Molecular and Cellular Mechanisms Involved in Adipose-derived stem cell and their extracellular vesicles in an Experimental Model of Cardio- renal Syndrome type 3: Histological and Biochemical Study.

A cardio-renal syndrome (CRS) is a medical condition in which kidney problems are accompanied by heart problems and diagnosed when acute kidney injury contributes to the development of acute cardiac injury. Regenerative medicine is becoming increasingly interested in adipose stem cells. We evaluated the effect of both adipose-derived stem cell extracellular vesicles (ADSCs-EVs) and adipose stem cells (ADSCs) on an experimental model of CRSIII. In this study, isolation, and further identification of ADSCs and ADSCs-EVs by transmission electron microscopy and flow cytometric analysis. Cardio-renal syndrome in rats was induced by renal artery ligation RAL followed by a single dose injection of both ADSCs and ADSCs-EVs in separate groups. The effects of ADSCs-EVs and ADSCs against induced CRSIII were evaluated by both renal and cardiac oxidant/antioxidant biomarkers, renal function, and mRNA gene expression quantitation for atrial natriuretic peptide (ANP), p300, and myocyte enhancer factor 2 (MEF2C and MEF2A), as well as myocardin (MYOCD), as molecules associated with cardiac hypertrophy. Additionally, histological and immunohistochemical studies of cardiac and renal tissues were done. ADSCs-EVs were effectively isolated and characterized. ADSCs-EVs and ADSCs reversed induced CRSIII, evidenced by considerably decreased serum urea and creatinine levels. Returned oxidant/antioxidant stability, and decreased caspase 3-mediated apoptotic programmed cell death in cardiac and renal tissues. Additionally, they led to successful down-regulation of hypertrophic cardiac genes levels and reversed histopathological cardiac and renal injures. ADSCs-derived extracellular vesicles and ADSCs injection restored damaged cardiac and renal tissue and improved its function greatly following induced CRSIII. They could therefore be useful as a means of protecting the heart from the deleterious effects of acute renal injury and reprogramed injured cardiac cells by activating regenerative processes. SIMPLE SUMMARY: Cardiorenal syndrome (CRS) type III is a subcategory of CRS whereby acute kidney injury (AKI) could contribute to the development of acute cardiac dysfunction. This study provided innovatory data regarding the role of adipose-derived stem cell extracellular vesicles ADSCs-EVs and adipose stem cells (ADSCs) in acute renal and cardiac dysfunction and renal biopsy specimens in the form of interstitial inflammation/tubular degeneration. The main cause of renal and cardiac dysfunction is identified to be the activation and accumulation of inflammatory cells and oxidants in the interstitium, surrounded by increased amounts of extracellular matrix, and ADSCs-EVs have been proposed as a contributor factor. The study has evidenced that both ADSCs-EVs and adipose stem cells display beneficial effects on renal and cardiac tissues survival in terms of the frequent occurrence of cardio-renal syndrome, ADSC-EVs treatment repaired damaged renal and cardiac tissues and recovered their function. ADSC-EVs reversed the effects of cardio-renal syndrome and reprogramed injured cells by activating regenerative processes. The clinical significance of the results presented in future studies needs to be investigated further.

Anatomical study of the latissimus dorsi muscle flap / Estudio anatómico del colgajo del músculo latísimo del dorso

SUMMARY: The latissimus dorsi is a broad muscle that originates from the inferior thoracic spinous processes, thoracolumbar fascia, iliac crest, and inferior ribs. It inserts on the inferior aspect of the intertubercular groove of the humerus through a thin tendon. The study was conducted on 10 cadavers (7 male and 3 female). These specimens were dissected and examined to study the gross anatomical characteristics of the latissimus dorsi muscle. The dimensions of the latissimus dorsi muscle and its surface area were measured in all the cadavers. The branching pattern of the thoracodorsal vessels was recorded. The pedicle length and caliper were measured using Vernier calipers. On the 20 dissected sides, the thoracodorsal artery was found to be one of the terminal branches of the subscapular artery that originates in the axillary region. In 19 (95 %) cases, the thoracodorsal artery terminated in a bifurcation, giving off a medial and a lateral branch. The average size of the elevated flap of the latissimus dorsi muscle was 18 cm x 36 cm. The average pedicle length was 9.5 cm (range: 5 cm-14 cm), and the average diameter at its origin was 2.5 mm (range: 1.5 mm-3.5 mm). The average diameter of the vena comitans was 3.3 mm. The current study focuses on the anatomical features of the latissimus dorsi muscle and its blood supply to increase the success rate of operations in clinical practice.

