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Background Methimazole is an antithyroid drug. It has side effects on many tissues. Bone marrow-derived mesenchymal stem cells (BM-MSCs) are promising in the field of tissue regeneration. Objective To investigate the capacity of BM-MSCs on odontogenic differentiation and dentin regeneration at different time intervals in methimazole treated rats. Methods Twenty-eight male albino rats were classified as: Group I: got distilled water. Group II: obtained therapeutic dosage of methimazole as pro-drug “Neo-Mercazole®”. Group III: received methimazole then solitary injection of BM-MSCs at day 21. Group IV: obtained methimazole and single injection of BM-MSCs at the beginning of the experiment. Light microscope was used to examine specimens. Recently formed collagen and β-catenin-immunoreactivity area% were appraised histomorphometrically and statistically. Results Histological examination of odontoblasts and dentin illustrated normal structure in Group I and nearly normal features in Group IV. Group II demonstrated discontinuation of odontoblastic layer and areas of different stainability in dentin. Group III showed an evidently wide layer of odontoblast-like cells and distinct dentinal tubules. Masson's trichrome results of dentin in Groups I &IV showed apparently equal areas of new and old collagen. Group II illustrated old collagen mainly. Group III explored new collagen only. β-catenin-immunoreactivity was strong in Groups I & IV, mild in Group II and moderate in Group III. Statistical results revealed that the highest mean of newly formed collagen area% was in Group III, followed by Group I, Group IV then Group II respectively. Regarding β-catenin-immunoreactivity area%, the highest mean was recorded in Group I, subsequently Group IV, next Group III then Group II. Conclusions Methimazole has destructive consequences. BM-MSCs have a time-based increased capacity for odontogenic differentiation and regeneration of dentin.

Background: Osteoarthritis (OA) is the most common degenerative joint disease for which there is no permanent treatment. The most affected structures are articular cartilage and subchondral bone. Knees are the most affected by OA, thus leading to severe disability. N-Acetyl phenylalanine glucosamine (NAPA) is a derivative of glucosamine (GlcN), which is one of the components of glycosaminoglycans (GAGs) of cartilage extracellular matrix (ECM).Aim: This work aimed to study the possible curative effect of N-acetyl phenylalanine glucosamine on surgically induced osteoarthritis in the knee joint of the adult male albino rat.Materials and Methods: This study was performed on 30 adult male albino rats. They were divided into three groups: group I included ten animals that were subdivided into two subgroups, IA (control) & IB (underwent Sham operation), group II included ten animals underwent destabilization of medial meniscus (DMM) surgery without receiving any treatment, group III included ten animals underwent DMM surgery then received a single intraarticular injection of NAPA after 4 weeks from the surgery and sacrificed 4 weeks later. At the end of the experiment, knee joints were obtained and processed for histological and immunohistochemical studies.Results: Hematoxylin and Eosin-stained sections showed a moderate degree of OA in group II that was improved in group III. Mallory’s trichrome stained sections revealed decreased collagen content in group II that was also improved in group III. The same results were found as regards the GAGs content of the cartilage matrix that was clearly decreased in group II and restored in group III.Conclusion: From the present study, it could be concluded that the osteoarthritic changes induced by surgical DMM in adult male albino rats can be improved by subsequent intra-articular administration of NAPA.

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.

The aim of this study was to assess the histological effect of Low Level Laser Therapy (LLLT) (904 nm) on the repair of standardized bone defects on the femur of rats. Sixty male wistar rats were assigned into two equal groups. Group (A: laser group) and group (B: control group). A surgical fracture was done in middle third of femur of all rats. In group (A) a continuous wave 904 nm infrared laser was applied at dose 4 j/cm2 at fracture site immediately post operative for 7 sessions, each session was 5 minutes. The animals were killed by over dose of general anesthesia on the 15th,30th and 45th days after surgery, The specimens were processed and stained with Hematoxylin-eosin (H/E), special stain Masion trichrome and analyzed by light microscopy. The descriptive analysis of histological imaging showed greater degree of new bone formation, osteoblastic surface and collagen fiber in the irradiated group when compared with the control group. Based on the obtained results, this study concluded that LLLT was efficient in promoting bone healing, and increasing new bone formation in the process of surgically fractured femur in animal study.

Background: Oral submucous fibrosis (OSMF) is graded according to various histological factors which include the epithelial changes and the connective tissue changes. These features could be identified in routine Hematoxylin and Eosin (H and E) staining in shades of pink. However, the use of special stains provides a contrast to various connective tissue components thereby aiding in better visualization of these connective tissue changes in advanced OSMF cases. Objective: To compare and evaluate muscle involvement and degeneration in advanced oral submucous fibrosis using three different stains namely, H&E, Van Gieson, and Masson’s Trichrome. Material and Methods: 10 Formalin-fixed paraffin-embedded tissue sections of advanced OSMFwere stained using 3 different stains namely Hematoxylin and Eosin (H&E), Van-Gieson, and Masson trichrome. The results obtained were tabulated and statistically analyzed using Kruskal Wallis ANOVA. Results: The hyalinization and fibrosis involving the skeletal muscle were better visualized in Masson’s Trichrome but were not statistically significant. The muscle degeneration in deeper areas was better visualized in Masson’s trichrome when compared to the H&E and Van Gieson. Conclusion: Hematoxylin and eosin stains all the connective tissue components in various shades of pink, use of special stains bestows contrast between different components of connective tissue, thus improvising grading of OSMF. Masson’s trichrome stain can be used as a single adjunct to hematoxylin and eosin stain as changes in the superficial and deeper connective tissue could be ascertained. Keywords Fibrosis; Masson’s trichrome stain; Muscle degeneration; Oral submucous fibrosis grading.

Background and Objectives: Cardiovascular diseases is a primary health concern. Recent studies have demonstrated that high-fat diet (HFD)-induced obesity could lead to cardiac dysfunction, hypertrophy, fibrosis and even heart failure. Quercetin (Q) is a common nontoxic polyphenol having a broad range of pharmacological and biological activities. The present study aimed at evaluating the histological and ultrastructural effects of high fat diet on cardiac muscle of adult male albino rat and the possible protective role of quercetin.Materials and Methods: Forty adult male albino rats were used and divided into four groups. Group I: Control group (10% of energy from fat). Group II: Quercetin group (10% of energy from fat + 60 mg/kg quercetin). Group III: HFD group (60% of energy from fat). Group IV: HFDQ group (60% of energy from fat + 60 mg/kg quercetin).At the end of experiment after12 weeks, all rats were sacrificed, and hearts were taken and processed for light (H&E and Mallory’s trichrome stains) and electron microscopic examination. Histological, morphometric and statistical studies were performed. Results: Sections from rats of group III showed widely separated cardiomyocytes, some atrophic degenerated fibers, vacuoles in the cytoplasm, engorged capillaries and marked increase in collagen deposition. Electron microscopic examination showed disorganized fragmented myofibrils, few swollen mitochondria and bizarre indented nuclei. Sections from rats of group IV revealed obvious improvement of the previous alterations. There was highly statistically significant difference between group I and group III and between group II and group III as regard collagen area percent. Conclusion: The results obtained from the current study confirmed the negative influence of high fat diet intake on cardiomyocytes. Concomitant administration of quercetin markedly improved myocardial damage so, it could be used as an effective cardioprotective agent against hyperlipidemia and aging insults.

