreased cellular proliferation and increased apoptosis histologically. Conclusions: As compared to wild type, the hearts of G6PC3 knockout mice demonstrated significantly decreased collagen globally, a crucial component for adequate strength and contractility of myocardial tissue. More investigation should be done to further explore the functional effects of such alterations via echocardiograms.

Neoplasia is rarely reported in decapod crustaceans, and sarcoma has not been previously reported in any crab species. A California king crab Paralithodes californiensis with a recent history of autotomy (4 legs lost) and anorexia was found dead. Grossly, the crab had a pigmented ulcer on the right cheliped merus. Necropsy tissue samples were placed in 10% neutral buffered formalin and processed routinely for histology. Both histochemical (i.e. Brown and Brenn Gram, Fite-Faraco acid fast, Fontana-Masson, Giemsa, hematoxylin and eosin, Masson’s trichrome, periodic acid-Schiff [PAS], phosphotungstic acid-hematoxylin, and von Kossa) and immunohistochemical (i.e. cytokeratin, vimentin, and lysozyme) stains were performed. The body wall (presumably of the right cheliped merus) was ulcerated and subtended by a densely cellular, unencapsulated, invasive neoplasm composed of spindle cells arranged in intersecting streams and bundles embedded in a small to moderate amount of fibromatous stroma. Neoplastic cells were oval to elongate with fibrillar, pale eosinophilic cytoplasm that occasionally contained moderate numbers of small, spherical, brightly eosinophilic granules that were highlighted with PAS and Giemsa stains. Neoplastic cells had mild atypia and no evident mitoses. Immunohistochemical stains were noncontributory. This neoplasm is consistent with hemocytic sarcoma of semi-granulocytic origin. Decapod crustaceans have 3 types of hemocytes: hyalinocytes, granulocytes, and semi-granulocytes. Neoplastic cells had PAS- and Giemsa-positive granules, which are present in both semi-granulocytes and granulocytes. Semi-granulocytes can elongate and are associated with deposition of extracellular matrix during some immune responses. Neoplastic cells were elongate and associated with deposition of matrix. These findings suggest neoplastic cells were of semi-granulocytic origin.

Background and study aim: Regarding the increased number of chronic liver diseases (CLD) patients suffering from ‎many manifestations e.g. diarrhea, this withdrew our attention to try to find a relation ‎between CLD and parasitic infections in those patients‎. Patients and Methods: A case-control study was performed on 190 participants suffering from gastrointestinal ‎complaints especially diarrhea attending Gastroenterology and Hepatology Outpatient ‎Clinics at Zagazig University Hospital, divided into 2 groups GI: 95 CLD patients. GII: 95 ‎non-CLD Control group patients. Cases underwent clinical, abdominal ultrasonographic, ‎and laboratory liver function assessments. Also, collected patients’ stool samples were ‎microscopically examined using iodine, Trichrome and Modified Ziehl-Neelsen stains. To ‎confirm accurate diagnosis to parasitic infections causing diarrhea, RIDA-ELISA for ‎Cryptosporidium spp. and Giardia lamblia copro-antigens detections was performed. Results: Parasitic infections among both studied groups were (47%), comprising (58%) in CLD and ‎‎(36%) in non-CLD as follow: Cryptosporidium (28%, 14%), Giardia (15%, 12%), mixed ‎‎(5%, 3%), E.histolytica spp. (5%, 4%), Blastocystis hominis (3%, 1%) and H. nana (1%, ‎‎2%) respectively. The sensitivity, specificity, PPV, NPP and accuracy of ELISA regarding ‎Giardia and Cryptosporidium infections were (100% and 89.6%, 97.5% and 100%, 87.9% ‎and 100%, 100% and 96.6%, 97.9% and 98.4%) respectively. In GI, most giardial cases ‎had normal ALT and AST levels (74%, 63%), but elevated in cryptosporidial infection ‎‎(59%, 66%) respectively, with statistically significant difference‎. Conclusion: Presence of intestinal parasitic infections; mainly Cryptosporidia and Giardia protozoa ‎among CLD patients was striking when compared to diarrheic non-CLD control group and ‎this may be attributed to impaired immune status.

