Morphological, Glycohistochemical, and Immunohistochemical Studies on the Embryonic and Adult Bovine Testis

In the present study, the testes of 32 bovine embryos with different crown-rump length (2.5- 90 cm CRL) and of 15 sexually mature bulls (Deutsches Fleckvieh) were investigated using light- and electron microscope as well as glycohistochemical and immunohistochemical methods. The gestation period was divided into 3 stages; early, mid, and late gestation. Developmental changes in the testicular morphogenesis were therefore analyzed in details during these phases. Generally, embryonic development of bovine testis involves the same mechanism described in other mammals. At the first stage of this study (2.5 cm CRL/43 dpc), the anlage of the testes protruded to the coelomic cavity as paired bean-shaped structures on either side of the dorsal mesentery medial to the mesonephros. It consists of primitive testicular cords, interstitium, and rete testis blastema. Proceeding with fetal age, these basic testicular structures are further differentiated. The tunica albuginea is separated into two layers: an outer fibrous layer (tunica fibrosa) with some mesenchymal cells, numerous fibroblast, and much fibrous content and an inner cellular layer with several blood vessels (tunica vasculosa). The testicular cords are surrounded by a marked basal lamina and peritubular cells and lined by two types of cells: a large number of dark polygonal cells with irregular nuclei, pre-Sertoli cells and small number of large light round cells with relatively round nuclei, the prespermatogonia. The average number of the germ cells per cross section of cord increases, particularly form 3.5 to 14 cm CRL, resulting in a germ cell maximum at the end of this stage (14 cm CRL). Although most of the germ cells are located toward the periphery of the cord, some are also found in the center. Pre-Sertoli cells form a complete layer at the periphery of the cords. Generally, these cells are irregular in shape and numerous but considerably smaller than the germ cells. Unlike prespermatogonia, mitotic figures are seen in pre-Sertoli cells during the whole embryonic life. As a consequence of the expansion in the interstitium, the seminiferous cords are progressively separated from each other. The testicular interstitium is rapidly differentiated and is composed of several islets or clusters of polygonal Leydig cells, peritubular flattened cells surrounding the testicular cords, connective tissue cells, and numerous blood vessels. In the present study, fetal Leydig cells were first recognized at 3.5 cm CRL. Thereafter, the average number of these cells is rapidly increased to attain their maximum with the end of the first gestation period (14 cm CRL). This generation of Leydig cells however dedifferentiates progressively with developmental age. A continuous system of basal lamina joins the testicular cords with rete strands from 10 cm CRL and onwards. This system establishes the first connection between these two testicular components via ill-developed uncanalized straight tubules (tubuli recti). Rete testis channels are lined by simple layer of cuboidal epithelium with round nuclei occupying most of the cytoplasm and enclosed by well-defined basal lamina. The adult bovine testis is enclosed by a connective tissue capsule, tunica albuginea, composed predominantly of collagen fibers and few elastic fibers. Most of the testicular parenchyma is made up of the convoluted seminiferous tubules (tubuli seminiferi contorti), two-ended convoluted loops, with both ends opening into the rete testis via specialized terminal segments. The seminiferous tubules of sexually mature bulls are enclosed by a distinct lamina propria and are lined by two cell populations, non-proliferating Sertoli cells and highly proliferating spermatogenic cells. The bovine lamina propria consists of basal lamina, collagen and elastic fibers, and 3-5 layers of partially overlapping myofibroblasts. Additionally, fibrocytes, collagen fibrils, and fibroblasts-like cells form the outermost border of the tubulus. Sertoli cells are easily identifiable elements of the seminiferous epithelium. Adult Sertoli cells are large irregularly shaped cells with their broad bases resting on the basal lamina while the remaining cytoplasmic processes extend upward to the tubular lumen. They are characterized by round or oval euchromatin-rich nuclei situating in the basal portion near the basal lamina of the seminiferous tubules. Adult bovine germ cells are present in four morphologically different groups, i.e., spermatogonia, spermatocytes, spermatids, and spermatozoa. The seminiferous cycle stages are identified using changes in the germ cell nuclei as well as location and shape of spermatids. According to this method, eight stages are defined in the seminiferous epithelium of bovine. The interstitial or intertubular tissue of adult bovine testis consists of Leydig cells, macrophages, scattered lymphocytes and plasma cells, and contains numerous blood and lymph vessels. Not all Leydig cells have contact to blood or lymph capillaries. The excurrent duct system of the adult bovine testis consists