RESUMEN: El músculo latísimo del dorso se origina en los procesos espinosos de las vértebras torácicas inferiores, la fascia toracolumbar, la cresta ilíaca y las costillas inferiores y se inserta en el surco intertubercular del húmero a través de un delgado tendón. El estudio se realizó en 10 cadáveres (7 mujeres y 3 hombres). Estos especímenes fueron disecados y examinados para estudiar las características anatómicas macroscópicas del músculo latísimo del dorso. En todos los cadáveres se midieron las dimensiones del músculo y su superficie. Se registró el patrón de ramificación de los vasos toracodorsales. La longitud del pedículo y el calibre se midieron con paquímetro Vernier. En los veinte lados disecados, se encontró que la arteria toracodorsal era una de las ramas terminales de la arteria subescapular que se originaba en la región axilar. En 19 (95 %) casos, la arteria toracodorsal terminaba bifurcándose en dos ramas, una rama medial y otra lateral. El tamaño promedio del colgajo elevado del músculo latísimo del dorso era de 18 cm x 36 cm. La longitud promedio del pedículo era de 9,5 cm (rango: 5 cm-14 cm), y el diámetro promedio en su origen era de 2,5 mm (rango: 1,5 mm-3,5 mm). El diámetro medio de la vena comitans era de 3,3 mm. El estudio actual se centra en las características anatómicas del músculo latísimo del dorso y su irrigación para aumentar la tasa de éxito de las operaciones en la práctica clínica.

Exosomes Derived from BM-MSCs Mitigate the Development of Chronic Kidney Damage Post-Menopause via Interfering with Fibrosis and Apoptosis.

The rate of chronic kidney disease (CKD) is increasing globally, and it is caused by continuous damage to kidney tissue. With time the renal damage becomes irreversible, leading to CKD development. In females, post-menopause lack of estrogen supply has been described as a risk factor for CKD development, and studies targeting post-menopause CKD are scarce. In the present study, we used exosomes isolated from bone marrow mesenchymal stem/stromal cells (BM-MSCs) to test their therapeutic potential against the development of CKD. At first, the menopause model was achieved by surgical bilateral ovariectomy in female albino rats. After that, 100 µg of exosomes was given to ovariectomized rats, and the study continued for 2 months. Changes in urine volume, urine protein content, kidney function biochemical parameters (creatinine and BUN), kidney antioxidant parameters (SOD, GPx and CAT), histological changes, immunohistochemical levels of caspase 3, and the gene expression of NGAL (related to kidney damage), TGFß1 and αSMA (related to fibrosis and EMT), and caspase 3 (related to apoptosis) were studied. After the ovariectomy, the occurrence of CKD was confirmed in the rats by the drastic reduction of serum estrogen and progesterone levels, reduced urine output, increased urinary protein excretion, elevated serum creatinine and BUN, reduced GPx SOD, and CAT in kidney tissue, degenerative and fibrotic lesions in the histopathological examination, higher immunohistochemical expression of caspase 3 and increased expression of all studied genes. After exosomes administration, the entire chronic inflammatory picture in the kidney was corrected, and a near-normal kidney structure and function were attained. This study shows for the first time that BM-MSCs exosomes are potent for reducing apoptosis and fibrosis levels and, thus, can reduce the chronic damage of the kidneys in females that are in their menopause period. Therefore, MSCs-derived exosomes should be considered a valuable therapy for preserving postmenopausal kidney structure and function and, subsequently, could improve the quality of females' life during menopause.