STUDY QUESTION Can ovarian tissue morphology be better preserved whilst enabling histological molecular analyses following fixation with a novel fixative, neutral buffered formalin (NBF) with 5% acetic acid (referred to hereafter as Form-Acetic)? SUMMARY ANSWER Fixation with Form-Acetic improved ovarian tissue histology compared to NBF in multiple species while still enabling histological molecular analyses. WHAT IS KNOWN ALREADY NBF fixation results in tissue shrinkage in various tissue types including the ovary. Components of ovarian tissue, notably follicles, are particularly susceptible to NBF-induced morphological alterations and can lead to data misrepresentation. Bouin’s solution (which contains 5% acetic acid) better preserves tissue architecture compared to NBF but is limited for immunohistochemical analyses. STUDY DESIGN, SIZE, DURATION A comparison of routinely used fixatives, NBF and Bouin’s, and a new fixative, Form-Acetic was carried out. Ovarian tissue was used from three different species: human (n = 5 patients), sheep (n = 3; 6 ovaries; 3 animals per condition) and mouse (n = 14 mice; 3 ovaries from 3 different animals per condition). PARTICIPANTS/MATERIALS, SETTING, METHODS Ovarian tissue from humans (aged 13 weeks to 32 years), sheep (reproductively young i.e. 3–6 months) and mice (10 weeks old) were obtained and fixed in 2 ml NBF, Bouin’s or Form-Acetic for 4, 8, and 24 h at room temperature. Tissues were embedded and sectioned. Five-micron sections were stained with haemotoxylin and eosin (H&E) and the percentage of artefact (clear space as a result of shrinkage) between ovarian structures was calculated. Additional histological staining using Periodic acid-Schiff and Masson’s trichrome were performed on 8 and 24 h NBF, Bouin’s and Form-Acetic fixed samples to assess the compatibility of the new fixative with stains. On ovarian tissue fixed for both 8 and 24 h in NBF and Form-Acetic, immunohistochemistry (IHC) studies to detect FOXO3a, FoxL2, collagen IV, laminin and anti-Müllerian hormone (AMH) proteins were performed in addition to the terminal deoxynucleotidyl transferase nick end labelling (TUNEL) assay to determine the compatibility of Form-Acetic fixation with types of histological molecular analyses. MAIN RESULTS AND THE ROLE OF CHANCE Fixation in Form-Acetic improved ovarian tissue morphology compared to NBF from all three species and either slightly improved or was comparable to Bouin’s for human, mouse and sheep tissues. Form-Acetic was compatible with H&E, Periodic acid-Schiff and Masson’s trichrome staining and all proteins (FOXO3a, FoxL2, collagen IV and laminin and AMH) could be detected via IHC. Furthermore, Form-Acetic, unlike NBF, enabled antigen recognition for most of the proteins tested without the need for antigen retrieval. Form-Acetic also enabled the detection of damaged DNA via the TUNEL assay using fluorescence. LARGE SCALE DATA N/A LIMITATIONS, REASONS FOR CAUTION In this study, IHC analysis was performed on a select number of protein types in ovarian tissue thus encouraging further studies to confirm the use of Form-Acetic in enabling the detection of a wider range of protein forms in addition to other tissue types. WIDER IMPLICATIONS OF THE FINDINGS The simplicity in preparation of Form-Acetic and its superior preservative properties whilst enabling forms of histological molecular analyses make it a highly valuable tool for studying ovarian tissue. We, therefore, recommend that Form-Acetic replaces currently used fixatives and encourage others to introduce it into their research workflow. STUDY FUNDING/COMPETING INTEREST(S) This work was supported by the Oxford Medical Research Council Doctoral Training Programme (Oxford MRC-DTP) grant awarded to B.D.B. (Grant no. MR/N013468/1), the Fondation Hoffmann supporting R.A. and the Petroleum Technology Development Fund (PTDF) awarded to B.V.A.

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

Background: Osteoarthritis (OA) is a common degenerative joint disease. Intra-articular injection of microcrystalline corticosteroid and platelet- rich plasma (PRP) are used for pain relief of knee OA.Aim of the Study: the goal of this study was to assess the possible therapeutic effects of corticosteroid compared to PRP in a rat model of induced OA.Material and Methods: Intra-articular injection of monosodium iodoacetate (MIA) was used for induction of OA in rats. Blood was collected from12 adult male rats for PRP preparation. 54 adult female albino rats were divided into control groups, OA model group and treated groups which received CS or PRP. All rats were euthanized after 28 days. Left knee joints were processed for histological examination by light microscope after staining by H& E, toluidine blue and masson’s trichrome stains. Right knee joints were grossly examined by stereomicroscope and scanning electron microscope (SEM). The obtained data were examined, further statistically analyzed and interpreted. Results: MIA caused gross anatomical and histological alterations in rat’s articular cartilage similar to human knee OA. Articular cartilage of corticosteroid and PRP treated knee joints showed thinning of the articular cartilage, fibrillation and presence of pannus- like tissue. These groups also showed significant increase in histopathological scores compared to the control group and OA model group in addition to gross macroscopic changes that were confirmed by SEM.Conclusion: This study showed that neither corticosteroid nor PRP could limit the structural changes in osteoarthritis and that different PRP preparation techniques have yielded varying results.

Highlights • Incisional wound closure is a critical surgical step to facilitate tissue healing. • Cyanoacrylates (Histoacryl®) have become a popular veterinary wound closure practice. • Effects of Histoacryl® over traditional suture on wounds rest undetermined. • Histoacryl® vs. Poliglecaprone (Monocryl®) have similar regenerative results in mice. • Histoacryl® or Monocryl® could be used to treat wounds in rodents., Incisional wound closure is a key surgical step to facilitate tissue healing, reduce the risk of infection and obtain esthetic and functional recovery. Cyanoacrylates such as Histoacryl® have become a popular choice in surgical veterinary practice. However, how Histoacryl® is affecting tissue regeneration and bacterial load in the wound in comparison to poliglecaprone (Monocryl®) traditional suture methods remains to be determined. This work aimed to evaluate how wounded tissue responds to traditional suture with Monocryl® (poliglecaprone 25/4-0) and Histoacryl®, as well as provide evidence of their effects on wound healing in mice. Fortyeight hours after the incisional procedure, wound tissue biopsies were prepared for histological and microbiological analysis. Biopsies were fixed and colored with Mallory's trichrome and hematoxylin-eosin stains. For microbiological assays, biopsies were suspended in tryptic soy broth (TSB) and 1/10 diluted to evaluate the number of CFU in nutrient agar plates. Our results show no differences between Histoacryl® and Monocryl® traditional suture suggesting that both methods could be used to treat wounds in small animals such as rodents.