of terminal segment of the convoluted seminiferous tubules, straight tubules, and rete testis. The terminal segment can be further subdivided into a proximal (transitional) region, middle portion, and distal part (terminal plug). The proximal region is lined by typical Sertoli cells while the last two parts are lined by modified Sertoli cells. The tubulus rectus of adult bovine testis is composed of three morphologically different regions: a proximal cup-shaped region, a middle narrow stalk, and a distal festooned portion. The rete testis is a complicated centrally positioned meshwork of intercommunicating channels that lies within the mediastinum testis parallel to the long axis of epididymis. The simple cuboidal epithelium of straight tubules and rete testis is shown to contain some lymphocytes and macrophages. The cellular distribution of glycoconjugates within the fetal and adult bovine testis was investigated using thirteen (ConA, PSA, LCA, PNA, GSA-I, ECA, DBA, SBA, HPA, VVA, WGA, UEA-I, LTA) different fluorescein isothiocyanate (FITC) conjugated lectins. In fetal testes, detection of sugar moieties by lectins was carried out on Bouin õ s-fixed paraffin-embedded sections while in adult it was performed on both Bouin õ s-fixed paraffin-embedded and acetone-fixed frozen sections. Only five lectins (PSA, PNA, GSA-I, DBA, WGA) showed a positive reaction in the embryonic testes. PNA, GSA-I, DBA, and WGA were detected in the germ cells whereas PSA, DBA and WGA labeled the fetal Leydig cells. None of the lectins used was observed in the pre-Sertoli cells. Further on, some lectins were seen in tunica albuginea (PSA, PNA, GSA-I, WGA), basal lamina of testicular cords (PSA, WGA), interstitial blood vessels (PSA, GSA-I, WGA), mediastinum testis (PSA, PNA, WGA) and rete testis epithelium (PNA). In adult animals, spermatogonia and spermatocytes were positively stained with PSA, LCA, DBA, SBA, and VVA. All the lectins investigated except that of the fucose-binding lectin (UEA-I and LTA) were definitely detected in the acrosome of round and elongated spermatids. These results indicate a role for carbohydrates in spermiogenesis. Apical Sertoli cells processes and Leydig cells were weakly stained with PSA and LCA as well. DBA binding sites were also seen in the Leydig cells. Immunohistochemical studies were performed using the Avidin-Biotin-Peroxidase Complex (ABC) method for localization of fibroblast growth factor-1 (FGF-1), fibroblast growth factor-2 (FGF-2), S-100, laminin, alpha-smooth muscle actin (á -SMA), vascular endothelial growth factor (VEGF), connexin 43 (Cx43), CD4, CD8, CD68, angiotensin-converting enzyme (ACE), and galactosyltransferase (GalTase) in the bovine testis. The expression of FGF-1 and FGF-2 was further investigated in the adult bovine testis using in situ hybridization and PCR. Immunohistochemically, FGF-1 was seen in the Sertoli cells, Leydig cells, endothelium of the blood vessels, and epithelium of straight tubules and rete testis of fetal and adult testis. It was additionally detected in spermatogonia and spermatids of sexual mature animals. FGF-2 exhibited a striking positive reaction in fetal (from 6 to 30 cm CRL) and adult Leydig cells. Moreover, it showed marked reaction in the endothelium of blood vessels and in the epithelium of tubulus rectus and rete testis. FGF-2 was also localized in some spermatogonia, and myofibroblasts. By means of in situ hybridization, FGF-1 and FGF-2 mRNA were found in Leydig and Sertoli cells as well as in the modified Sertoli cells of the terminal segment. FGF-1 transcripts were additionally recognized in the straight tubules and rete testis epithelium. Distinct S100 immunostaining was observed in the Sertoli cells, endothelium of blood vessels and in the rete testis epithelium of fetal and adult testis. Laminin was localized to the basal lamina of seminiferous tubules, blood vessels, myofibroblasts, and rete testis. Although á -SMA was detected in smooth muscle cells of the blood vessels, no immunoreactivity was seen in the peritubular cells during the whole gestation period. The myofibroblasts surrounding the seminiferous tubules and rete testis showed intense positive reaction for á -SMA in the adult testis. VEGF was detected in the acrosomes of the elongating spermatids. Connexin 43 was localized to gap junctions between Leydig cells in the fetal and adult life as well as to the seminiferous epithelium apical to spermatogonia and basal to spermatocytes, a position correlating with Sertoli-Sertoli cell junctions. The detection of cells positive for CD4, CD8, CD68 within the adult testis interstitium clearly indicate the presence of lymphocytes and macrophages within this testicular compartment. GalTase showed striking positive reaction in the Golgi complex of Sertoli cells, Leydig cells, and some spermatocytes as well as at the cell membrane of elongating spermatids and in the simple cuboidal epithelium of rete testis. ACE positive reaction was found in the prespermatogonia (only at 6-10 cm CRL) and in fetal and adult testicular blood vessels. The functional significance of these immunocytochemically-demonstrated proteins is discussed