Adipose-derived stem cell and their extracellular vesicles ameliorates immune function, and cardiac markers in experimental model of cardiorenal syndrome type III: TNF-α, IFN-γ and IL-10 cytokine production and their correlation with genotype.

Cardio-renal syndrome (CRS) denotes the convergence of heart-kidney interactions across several mechanisms. The current study is conducted to evaluate the anti-inflammatory role of adipose tissue-derived stem cells (ASCs) versus adipose stem cell-derived extracellular vesicles (ADSCs-EVs) in experimental model of cardiorenal syndrome type III. The study was conducted on 50 male rats that were equally divided to: group I (control group); Group II (experimental cardiorenal syndrome group) which induced by right renal artery ligation (ICRSIII); Group III (Sham-operated control group) which underwent surgical incision without renal artery ligation; Group IV (ICRSIII which received ADSCs-extracellular vesicles (ADSCs-EVs); Group V (ICRSIII which received adipose tissue stem cells (ASCs). Assessment of pro-inflammatory cytokines; IL-10, IL-1α, IL-6, IL-1 ß, IFN-γ, NF-α and their mRNA gene expression quantitation, (NGAL), and brain natriuretic peptide (BNP) as markers of cardiac dysfunction, as well as histopathological examination of renal tissue was examined by H& E, Masson trichrome and periodic acid-Schiff stains (PAS). The ICRS group exhibited significant acute tubular injury with tubular dilation, loss of brush borders, epithelial flattening, and occasional sloughed cells in lumen. Use of either ADSCs-EVs or ASCs significantly ameliorated the histological findings of tubular injury. Proinflammatory cytokines, BNP and NGAL were significantly elevated in ICRSIII group as compared to all other studied groups. Administration of ADSCs-EVs or ASCs led to significant decrease in all proinflammatory cytokines as well as BNP and NGAL levels with no significant difference between them. In conclusion, ADSCs-EXs and ASCs exhibited significant repairing effects in experimental-induced cardiorenal syndrome type III as evidenced by amelioration of histological findings of tubular injury, anti-inflammatory effects, and the significant decrease in markers of cardiac dysfunction. ADSC-EVs reprogramed injured cardiac cells by activating regenerative processes.

Mesenchymal stem-cells' exosomes are renoprotective in postmenopausal chronic kidney injury via reducing inflammation and degeneration.

Chronic kidney disease (CKD) is an important global disease its rates are increasing worldwide. CKD is caused by injuries to kidney tissue that exceeds the rate of regeneration, which with time lead to irreversible renal damage and CKD become evident. In females, diminished estrogen supply in the postmenopausal period is associated with greater risk for developing CKD. In this study we isolated exosomes from bone marrow mesenchymal stem/stromal cells (BM-MSCs) and tested their therapeutic effects on post-menopause CKD (PM-CKD) and compared their effects with BM-MSCs. The menopause model was achieved by bilateral ovariectomy in 8-months-old female albino rats, then no treatment, 2 million BM-MSCs or 100 µg of exosomes (Exo) was given intravenously in tail vein to ovariectomized rats and the study continued for 8 weeks post-ovariectomy. Changes in weight, urine volume, urine protein content, kidney function biochemical parameters (creatinine and BUN), Kidney oxidative stress (MDA), kidney antioxidant parameters (SOD, GPx and CAT), histopathological changes, immunohistochemical expression of KIM-1 and, finally, genes related to renal damage (peroxiredoxin-3, KIM-1 and ICAM-1) and inflammation (TNF-α, Cox2 and IL-6) were recorded for all study groups. Post-ovariectomy there was an increased body weight, drastic reduction of estrogen and progesterone levels, reduced urine output, increased urinary protein excretion, elevated serum creatinine and BUN, increased MDA and reduced GPx SOD, and CAT in kidney tissue, chronic inflammation, degenerative and fibrotic lesions in histopathological examination, high expression of KIM-1 immunohistochemically and changes in gene expression analyses all pointing to the development of CKD in the study rats. In the PM-CKD groups receiving BM-MSCs or Exo, the whole chronic inflammatory picture was completely reversed towards a much normal kidney structure and function. The improvements were more observable with Exo compared to BM-MSCs. Overall, our results show for the first time that exosomes isolated from BM-MSCs are more potent in reducing chronic inflammatory changes in the kidney of postmenopausal females compared to the cell-based approach using BM-MSCs. Therefore, MSCs-derived exosomes are a promising therapeutic approach for preserving postmenopausal kidney structure and function and, subsequently, should improve the quality of life of postmenopausal females.