Purpose: Dipotassium glycyrrhizinate (DPG) has anti-inflammatory properties, besides promoting the regeneration of skeletal muscle. However, it has not been reported on skin wound healing/regeneration. This research aimed to characterize the effects of DPG in the treatment of excisional wounds by second intention. Methods: Male adults (n=10) and elderly (n=10) Wistar rats were used. Two circular wounds were excised on the dorsal skin. The excised normal skins were considered adult (GAN) and elderly (GIN) naïve. For seven days, 2% DPG was applied on the proximal excision: treated adult (GADPG) and elderly (GIDPG), whereas distal excisions were untreated adult (GANT) and elderly (GINT). Wound healing areas were daily measured and removed for morphological analyses after the 14th and the 21st postoperative day. Slides were stained with hematoxylin-eosin, Masson’s trichrome, and picrosirius red. Results: Histological analysis revealed intact (GAN/GIN) and regenerated(GANT/GINT/GADPG/GIDPG) skins. No differences of wounds’ size were found among treated groups. Epidermis was thicker after 14 days and thinner after 21 days of DPG administration. Higher collagen I density was found in GIDPG (14th day) and GADPG (21st day). Conclusions: DPG induced woundhealing/skin regeneration, with collagen I, being more effective in the first 14 days after injury.

Background Proper assessment of dermal collagen fibers by dermatologists and researchers is essential. Histologic evaluation methods have limitations. We present a simple method for measurement of collagen fibers in human skin using Masson's trichrome staining. Materials and methods Normal skin specimens from a cadaver were processed with Masson's trichrome, which can effectively stain collagen fibers blue with aniline dye. Optical photomicrographs of these slides were analyzed using ImageJ software. Color image processing, a histogram-based function of ImageJ for image segmentation, was performed with color moments thresholding technique. We selected blue areas by adjusting the blue channel to include specific values. The selected areas were highlighted and evaluated. We divided the image into layers of 0.09-mm2 areas from the top to bottom of the dermis. Each area was cropped and evaluated. Results Quantitative assessment yielded the quantitative size occupied by collagen fibers in an area of 0.09 mm2 . Calculation of the percentage in each area can be used to determine the density of collagen fibers. Conclusion Measurements obtained with our method can be applied to research on dermal collagen fibers. We present a convenient quantitative assessment method for the dermal constituents in Masson's trichrome-stained slides.

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%).

Preoperative radiation is associated with an increased risk of wound complications. However, the influences of radiation on musculofascial wound healing remains unclear. The purpose of the study was to investigate the short-term effects of preoperative local radiation on the musculofascial healing of laparotomy incisions in a rat model. Eighteen Fischer 344 rats received radiation doses of 0, 10, or 20 Gy to the abdominal wall and underwent laparotomy 4 weeks later. Two weeks after laparotomy, samples of irradiated muscle were harvested for mechanical tests, histological (Hematoxylin & Eosin, and Masson’s Trichrome) and immunohistochemical analyses using KI67, CD31, TGF-β, and MYOD1 antibodies. The elastic modulus (EM), maximum strain (MS), and ultimate tensile strength (UTS) in the 20-Gy group were significantly weaker than those in the 0-Gy group. The EM and UTS in the 20-Gy group were significantly lower than those in the 10-Gy group. The UTS and MS in the 10-Gy group were significantly lower than those in the 0-Gy group. The mean number of inflammatory cells per mm2 in the 20-Gy group was significantly larger than those in the 10- and 0-Gy groups. The mean numbers of CD31-, KI67-, and MYOD1-positive cells, the optical density of TGF-β, and the microvessel density in the 20-Gy group were significantly smaller than those in the 10- and 0-Gy groups. These results indicated that radiation delays musculofascial healing and decreases mechanical strength of the laparotomy incision by creating a chronic inflammatory environment, inhibiting cell proliferation, angiogenesis, granulation maturation, collagen deposition, and muscular regeneration in a dose-dependent manner. The impaired biomechanical, histological and molecular properties may be associated with the higher risk of wound complications in patients who undergo radiotherapy prior to laparotomy.

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.

Safety concerns on the toxic and carcinogenic effects of formalin exposure have drawn increasing attention to the search for alternative low risk fixatives for processing tissue specimens in laboratories worldwide. Alcohol-based fixatives are considered some of the most promising alternatives. We evaluated the performance of alcohol-fixed paraffin-embedded (AFPE) samples from Sprague-Dawley (SD) rats analyzing tissue morphology, protein and nucleic acid preservation after short and extremely long fixation times (up to 7 years), using formalin-fixed paraffin-embedded (FFPE) samples as a comparator fixative. Following short and long-term alcohol fixation, tissue morphology and cellular details in tissues, evaluated by scoring stained sections (Hematoxylin-Eosin and Mallory’s trichrome), were optimally preserved if compared to formalin fixation. Immunoreactivity of proteins (Ki67, CD3, PAX5, CD68), evaluated by immunohistochemistry, showed satisfactory results when the fixation period did not exceed 1 year. Finally, we confirm the superiority of alcohol fixation compared to formalin, in terms of quantity of nucleic acid extracted from paraffin blocks, even after an extremely long time of alcohol fixation. Our results confirm that alcohol fixation is a suitable and safe alternative to formalin for pathological evaluations. There is a need for standardization of formalin-free methods and harmonization of diagnosis in pathology department worldwide.

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

Introduction: The material we have in our hands represents a commonly used additive for enhancing the savor of different foods producing an exceptional kick. The oxidative stress might be augmented by high intake of MSG and results in cytotoxicity of various tissues. Aim: To evaluate the effects of MSG on the buccal mucous membrane of rats (male, adult, albino) through histological and immune-histochemical investigations. Material and methods: Thirty adult male albino rats (n=30) were used in this study. Random division of the samples were done, grouping those samples within 2 alike groups; group I and group II (n=15 rats). Group I (Control); rats were treated by distilled water (2 ml) daily by gastric tube, and Group II (MSG treated); rats were given MSG daily using a 30 mg/kg dosage which is thawed in (2 ml) distilled water by gastric tube. Rats of all groups, were euthanized and sacrificed six weeks after the start of experiment. Specimens from buccal mucosa were dissected out and processed where, Hematoxylin and Eosin and Mallory trichrome stains were used at one part, and in the other part immuno-histochemical stains were utilized for identifying the expression of (PCNA) gene. Results: The histological results of the buccal mucous membrane of MSG- supplemented group as compared, to control group, exposed shrivelled epithelial alterations, alongside degenerated cells having pyknotic nuclei. While, the collagen fibers were strongly deposited at lamina propria with dilated and congested blood vessels and immunohistochemical results revealed many epithelial cells with a strong brown positive PCNA. Conclusion: Low doses of monosodium glutamate-induced degenerative effects on the buccal mucosa of rats. Subsequently, we recommend, to restrict, the using of MSG as food flavor enhancer.