Surgical Management and Outcome of Patients with Thyroid Diseases during the COVID-19 Pandemic

Background: Globally, healthcare authorities have reacted by restricting medical care to emergency cases and postponing elective surgical procedures of all types. Elective surgeries have been almost totally postponed keeping to a minimum the risk of transmission of SARS-CoV-2 and to allow a better allocation of resources. Since thyroid surgery usually does not cover immediate surgical interventions, during this pandemic nearly all the patients who require thyroid surgery care are experiencing delays in the operation planning procedure. Objective: The aim of the work was to investigate the impact of COVID-19 pandemic on thyroid surgery.Subjects and methods: We conducted prospective study from August 2020 to February 2021 among all patients who attended to Damanhur Medical National Institute Hospital. Total sample size was 60 patients and patients with positive COVID-19 were enrolled in group 1 (cases) and those who were negative COVID-19 were enrolled in group 2 (controls). Results: A total of 60 patients were seen during the study period. Males were 35 (58.3%) while females were 25 (41.7%). The mean age 39.5± 9.5 years. There is no statistically significant difference between patients with COVID-19 and patients without COVID-19 regarding developing complications from thyroid surgeries.Conclusion: The COVID-19 pandemic has created a complicated set of issues for patients with endocrine-related cancers. Most thyroid cancers are indolent and so postponement of surgery in the era of COVID-19. There is no significant association between COVID-19 and complications of surgery

Postoperative Outcomes after Laparoscopic Cholecystectomy in Elderly Patients

Background: Age is a critical factor influencing the death and morbidities afterward cholecystectomy. It was revealed that laparoscopic cholecystectomy (LC) in the elderly has similar security and effectiveness to those in young people. The gold-standard in the management of cholelithiasis is LC. Objectives: This work was aimed to assess post-operative outcomes of LC between elderly cases and for comparing the post-operative outcome of LC among ages groups >60yrs and <60yrs. Patients and Methods: This prospective study conducted among 68 cases who experienced LC at Damanhur Medical National Institute Hospital. Cases were allocated into 2 groups according to ages: elderly (≥60yrs, n= 38) and young (<60 yrs, n= 35). Results: 68 cases divided into two groups. The mean age is 68.4± 5.7 and 49.1± 10.5 among elderly and younger groups. There is significant difference between groups regarding age and comorbidities. The mean operative time was 62.8± 21.2 and 59.5± 18.7 among elderly and young groups. There is no statistically significant difference between two groups regarding intraoperative data. There is nonsignificant change regarding postoperative outcomes after LC. Conclusion: It could be concluded that laparoscopic cholecystectomy in elderly cases is similar with young cases. Consequently, LC is secure even in the elderly

Distinct cytokine patterns in Occult Hepatitis C and Chronic Hepatitis C Virus Infection

Background & Aim: 
The immunopathogenesis of chronic hepatitis C virus (HCV) infection is a matter of great controversy. The imbalance of T-helper lymphocyte cell cytokine production was believed to play an important pathogenic role in chronic viral hepatitis. Occult hepatitis C infection is regarded as a new entity that should be considered when diagnosing patients with a liver disease of unknown origin. The aim of this study was to determine serum T-helper 1 and T-helper 2 cytokine production in patients with occult HCV infection and its role in pathogenesis versus chronic viral hepatitis C infection.

Methods: 
Serum levels of cytokines of T-helper 1 (IL-2, IFN-[gamma]) and T-helper 2 (IL-4) were measured in 27 patients with occult HCV infection and 50 patients with chronic hepatitis C infection.

Results: 
The levels of the T-helper 1 cytokines, IL-2 and IFN-[gamma], were highly and significantly increased in patients with chronic HCV infection as compared with occult HCV infection (p<0.001). The T-helper 2 cytokine IL-4 was highly and significantly increased in occult HCV infection as compared with chronic HCV infection (p<0.001). Necroinflammation (P<0.001) fibrosis (P<0.001) and cirrhosis (P =0.03) were significantly increased in chronic HCV than occult HCV. 