Giant anteater (Myrmecophaga tridactyla) is a wild mammal distributed in Central and South America; nowadays, it is classified as an endangered species. Research about the macroscopic and histomorphological aspects of its respiratory tract is scarce, and, sometimes, it limits the treatment provided to sick animals and impairs species preservation. Thus, the present study aims to describe the macroscopic and microscopic morphology of its lower respiratory tract, including trachea and lungs. To do so, 12 adult giant anteaters from “Centro de Triagem de Animais Silvestres de Goiânia” (CETAS-GO), Goiás State, Brazil, were used in the research after natural death or euthanasia. Three of these animals were used for macroscopic assessments; they were fixed in 10% buffered formalin and dissected. Trachea and lung tissue samples were collected from nine animals right after death and fixed in 10% buffered formalin for histomorphological analysis; they were processed, embedded in paraffin, and inked with hematoxylin-eosin (HE), periodic acid-Schiff (PAS), and Masson’s trichrome. The macroscopic analysis showed that the trachea in this species is proportionally short and presents from 19 to 27 tracheal cartilages. The right lung presents four lobes and the left one, two. The microscopic analysis evidenced respiratory epithelium of the ciliated cylindrical pseudostratified type, without evident goblet cells in the mucosa layer of the trachea and bronchi. The pulmonary visceral pleura is thick, similar to other large domestic mammals - complete septa extend from the pulmonary visceral pleura. In conclusion, the macroscopy and histomorphology of giant anteater’s lower respiratory tract, represented by trachea and lungs, are similar to that of other domestic and wild mammals. Pulmonary histomorphology is mainly similar to that of pigs and ruminants: it has thick visceral pleura that emits complete septa of conjunctive tissue, which enable lobular parenchymal architecture.

Graphical abstract, Introduction and aim Blastocystis is a common enteric parasite, having a worldwide distribution. Many antimicrobial agents are effective against it, yet side effects and drug resistance have been reported. Thus, ongoing trials are being conducted for exploring anti-Blastocystis alternatives. Proteases are attractive anti-protozoal drug targets, having documented roles in Blastocystis. Serine proteases are present in both hepatitis C virus and Blastocystis. Since drug repositioning is quite trendy, the in vitro efficacy of simeprevir (SMV), an anti-hepatitis serine protease inhibitor, against Blastocystis was investigated in the current study. Methods Stool samples were collected from patients, Alexandria, Egypt. Concentrated stools were screened using direct smears, trichrome, and modified Ziehl-Neelsen stains to exclude parasitic co-infections. Positive stool isolates were cultivated, molecularly subtyped for assessing the efficacy of three SMV doses (100,150, and 200 μg/ml) along 72 hours (h), on the most common subtype, through monitoring parasite growth, viability, re-culture, and also via ultrastructure verification. The most efficient dose and duration were later tested on other subtypes. Results Results revealed that Blastocystis was detected in 54.17% of examined samples. Molecularly, ST3 predominated (62%), followed by ST1 (8.6%) and ST2 (3.4%). Ascending concentrations of SMV progressively inhibited growth, viability, and re-culture of treated Blastocystis, with a non-statistically significant difference when compared to the therapeutic control metronidazole (MTZ). The most efficient dose and duration against ST3 was 150 µg/ml for 72 h. This dose inhibited the growth of ST3, ST1, and ST2 with percentages of 95.19%, 94.83%, and 94.74%, successively and viability with percentages of 98.30%, 98.09%, and 97.96%, successively. This dose abolished Blastocystis upon re-culturing. Ultra-structurally, SMV induced rupture of Blastocystis cell membrane leading to necrotic death, versus the reported apoptotic death caused by MTZ. In conclusion, 150 µg/ml SMV for 72 h proved its efficacy against ST1, ST2, and ST3 Blastocystis, thus sparing the need for pre-treatment molecular subtyping in developing countries.