The purpose is to examine early wound healing through histological analysis by characterizing connective tissue distribution and organization in the treated periodontium following nonsurgical therapy. Periodontal disease is a multifactorial pathological process that leads to the loss of the surrounding periodontium. Traditional periodontal therapies have proven beneficial in halting the progression of disease. The aim of this study is to investigate early wound healing in periodontal patients following hand/ultrasonic instrumentation alone, erbium‐doped yttrium aluminum garnet laser instrumentation alone, or a combination of hand/ultrasonic instrumentation and Er:YAG laser instrumentation for the nonsurgical treatment of periodontitis by histologic evaluation. Twenty‐one patients were randomized to receive nonsurgical therapy for the treatment of chronic periodontitis with three modalities prior to surgical therapy. Baseline clinical measurements were obtained prior to treatment. Wound healing was assessed by obtaining an otherwise discarded tissue sample following nonsurgical therapy of the selected study site. Samples were obtained at 2 or 6 weeks following initial therapy with a step‐back incision and fixated for histological and immunohistochemical analysis. There were minimal between‐group differences in the amount of collagen distribution when analyzing the Mallory–Heidenhain Azan trichrome, Picrosirus Red stain, and proliferating cell nuclear antigen at both time points. Descriptive analysis of baseline measurements showed no differences in probing depth change, bleeding on probing, and clinical attachment level following initial therapy between the three treatment groups at 2 or 6 weeks. Each treatment modality was effective in treating moderate to severe chronic periodontitis; however, the results of this study are inconclusive regarding superiority of any one treatment approach from a histologic and immunohistochemical perspective. Based on this assessment, there was increased fibroblast proliferation and collagen maturation between the 2‐ and 6‐week time point after treatment in all treatment groups, with few apparent differences between treatment groups. This pilot study qualitatively evaluated early wound healing in periodontal patients following non surgical therapy with various treatment modalities. When comparing descriptive outcomes of Er:YAG laser therapy and hand/ultrasonic instrumentation there were minimal differences in collagen distribution and density between groups. The evaluated modalities were each effective treating periodontal patients with non surgical therapy.

Background: Functional impairment of the muscle-tendon unit is one of the most remarkable effects of aging. The function of the peroneus longus muscle is to stabilise the foot and ankle joint. A deterioration of the structural and mechanical properties of this muscle can potentially lead to foot problems in older adults. This study aimed to investigate the effects of age on structural, histological, and mechanical features in peroneus longus muscle samples taken from embalmed cadavers of two different age groups; young adult (30–60 years) and old adult (over 60 years). Materials and methods: The mechanical property was analysed through the results of cross-sectional area, tensile, tensile stress, and modulus of elasticity. The arran- gement of the collagen in the perimysium and tendon was examined by scanning electron microscopy. Fatty infiltration within the musculotendinous junction was evaluated by Masson’ trichrome stained muscle sections. Results: This study thus provides evidence that there are indeed age-induced mechanical property changes in the peroneus longus muscle, which include reductions in the tensile force, tensile stress, and modulus of elasticity, and is related to the malformation of collagen fibrils and the massive fat accumulation in the musculotendinous junction. Conclusions: These alterations may further result in a reduction of muscle strength and quality in an older person.

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.

Trichrome painted wares spread out during the Middle Neolithic (between 5000 and 4500 cal. BC) along the Adriatic side of the Italian Peninsula. The pottery production was characterized by a very fine-granulated paste defined figuline and by a decoration painted with red and black colors. This research reports the results of X-Ray Fluorescence (XRF) and micro-Raman spectroscopy analysis focused on the decoration of the ceramics from two distinct geographical groups, corresponding to the village of Ripoli (Abruzzo, Central Italy) and to the Matera's caves (Basilicata, Southern Italy). A representative set of samples has been studied. XRF and micro-Raman analyses reveal that the black decoration was obtained using a black pigment based on manganese-iron oxide, whereas the red one includes iron oxides. To investigate the multilayer structure of the samples, a Monte Carlo simulation has been performed on the XRF data. The results, together with the data achieved on samples of the Serra d'Alto Culture from Matera's sites, allow to confirm previous hypotheses on the figuline production. In particular, a technological evolution is proven by the selection and control of the raw materials associated to a technologically advanced firing system. The choice of a manganese pigment is a further technological connotation of figuline potteries.

Extracellular matrix (ECM) scaffolds have been shown to promote constructive remodeling response into a host. Minimal disruption of ECM ultrastructure during tissue processing is typically desirable. The purpose of the study was to prepare and characterize bubaline aortic matrix (BAM) for abdominal hernia repair in cattle. The bubaline aorta was decellularized with 1% sodium dodecyl sulfate (SDS) for 24 h followed by 0.25% trypsin for 2 h and again with 1% SDS for 24 h. Native and decellularized aortae were examined for residual cells, collagen structure and integrity by histologic examination and scanning electron microscopy (SEM). Masson’s trichrome and Weigert’s staining, DNA quantification and Fourier transform infrared (FTIR) spectroscopy were used for further characterization of BAM. BAM was used for repair of abdominal hernia in six cattle having mean weight of 132.50 ± 21.86 kg and hernial ring size of 40.55 ± 15.42 cm2. Clinical, hemato-biochemical and antioxidant parameters were evaluated to assess biocompatibility of xenogenic BAM. Histologically, the absence of cells and orderly arranged collagen fibers were observed in treated aorta. SEM confirmed the preservation of collagen structure and integrity. DNA content was significantly (P

Background: Skin is the largest organ of the body in surface area and weight. One of the most important side effects of topical glucocorticoids therapy is skin atrophy. This study aimed to evaluate the possible therapeutic effects of curcumin versus aftamed gel (Hyaluronic Acid) on glucocorticoid-induced skin atrophy, as well as on subsequently induced abrasion wounds. Methods: Forty two adult male albino rats were divided into 4 groups: Group1 (Control group) of 6 rats, Group II (Steroid-Exposed Group), Group III (Curcumin-Treated Group) and Group IV (Aftamed-treated group). Each of group II, III and IV were subdivided into two subgroups IIA, IIIA, IVA (subjected to clobetasol propionate cream) each included 6 rats and IIB, IIIB, IVB (subjected to clobetasol propionate cream, then exposed to skin abrasion) each included 6 rats. Skin biopsies were taken from the dorsum of rats 12th days from the stoppage of clobetasol propionate application. Sections were stained with hematoxylin and eosin, Masson's trichrome and immunohistochemichal stain for PCNA & CK19. Epidermal thicknesses, mean area percent of collagen fibers and mean number of PCNA & CK19 immunopositive cells were measured. Results: Steroid-exposed group showed thinning of epidermal and dermal layers with scanty hair follicles and sebaceous glands as well as delay in wound healing. Increased epidermal thickness and reorganized dermal structural elements with increased PCNA & CK19 immunopositive cells were more obvious in curcumin treated group than aftamed treated group. Conclusion: topical application of curcumin had more improving effect on skin and its abrasions rather than aftamed gel.