Conclusion: 
Patients with occult HCV infection exhibited distinct immunoregulatory cytokine patterns, favoring viral persistence in the liver in spite of its absence from peripheral blood and explaining the less aggressive course of this disease entity than chronic hepatitis C virus infection

Fixed gonadotropin-releasing hormone antagonist protocol versus gonadotropin-releasing hormone agonist long protocol in patients with polycystic ovary syndrome treated for intracytoplasmic sperm injection cycles

Background: Women with polycystic ovary syndrome (PCOS) are at risk of developing ovarian hyperstimulation syndrome (OHSS) during ovarian stimulation. Use of GnRH antagonist in the general sub fertile population is associated with lower incidence of (OHSS) than agonists and similar probability of live birth but it is unclear. Our Objective was to compare the fixed GnRH antagonist and GnRH agonist long protocols in patients with PCOS undergoing IVF.Methods: In this randomized controlled trial (RCT), 200 patients with PCOS were randomly allocated in two groups: long GnRH (n = 100) and fixed GnRH antagonist protocol (n = 100).           Results: There is significant difference was observed in chemical pregnancy rate (46.0% versus 31.0%), and clinical pregnancy rate (43.0% versus 29.0%) in agonist and antagonist protocols, respectively. Duration of stimulation was significantly higher in agonist group (13.58 versus 12.381 days), respectively.  Total number of ampoules of gonadotrophin is comparable in both groups (t=1.914, p=0.057).Conclusions: The use of GnRH antagonists is more advantageous than GnRH agonists in relation to shorter duration of stimulation thus allowing a reduction in the treatment time that makes COS less costly and better patient compliance. In this study GnRH agonist shows higher pregnancy rate than antagonist, so larger studies needed to clarify their roles

Effect of induced hyperthyroidism on pancreas of adult female albino rats

Introduction: A condition known as hyperthyroidism can lead to a wide range of health issues, including osteoporosis, oxidative liver damage, diabetes mellitus, as well as cardiovascular disease.Objective: To examine the detrimental effects of hyperthyroidism on the pancreatic structure of adult female albino rats.Materials and Methods: twenty-one adult virgin female albino rats were assigned to 2 groups; control and induction of hyperthyroidism. Serological analysis to assess thyroid functions, oxidative stress and lipid peroxidation analysis was carried out. Pancreatic samples were processed for light microscopic examination.Results: Hyperthyroidism caused both biochemical and histological changes on pancreas of adult female albino rats. The biochemical changes in the form of significant decreased in the thyroid stimulating hormone serum level, significant reduction in level of serum superoxide dismutase and the malondialdehyde level was significantly increased in the hyperthyroid rats. The histological changes were loss of its general architecture. The pancreatic acini appeared irregular, vacuolated and had dark nuclei. Some islets of Langerhans appeared shrunken and the acini did not have the normal smooth demarcation from the surface.Conclusion: Hyperthyroidism seriously affected the histological structure of the pancreas with subsequent disturbance in the biochemical markers

Validation of a method to elute viruses from different types of face masks

Due to the SARS-CoV-2 pandemic, it is crucial to study the efficiency of face masks in retaining viruses for the upcoming years. The first objective of this study was to validate a method to elute viruses from polyester and cotton face masks. We observed that deionized water followed by 3% beef glycine (pH 9.5 or pH 7.2) was significantly more efficient (p < 0.05) in eluting the bacteriophage phiX174 virus from polyester (4.73% ± 0.25% to 28.67% ± 1.89%), polyester/cotton (3% ± 0.33%), and cotton (1.7% ± 0.21%) face masks than 3% beef glycine only (pH 9.5 or pH 7.2) as a single eluent (3.4% ± 0.16% to 21.33% ± 0.94% for polyester, 1.91% ± 0.08% for polyester/cotton, and 1.47% ± 0.12% for cotton face masks). Also, deionized water was significantly less efficient as a single eluent for eluting bacteriophage phiX174 from all the studied face mask types. The polyethylene glycol (PEG) precipitation method was substantially more efficient (p < 0.05) as a second step concentration method for the viruses in the eluates than the organic flocculation (OF) method. Higher viral loads were eluted from polyester face masks than cotton ones. We also found varying viral loads in the eluate solutions from different commercial polyester face masks, with the highest percentage seen for the N95 face mask. The second objective was to apply the validated method to study the effect of autoclaving on the different face mask materials. Results of the study did not show any significant differences in the viral loads eluted from the studied face masks before and after one and five autoclaving cycles. Moreover, a scanning electron microscope (SEM) analysis revealed no changes in the yarns, elongation, tensile strength, and contact angle measurements of the polyester or cotton materials after one or five autoclaving cycles

Amelioration effect of 18β-Glycyrrhetinic acid on methylation inhibitors in hepatocarcinogenesis -induced by diethylnitrosamine