The maned wolf, Chrysocyon brachyurus, is a near-threatened carnivore inhabiting the Brazilian Cerrado. Few studies have been conducted on this species, and even fewer have explored its ophthalmological characteristics. Vision is critical to wild canids; thus, this study aimed to provide a morphoquantitative description of the bulbar conjunctiva of the maned wolf using cytological and histological analyses. Ten healthy maned wolves from a conservational centre, including 4 females and 6 males aged 1-12 years (6.5 ± 2.8), were included in the study. The samples for cytological analysis were collected from the inferior conjunctival sac using a cytobrush, and conjunctival tissue was collected for histological analysis from the temporal canthus zone. The cytological samples were stained using the Papanicolaou method, and the histological sections were stained using haematoxylin and eosin, Periodic acid-Schiff, picrosirius red and Masson's trichrome stains. The cytological samples were studied for stain quality, and the different cell types were counted. Histological examination was used to determine tissue types in the conjunctiva and their proportions. Analyses revealed a stratified squamous epithelium with some goblet cells and eventual pigmentation in the basal layer. Loose connective tissue with the presence of some mononuclear and polymorphonuclear cells was also observed. The epithelium of the maned wolf's bulbar conjunctiva resembles that of dogs and other carnivore species; furthermore, its physiological and pathological responses were similar to those of other carnivore species.

Amaç: Bağırsak parazitlerinin neden olduğu hastalıklar dünya genelinde olduğu gibi ülkemizde de önemli bir halk sağlığı problemidir. Bu çalışmada çeşitli gastrointestinal sistem yakınmaları ile Dokuz Eylül Üniversitesi Hastanesi’ne (DEÜH) başvuran 1hastalardaki bağırsak parazitleri araştırıldı.Yöntemler: Çalışmaya Ocak 2011-Aralık 2018 tarihleri arasında DEÜH Merkez Parazitoloji Laboratuvarı’na gönderilen 18460 hasta dahil edildi. Hastalara ait dışkı örnekleri Nativ-lugol yöntemiyle incelendikten sonra bu örneklere formol etil-asetatçöktürme yöntemi uygulandı. Gerekli görülen örneklere trikrom ve kinyoun asit-fast boyama yapıldı. Hastalara ait demografik verilere ise hastane ve laboratuvar bilgi işletim sistemi üzerinden ulaşıldı.Bulgular: İncelenen toplam 18460 hastanın %6’sında (1128) bir veya birden fazla parazit saptandı. Parazit saptanan olguların yaş ortalaması 39,7 (±23,1) yıl olmakla birlikte bunların %53,3’ü erkek, %47,6’sı kadındı. Pozitiflik oranının parazitlere göre dağılımı incelendiğinde ise; %4,8’i (879) Blastocystis hominis, %0,7’si (135) Entamoeba histolytica/dispar harici diğer amipler, %0,4’ü (70) Giardia intestinalis, %0,3’ü (49) Enterobius vermicularis, %0,1’i (21) Entamoeba histolytica/dispar ve %0,01’i (10) nadir görülen diğer parazitler şeklinde idi.Sonuç: Çalışmamız, bağırsak parazit enfeksiyonlarının bölgemizde halen önemli bir halk sağlığı sorunu olarak devam ettiğini göstermekle birlikte; görülme sıklıklarındaki düşüşü ortaya koyması açısından da önemlidir. Objective: Intestinal parasitic diseases are important public health problems in our country as well as in the world. In this study,intestinal parasites were investigated in patients admitted to Dokuz Eylül University Hospital (DEUH) with various gastrointestinal system complaints.Methods: Patients (n=18460) who were referred to the DEUH Central Parasitology Laboratory between January 2011 and December 2018, were included in the study. Fecal samples were examined with Nativ-lugol method and then formol ethylacetateprecipitation method was applied. Trichrome and kinyoun acid-fast stainings were performed on the necessary samples.Demographic data of the patients were obtained from the hospital’s and laboratory’s information operating system.Results: One or more parasites were detected in 6% (1128) of 18460 patients examined. The mean age of the patients with parasites was 39.7 (±23.1) years, of which 53.3% were male and 47.6% were female. The distribution of parasites detected were as follows; 4.8% (879) Blastocystis hominis, 0.7% (135) amoebas other than Entamoeba histolytica/dispar, 0.4% (70) Giardia intestinalis, 0.3% (49) Enterebius vermicularis, 0.1% (21) Entamoeba histolytica/dispar, and 0.01% (10) other rare parasites.Conclusion: Our study shows that intestinal parasitic infections are still an important public health problem in our region and that there is a decrease in their incidence