The histologic findings of a subretinal band and epiretinal membrane (ERM) excised from two patients who developed retinal detachments (RDs) after non-U.S. Food and Drug Administration-regulated intravitreal “stem cell” injections are reported. Both membranes were composed of fibrocellular tissue that stained positively with Smooth Muscle Actin and Masson's trichrome, consistent with collagenous and smooth muscle composition. CD34 immunostain (for hematopoietic cells) was negative for the subretinal band and minimally positive for the ERM. The authors speculate that the “stem cells” may cause RDs by differentiation into myofibroblasts that cause tractional membranes, though further studies are warranted. [ Ophthalmic Surg Lasers Imaging Retina. 2019;50:125–131.]

Knowledge of skeletal muscle adaptations is important to understand the functional deficits in cerebral palsy (CP). This study aimed to investigate the morphofunctional characteristics of skeletal muscle in a CP animal model. Initially, pregnant Wistar rats were injected intraperitoneally with saline or lipopolysaccharide over the last five days of pregnancy. The control group (n = 8) consisted of male pups born to females injected with saline. The CP group (n = 8) consisted of male pups born to females injected with lipopolysaccharide, which were submitted to perinatal anoxia [day of birth, postnatal day 0 (P0)] and sensorimotor restriction (P1‐P30). The open‐field test was undertaken on P29 and P45. On P48, the animals were weighed, and the plantaris muscle was collected and its weight and length were measured. Transverse sections were stained with haematoxylin‐eosin, NADH‐TR, Masson's trichrome and non‐specific esterase reaction for analysis. and transmission electron microscopy was performed. In the CP group, reductions were observed in mobility time, number of crossings and rearing frequency, body weight, muscle weight and length, and nucleus‐to‐fibre and capillary‐to‐fibre ratios. There was a statistically significant increase in the percentage area of the muscle section occupied by collagen; reduction in the area and increase in the number of type I muscle fibres; increase in myofibrillar disorganization and Z‐line disorganization and dissolution; and reduction in the area and largest and smallest diameters of neuromuscular junctions. Thus this animal model of CP produced morphofunctional alterations in skeletal muscle, that were associated with evidence of motor deficits as demonstrated by the open‐field test.

Cancer chemotherapy is confronted with insufficient drug penetration in tumors. "Solid tumor priming" is proposed to modulate the abnormal tumor microenvironment but suffers from limited digestion efficiency and underlying tumor metastasis. Losartan (Los) and telmisartan (Tel) are well-known antihypertensive agents and show capabilities in inhibiting collagen synthesis by cancer-associated fibroblasts. Up to now, no attempt has been made to achieve a local and sustained release of Los and Tel in tumors while alleviating the side effects after systemic administration. In the previous study, micelles were loaded into fiber fragments to achieve high drug accumulation in tumors after intratumoral administration. In the current study, Los and Tel are blend electrospun into fibers to retard the collagen synthesis and promote the tissue penetration of micelles released from fiber fragments. The loading of Los and Tel shows no effect on the micelle release, cellular uptake, and cytotoxicities of micelles released from fiber fragments. Because of the hydrophilicity, Los is almost released out after 5 day in pH 6.8 buffers, while hydrophobic Tel is gradually released for 30 days. Thus, fiber fragments with loaded Los and Tel are combined to achieve a sustained remodeling of collagen levels in tumors, and the combination with a ratio of 1/2 showed the most significant and consistent reductions of collagen I levels in tumors, as determined via Western blotting, Masson's trichrome, and immunofluorescence staining. A wide distribution of micelles is observed in the tumor tissues, as well as strong fluorescence in the distal sections of tumors during 14 days. Compared with pristine fiber fragments, the sequential release of Los and Tel induces stronger inhibition of tumor growth, lower expression of hypoxia-inducible factor-α (HIF-α), and fewer tumor metastases to lungs. Thus, this study demonstrates a feasible strategy to enhance the local retention and even distribution of chemotherapeutic agents in tumors in favor of therapeutic efficacy.

Background Patellar tendinopathy is difficult to successfully treat. This study aimed to characterize the pathological changes of the infrapatellar fat pad (IPFP) in patellar tendinopathy (PT), and to investigate the influence of PT on the development of fibrotic changes in the IPFP. Methods Forty male Wistar rats were randomly divided into PT (n = 20) and control groups (n = 20). Bacterial collagenase I (patellar tendinopathy group) or saline (control) was injected, intratendinous, into the patellar tendon. Rats were sacrificed at week 12. The whole knee joint was sagittally sectioned and stained with hematoxylin–eosin and Masson's trichrome. The IPFP samples were graded according to cellularity, fibrosis, and vascularity. The whole IPFP and blue-stained area was measured. Mann–Whitney U tests were used to compare the between-group differences of each score and quantitative value. Results Scores for cellularity were three (2–3) and 0 (0–1) in the PT and control groups, respectively (P Conclusion The IPFP in the patellar tendinopathy group showed greater cellularity, fibrosis, and vascularity than the control group.

To study the thyroid regeneration and injury of recurrent laryngeal nerve after irreversible electroporation (IRE). 12 pigs were divided into three groups: six pigs underwent IRE, other pigs were used as controls. IRE was performed near tracheoesophageal groove, to ablate most part of thyroid gland. Parathyroid and thyroid function, recurrent laryngeal nerve injury and thyroid computed tomography (CT) imaging were regularly investigated. The histopathology results were analyzed to detect thyroid regeneration. Masson's trichrome method for collagen and immunohistochemistry were performed for Soluble protein-100 (S100) and neurofilaments on nerve section. In IRE group, there were no symptoms of recurrent laryngeal nerve-related injury. No abnormalities of recurrent laryngeal nerve were shown on hematoxylin-eosin (HE) staining, Masson's trichrome staining, Neurofilament (NF) staining and S100 staining. There were no significant changes for thyroid and parathyroid function in all pigs. Immediately after IRE, CT showed hypoattenuation in the ablated thyroid gland and it became swelling. 14 days after IRE, thyroid CT showed hetergenous attenuation in the electroporation zone, and the size and attenuation of thyroid gland were normal after two months. There was cell apoptosis in the thyroid gland after IRE. Seven and 14 days after IRE, there was fragmentation of nucleus within the follicle, and some follicles were empty. Two months later, complete regeneration of thyroid tissue was shown. IRE was shown to be both effective and safe with complete regeneration of thyroid tissue and preservation of the function and structure of the recurrent laryngeal nerve.

Purpose this study was conducted between 2015 and 2017 at the faculty of Veterinary Medicine Cairo University and aimed to histologically compare between the effect of processed and unprocessed bone marrow aspirate on the quantity and quality of the newly formed bone. Material and methods 72 Adult male white New Zealand rabbits were used in this study and were divided equally into 2 groups A and B. Bone defects were created at the medial upper extremity of right and left tibia of all rabbits down to bone using trephine bur measuring 4 mm diameter to 5 mm depth. The bone defects were treated with whole bone marrow loaded on gel foam in group A right tibias and using mononuclear cell layer obtained through centrifuging bone marrow loaded on gel foam in group B right tibias. Bone defects in the left tibias in all rabbits treated with gel foam only. Scarification was done 3,6,12 weeks postoperative. Histological evaluation was done using H and E and Masson’s trichrome stain. Results the results showed that group A right tibial defects had better healing results than group B right tibial defects all over the follow-up period, but the differences were not statically significant. Conclusion the study findings indicate that the centrifugal concentration techniques provide better therapeutic outcomes over the whole BMA, but it is statistically insignificant.