Aimsuppression of methylation inhibitors (epigenetic genes) in hepatocarcinogenesis induced by diethylnitrosamine using glycyrrhetinic acid.MethodIn the current work, we investigated the effect of sole GA combined with different agents such as doxorubicin (DOX) or probiotic bacteria (Lactobacillus rhamanosus) against hepatocarcinogenesis induced by diethylnitrosamine to improve efficiency. The genomic DNA was isolated from rats’ liver tissues to evaluate either methylation-sensitive or methylation-dependent resection enzymes. The methylation activity of the targeting genes DLC-1, TET-1, NF-kB, and STAT-3 was examined using specific primers and cleaved DNA products. Furthermore, flow cytometry was used to determine the protein expression profiles of DLC-1 and TET-1 in treated rats’ liver tissue.ResultsOur results demonstrated the activity of GA to reduce the methylation activity in TET-1 and DLC-1 by 33.6% and 78%, respectively. As compared with the positive control. Furthermore, the association of GA with DOX avoided the methylation activity by 88% and 91% for TET-1 and DLC-1, respectively, as compared with the positive control. Similarly, the combined use of GA with probiotics suppressed the methylation activity in the TET-1 and DLC-1 genes by 75% and 81% for TET-1 and DLC-1, respectively. Also, GA and its combination with bacteria attenuated the adverse effect in hepatocarcinogenesis rats by altering potential methylomic genes such as NF-kb and STAT3 genes by 76% and 83%, respectively.ConclusionGA has an ameliorative effect against methylation inhibitors in hepatocellular carcinoma (HCC) by decreasing the methylation activity genes

Modeling the role of fomites in a norovirus outbreak

Norovirus accounts for a large portion of the gastroenteritis disease burden, and outbreaks have occurred in a wide variety of environments. Understanding the role of fomites in norovirus transmission will inform behavioral interventions, such as hand washing and surface disinfection. The purpose of this study was to estimate the contribution of fomite-mediated exposures to infection and illness risks in outbreaks. A simulation model in discrete time that accounted for hand-to-porous surfaces, hand-to-nonporous surfaces, hand-to-mouth, -eyes, -nose, and hand washing events was used to predict 17 hr of simulated human behavior. Norovirus concentrations originated from monitoring contamination levels on surfaces during an outbreak on houseboats. To predict infection risk, two dose-response models (fractional Poisson and 2F1 hypergeometric) were used to capture a range of infection risks. A triangular distribution describing the conditional probability of illness given an infection was multiplied by modeled infection risks to estimate illness risks. Infection risks ranged from 70.22% to 72.20% and illness risks ranged from 21.29% to 70.36%. A sensitivity analysis revealed that the number of hand-to-mouth contacts and the number of hand washing events had strong relationships with model-predicted doses. Predicted illness risks overlapped with leisure setting and environmental attack rates reported in the literature. In the outbreak associated with the viral concentrations used in this study, attack rates ranged from 50% to 86%. This model suggests that fomites may have accounted for 25% to 82% of illnesses in this outbreak. Fomite-mediated exposures may contribute to a large portion of total attack rates in outbreaks involving multiple transmission modes. The findings of this study reinforce the importance of frequent fomite cleaning and hand washing, especially when ill persons are present.12 month embargo; published online: 04 Feb 2019This item from the UA Faculty Publications collection is made available by the University of Arizona with support from the University of Arizona Libraries. If you have questions, please contact us at [email protected]

Xeno-free trans-differentiation of adipose tissue-derived mesenchymal stem cells into glial and neuronal cells.

Mesenchymal stem cells (MSCs) are undifferentiated cells that have the ability of self-renewal and trans-differentiation into other cell types. They hold out hope for finding a cure for many diseases. Nevertheless, there are still some obstacles that limit their clinical transplantation. One of these obstacles are the xenogeneic substances added in either proliferation or differentiation media with subsequent immunogenic and infectious transmission problems. In this study, we aimed to replace fetal bovine serum (FBS), the main nutrient source for MSC proliferation with xeno-free blood derivatives. We tested the effect of human activated pure platelet-rich plasma (P-PRP) and advanced platelet-rich fibrin (A-PRF) on the proliferation of human adipose derived-MSCs (AD-MSCs) at different concentrations. For the induction of MSC neural differentiation, we used human cerebrospinal fluid (CSF) at different concentrations in combination with P-PRP to effect xeno-free/species-specific neuronal/glial differentiation and we found that media with 10% CSF and 10% PRP promoted glial differentiation, while media with only 10% PRP induced a neuron-like phenotype