Background: Non-cirrhotic portal hypertension includes primarily the cases of non-cirrhotic portal fibrosis (NCPF) and extra-hepatic portal vein obstruction, both showing no significant parenchymal disease. NCPF is a disease of uncertain etiology characterized by periportal fibrosis with small and medium sized portal vein branches involvement. Most study done in the past are clinical, hence we decided to analyse its histo-pathological features with clinical correlation. Methods: This is four and half year’s retrospective study of liver biopsies of clinically diagnosed patients of NCPF. Fifteen patients of EHPVO served as controls. Biopsies were stained with Haematoxylin and eosin, reticulin and Masson’s trichrome. Result: Total 978 benign specimens (excluding space-occupying lesions) received, of which 45 diagnosed as NCPF (4.6%). Most patients (68.88%) were between 20-40 years (M:F ratio 1:1.5), commonest symptoms long-standing splenomegaly (95%), anemia (78%) and variceal bleeds (42%) with duration varying from 3 months to 11 years. Commonest histopathological findings were portal tract fibrosis (77.78%), phlebosclerosis (68.89%), portal tract remnants (24.44%), more frequently seen in NCPF than EHPVO, septal fibrosis was seen in five. Follow up available in 91.11% cases showed no mortality or liver failure, but significant morbidity due to repeated episodes of variceal bleeds and hypersplenism. Conclusion: NCPF is still common in India. Distinctive features on histology are portal tract fibrosis, portal tract remnants, with phlebosclerosis which is being statistically significant when comparing with EHPVO. NCPF can show advanced fibrosis, can also present with features of decompensation mimicking cirrhosis, however its course remains stable with good long-term survival.

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.

Prolonged healing and scar formation are two major challenges in treating wounds, resulting in physical, mental and socioeconomic burden to the affected subjects. Exosomes (Exos) are naturally secreted vesicles playing a vital role in the transfer of RNA to neighboring or distant recipient cells and there are numerous on-going studies on their role of in tissue regeneration and wound healing.Aim of the work: Evaluation of the efficiency of Exos derived from adipose stem cells on cutaneous wound healing in adult male albino rats.Materials and Methods: Thirty-four rats were divided into three groups; Group I: control. Group II: a dorsal cut wound (1.5 × 1.5 cm) was done to each rat. Group III: the same wound was done and rats were injected with 200μ PKH26-labeled Exos intravenously. Rats were sacrificed after 21 days. Skin sections were subjected to hematoxylin and eosin (HandE), Mallory's trichrome and orcein stains, alpha smooth muscle actin (α-SMA) and Ki67 immunohistochemical stains.Results: Skin sections from group II showed scar formation, lost epidermal layers and hair follicles. There was significant increase in collagen fibers and α-SMA immunoreaction, with significant decrease in elastic fibers and Ki67 immunoexpression compared to the control. In group III, the skin revealed almost normal histology, significant decrease in collagen fibers and α-SMA, with significant increase in elastic fibers and Ki67 immunoreaction compared to group II.Conclusion: Exos injection has demonstrated a beneficial effect on cutaneous wound healing and could prevent scar formation in adult male albino rats.

The corpus spongiosum reportedly occupies a larger proportion of the human glans penis than does the penile body, embedding the end of the corpus cavernosus (CC). However, anatomic descriptions about the fibrous structures of glans penis in the literature cause confusion during dissection and reconstructive surgery. Forty-five penises of formalin-embalmed cadavers were dissected sagittally along the course of the distal urethra and observed macroscopically. Dense connective tissues adjacent to the fossa navicularis and spongiosum parts of the glans were cropped, and underwent Masson's trichrome and Verhoeff-Van-Gieson staining. Most (55.5%) of the specimens had distinct fibrous bands toward the distal tips of the glans penis, which elongated from the tunica albuginea of the CC. They comprised longitudinal collagen bundles continuous to the outer longitudinal layer of the tunica albuginea covering the CC and were intermingled with sparse elastic fibres. This architecture either did not reach the distal end of the glans penis (35.5% of cases), or was obscure or dispersed in all directions (9.0% of cases). The structural dimorphism and the variations in the ratio of dense connective tissue components of the fibrous skeleton are considered to contribute to the varying degrees of flexibility, distensibility and rigidity of the human glans penis.

Objectives It is challenging to separate peritumoral fibrosis from fibrosis due to chronic liver disease in mass-directed liver biopsies. We evaluated the distance that peritumoral fibrosis extends from metastatic colorectal adenocarcinoma in liver. Methods Peritumoral and distant uninvolved liver trichrome stains from 25 cases were analyzed using digital image analysis. Fibrosis was quantitated at concentric intervals from each tumor and in uninvolved liver. Results There was a 3.9 fold (range 0.9-18.6) median increase in fibrosis in the first 0.5 mm of peritumoral liver compared to distant liver. Fibrosis levels returned to baseline at median 2.5 mm (interquartile range 1.5-5.0 mm) from tumor. Conclusions Fibrosis is markedly increased in peritumoral liver. Fibrosis levels returned to baseline by 5 mm from tumor in approximately 75% of cases. Pathologists should be cautious of fibrosis in mass-directed liver biopsies without at least 5 mm of liver tissue distal to the mass.

While therapies using mesenchymal stem cells (MSCs) to treat rotator cuff tendon tear (RCTT) have yielded some promising preliminary results, MSCs therapy has not yet completely regenerated full-thickness RCTT (FTRCTT). It has recently been reported that polydeoxyribonucleotide (PDRN) is effective in the treatment of chronic rotator cuff disease. We hypothesized that local injection of human umbilical cord blood-derived (UCB)–MSCs with PDRN would be more effective in regenerating tendon tear than UCB-MSCs alone. The purpose of this study was to evaluate the effects of UCB–MSCs combined with different doses of PDRN on the regeneration of RCTT in a chronic RCTT model by using a rabbit model. New Zealand white rabbits ( n = 24) with FTRCTT were allocated randomly into three groups (8 rabbits per group). Three different injectants (G1-S, 0.2 mL UCB-MSCs; G2-P1, 0.2 mL UCB-MSCs with one injection of 0.2 mL PDRN; G3-P4, 0.2 mL UCB-MSCs, and four injections of 0.2 mL PDRN per week) were injected into FTRCTT under US-guidance. After the rabbits were euthanized, we evaluated ross morphological and histological change. Motion analysis was also performed. There were significant differences in gross morphological changes between before, and at 4 weeks after injection, in all three groups, but no differences were found among the three groups. Masson’s trichrome (MT) or anti-type 1 collagen antibody (COL-1)-positive cell densities in G2-P1 and G3-P4 were improved significantly compared with those in G1-S, but showed no significant difference between G2-P1 and G3-P4. On motion analysis, walking distance and fast walking time in G2-P1 and G3-P4 were significantly longer/higher than those in G1-S, but showed no significant differences between G2-P1 and G3-P4. These results demonstrated that there was no significant difference in the gross morphologic change of tendon tear between UCB-MSCs only and combination with PDRN injection in rabbit model of chronic traumatic FTRCTT. Furthermore, there were no significant differences in the regenerative effects between high and low doses of (0.8 and 0.2) mL of PDRN.

Aim: the aim of the present investigation was to study the effect of mesenchymal bone marrowstem cells (BMSCs) on the histological structure of the dental pulp of rats with Streptozotocininduceddiabetes mellitus using hematoxylin & eosin and Masson’s trichrome stains. Furthermore,the apoptotic activity of dental pulp cells was detected through Caspase-9 localization.Material and Methods: Thirty, three- month old, adult male albino rats, 180-200 gram weightwere used in this investigation. They were divided into three groups. Group I: 10 animals servedas controls. Group II: 10 animals subjected to a single intraperitoneal injection of Streptozotocinin a dose of 60 mg/kg body weight for induction of diabetes mellitus that was verified by detectinghyperglycaemia (200mg/dl or more) in a blood sample. Group III: 10 animals treated the sameas those of group II and one week later these animals were intravenously injected with BMSCs.At the end of the experiment which lasted for two months, the animals of the different groupswere euthanized. Their jaws were dissected, decalcified and processed for hematoxylin and eosin,Masson’s trichrome stains and immunohistochemical stain for caspase-9 to study the dental pulpof the experimental rats.Results: The present work revealed that the treatment of diabetic rats with single intravenousinjection of bone marrow stem cells resulted in amelioration of the degenerative and apoptotic.effect of diabetes on the pulpal tissuesConclusion: Induction of diabetes in albino rats has degenerative effect on pulpal tissuesaccompanied by increased expression of caspase-9 (apoptotic marker). Intravenous injectionof BMSCs resulted in regeneration of pulp tissues and decreased apoptosis as detected by lowcaspase-9 expression.

Free living amoebas (FLA) are opportunistic pathogen found in different water sources in the environment. The aim of this study was to investigate the prevalence of free living amoeba in different samples of domestic water reserves (DWR) in Sfax region from Tunisia. It was a prospective study dealing with 486 water samples collected from different DWR. After filtration through a cellulose acetate membrane samples were cultured on non-nutrient agar and the FLA were detected and strained with Giesma, Trichrome and red nuclear stain for morphological and morphotypic studies. FLA were found in 62% of samples. The Acanthopodial morphotype was detected in 43%, Polytactic (38%), Monotactic (28%), Fan-shaped (17%), Rugose (11%), Dactilopodial (10%) and Eruptive (9%). These results demonstrate that domestic water reserves are a significant source of the FLA and maintenance of DWR is recommended.

The aim of this study was to evaluate the effect of different storage media on angiogenesis and maintaining autogenous bone graft volume in rabbits.Two grafts were removed bilaterally from the calvaria of 18 rabbits. One graft was removed and immediately fixed in the right mandibular angle (control group). The other graft was stored for 30 minutes in 1 of the following storage media (n = 6): saline solution (saline group), air exposure (dry group), or platelet-poor plasma (PPP group) and then retained by a screw in the right mandibular angle in the same animal. Four weeks later the animals were euthanized, and the grafted areas were harvested, fixed in 10% phosphate buffered formaldehyde solution, and embedded in paraffin. The 5-μm semi-serial sections were stained in hematoxylin and eosin and Mallory trichrome.Histologic analysis of all groups showed the bone graft was vascularized and well incorporated into the recipient site. The number of blood vessels decreased in the saline and dry groups compared with the control group (P .03); in contrast, the number of blood vessels increased in the PPP group (P .05). There were fewer osteoclasts in the saline group compared with the control group (P .05). Furthermore, the saline group showed larger numbers of blood vessels than the dry group (P .01). The PPP group showed larger bone graft volumes compared with the dry and saline groups (P .01). In addition, the saline group showed larger bone graft volumes than the dry group (P .01).PPP improved angiogenesis, maintained the volume of the autogenous bone graft, and was a better storage medium during the trans-surgical period than the dry and saline media.

This study aimed to evaluate the potential cytotoxic effect of oral administration of silver nanoparticles (Ag-NPs) on adult albino rats’ pulp tissue; due to the enormous uses of Ag-NPs in the medical and dental field. The Ag-NPs were synthesized via the green process using peels of pomegranate extract. The pomegranate-mediated Ag-NPs were subjected to morphological and spectral analysis through ultraviolet visible absorption spectra, transmission electron microscopy, Fourier transforms infrared, Zeta-potential measurements, and energy dispersive X-ray spectroscopy. The structural and morphological characterization techniques confirmed the proper synthesis of biosynthesized Ag-NPs with a size around 20 ​nm and the surface plasmon resonance peak within 400–450 ​nm. The oral cytotoxic effect of Ag-NPs was assessed through detecting the histological (hematoxylin & eosin, Masson’s trichrome) and immunohistochemical (vascular endothelial growth factor (VEGF), Caspase-3 proteins) variations. The data was analyzed statistically through using the SPSS software. Dental pulp tissues of albino rats-treated with Ag-NPs revealed that most of the odontoblasts with marked hydropic degeneration, vacuolization of their cytoplasm, loss of organization and apoptosis. Marked vasodilatation and cognition of blood vessels were detected. There was weak to moderate positive reactivity to Masson’s trichrome stain. There was statistically significant decrease in the expression of VEGF in the treated group and highly statistically significant increase in the expression of Caspase-3 in comparison with the control group. Conclusion Oral administration of Ag-NPs induced size and dose-dependent structural changes in the pulp tissue of adult male albino rats.

AIMS Machine learning in digital pathology can improve efficiency and accuracy via prescreening with automated feature identification. Studies using uniform histological material have shown promise. Generalised application requires validation on slides from multiple institutions. We used machine learning to identify glomeruli on renal biopsies and compared performance between single and multiple institutions. METHODS AND RESULTS Randomly selected, adequately sampled renal core biopsy cases (71) consisting of four stains each (haematoxylin and eosin, trichrome, silver, periodic acid Schiff) from three institutions were digitised at ×40. Glomeruli were manually annotated by three renal pathologists using a digital tool. Cases were divided into training/validation (n = 52) and evaluation (n = 19) cohorts. An algorithm was trained to develop three convolutional neural network (CNN) models which tested case cohorts intra- and inter-institutionally. Raw CNN search data from each of the four slides per case were merged into composite regions of interest containing putative glomeruli. The sensitivity and modified specificity of glomerulus detection (versus annotated truth) were calculated for each model/cohort. Intra-institutional (3) sensitivity ranged from 90 to 93%, with modified specificity from 86 to 98%. Interinstitutional (1) sensitivity was 77%, with modified specificity 97%. Combined intra- and inter-institutional (1) sensitivity was 86%, with modified specificity 92%. CONCLUSIONS Feature detection sensitivity degrades when training and test material originate from different sites. Training using a combined set of digital slides from three institutions improves performance. Differing histology methods probably account for algorithm performance contrasts. Our data highlight the need for diverse training sets for the development of generalisable machine learning histology algorithms.

We virtually generate multiple histological stains through a single deep-neural-network, using at its input autofluorescence images of the unlabeled tissue alongside a user-defined digital-staining-matrix. By feeding this digital-staining-matrix to the network, the user indicates which stain to apply on each pixel or region-of-interest, enabling virtual blending of multiple stains according to a desired micro-structure map. We demonstrated this technique by applying combinations of different stains (H&E, Masson’s Trichrome and Jones silver stain) on blindly-tested, unlabeled tissue sections. This technology avoids the histochemical staining process and enables newly-generated stains and stain-combinations to be used for inspection of label-free tissue microstructure.

Background: Chronic tubulointerstitial fibrosis is a common final pathway leading to end stage kidney disease in cats and has no effective treatment. The use of cell-based molecules to treat kidney fibrosis may be a promising approach. The objectives were to test the effects of intra-renal chemokine CXCL12 injection in a pre-clinical cat model of unilateral ischemia/reperfusion (I/R)-induced kidney fibrosis and then, within a clinical pilot study, test the safety/feasibility of CXCL12 injection in cats that might have early chronic kidney disease (CKD).Methods:Pre-clinical: Thirty cats received intra-renal injection of 100, 200, or 400 ng of recombinant human CXCL12, or sterile saline, into the I/R kidney 70 days post-injury, or were non-injured, non-injected controls (n = 6/group). Kidney collagen content was quantified 4 months post-treatment using Masson's Trichrome and Picrosirius Red (PSR) stained tissues. In a separate study (n = 2) exploring short-term effects of CXCL12, 200 ng CXCL12 was injected into I/R kidneys and then harvested either 30 min (n = 1) or 1 month (n = 1) post-injection. Kidney concentrations of CXCL12, matrix metalloproteinase 1 (MMP-1), and lysyl oxidase-like enzyme 2 (LOXL-2) were quantified via ELISA. Clinical Pilot: 14 client-owned cats with potential early kidney disease received a single-treatment, bilateral intra-renal injection of 200 ng CXCL12 (n = 7), or received no injection (n = 7). Blood/urine samples were collected monthly for 9 months to assess renal function and CKD staging.Results:Pre-clinical: I/R increased the affected kidney collagen content, which both mid and high doses of CXCL12 restored to normal (ps < 0.05 vs. untreated). I/R increased collagen fiber width, which both mid and high doses of CXCL12 restored to normal (p < 0.001 vs. untreated). Early changes in kidney MMP-1, associated with collagen breakdown, and subsequent decreases in LOXL-2, associated with collagen cross-linking, in response to CXCL12 treatment may contribute to these findings. Clinical Pilot: Bilateral intra-renal injection of CXCL12 using ultrasound guidance in cats with CKD was feasible and safe in a general practice clinical setting with no obvious side effects noted during the 9-month follow-up period.Conclusions: Intra-renal injection of CXCL12 may prove to be an effective treatment for kidney fibrosis in cats with CKD. Additional mechanistic and clinical evaluations are needed.

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.

To study the features of reparative dentinogenesis when using biomaterials during accidental opening of the pulp in an experiment.In an experiment on 3 dwarf pigs of the Svetlogorsk breed a traumatic opening of the tooth cavity was carried out in 18 teeth (6 molars, 6 premolars, 6 incisors), then Biodentin (9 teeth) and Aurosil (9 teeth) were applied on the opened pulp horn. After 14, 30 and 60 days, dwarf pigs were euthanized, and jaw blocks taken. Histological examination of the teeth was carried out using the method of embedding in methyl methacrylate (OsteoBead, Sigma, USA). Thin sections of 35-50 microns were made from the samples and stained using the «Heavenly Trichrome» method. Documentation was carried out using a Leica DM100 microscope and an EC3 digital camera in reflected and transmitted light.Histological examination of the thin sections of teeth stained according to the «Heavenly Trichrome» method revealed that 2 weeks after the pulp opening and applying of Aurosil and Biodentin, comparable processes occurred in the tooth cavity. Thus, the proliferation of odontoblasts was noted on the lateral surfaces of the chamber. New dentin layers and odontoblasts spread over the lower surface of the material with the formation of dentinal bridges by 4 weeks of observation in both groups. By week 12, there was a decrease in the proliferative activity of odontoblasts and dentin maturation in all cases. At the initial stages of the experiment, some pulp edema and moderate plethora were noted, which was no longer observed at 4 weeks. By 12 weeks, because of the proliferation of odontoblasts, the volume of the tooth cavity was reduced by a third. By the 6th month in the experiment, no significant changes in the structure of dentin and pulp were revealed.According to morphological data the studied biomaterials (Aurosil and Biodentin) contribute to dentin regeneration equally.Изучить особенности репаративного дентиногенеза при применении биоматериалов при случайном вскрытии пульпы в эксперименте.В эксперименте на 3 карликовых свиньях светлогорской породы проведено травматическое вскрытие полости зуба, в области 18 зубов (из них 6 моляров, 6 премоляров, 6 резцов) на вскрытый рог пульпы наносили препарат Биодентин (9 зубов) и Ауросил (9). Через 14, 30 и 60 дней карликовые свиньи подвергались эвтаназии, был проведен забор блоков челюстей. Гистологическое исследование зубов карликовых свиней проводили с использованием метода заливки в метилметакрилат (OsteoBead, «Sigma», США). Из образцов изготавливали шлифы толщиной 35—50 мкм и окрашивали по методу «Небесный трихром». Документирование осуществлялось с помощью микроскопа Leica DM100 и цифровой камеры EC3 в отраженном и проходящем свете.Гистологическое исследование шлифов зубов, окрашенных по методу «небесный трихром», выявило, что через 2 нед после вскрытия пульпы и нанесения биоактивных цементов в полости зуба происходили сопоставимые процессы. Так, на боковых поверхностях камеры отмечали пролиферацию одонтобластов. Напластования нового дентина и одонтобласты распространялись по нижней поверхности материала с образованием дентинных мостиков к 4 нед наблюдения в обеих группах. К 12 нед отмечали снижение пролиферативной активности одонтобластов и созревание дентина во всех случаях. На начальных этапах эксперимента отмечали некоторый отек пульпы и умеренное полнокровие, которое уже не наблюдалось на сроке 4 нед. К 12 нед в результате пролиферации одонтобластов произошло сокращение объема полости зуба наПо данным морфологического исследования установлено, что биоматериалы Ауросил и Биодентин способствуют регенерации дентина в равной степени.

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.