Effect of vanillic acid on pentylenetetrazole-kindled rats: Nrf2/HO-1, IGF-1 signaling pathways cross talk.

Vanillic acid (VA) exhibited antioxidant and neuroprotective properties in some neurodegenerative disorders. So, the current study examined the neuroprotective potential of VA as an antiepileptic agent in pentylenetetrazole (PTZ)-induced epileptic rats and the prospective role of Insulin like growth factor-1 (IGF-1) and nuclear factor-2 erythroid-related factor-2 (Nrf2)/heme oxygenase-1 (HO-1) pathway in this respect. Thirty male albino rats were equally subdivided into 3 groups; (1) normal control (NC) group, (2) PTZ-group: received PTZ (50 mg/Kg, i.p. every other day) for 14 days, and (3) PTZ + VA group: received PTZ and VA (50 mg/Kg daily for 2 weeks). The seizure score and latency were evaluated after PTZ injection. Also, the markers of oxidative stress (malondialdehyde (MDA), catalase, and reduced glutathione (GSH)), histopathological examination, the expression of glial fibrillary acidic protein (GFAP) (a marker of astrocytes) IGF-1, Nrf2, and HO-1 were assessed in the brain tissues by the end of the experiment. PTZ caused significant decrease in seizure latency and significant increase in seizure score by the end of the experiment (p < 0.01). This was associated with significant increase in MDA and GFAP with significant decrease in GSH, total antioxidant capacity (TAC) and IGF-1 in brain tissues compared to normal group (p < 0.01). On the other hand, treatment with VA caused significant attenuation in PTZ-induced seizures which was associated with significant improvement in oxidative stress markers and downregulation in GFAP and upregulation of Nrf2, HO-1 and IGF-1 in CA3 hippocampal region (p < 0.01). VA showed neuroprotective and anti-epileptic effects against PTZ-induced epilepsy which probably might be due to its antioxidant properties and upregulation of Nrf2/HO-1 pathway and IGF-1.

Effects of Stevia rebaudiana Bertoni extracts in the rat model of epilepsy induced by pentylenetetrazol: Sirt-1, at the crossroads between inflammation and apoptosis.

The current study investigated the effects of stevia extracts on a PTZ-induced epileptic rat model and its potential mechanism. Thirty male Sprague-Dawley rats were equally subdivided into 3 groups; (1) normal control (NC) group, (2) PTZ-group: received PTZ (50 mg/kg, i.p. every other day) for 2 weeks, and (3) PTZ+ Stevia group: received PTZ and stevia (200 mg/kg orally daily) for 4 weeks (2 weeks before the start of PTZ treatment and 2 weeks with PTZ administration). The first jerk latency and the seizure score were assessed in rats. Also, brain tissue samples were collected by the end of the experiment, and oxidative stress markers (catalase, MDA, and total antioxidant capacity (TAC)) were measured by biochemical analysis in hippocampal brain homogenates. Also, in the hippocampus, the expression of IL6 and Bcl-2 at the mRNA level and expression of Sirt-1, P53, caspase-3, GFAP, and NF-kB in CA3 hippocampal region by immunohistochemistry was investigated. PTZ substantially increased the seizure score and decreased the seizure latency. Also, PTZ significantly increased MDA, GFAP, IL-6, NF-kB, caspase-3, and p53 and significantly reduced Sirt-1, TAC, and Bcl-2 in hippocampal tissues compared to the control group (p < 0.01). However, Stevia Rebaudiana Bertoni (Stevia R.) significantly attenuated the PTZ-induced seizures, improved oxidative stress markers, downregulated GFAP, IL-6, NF-kB, caspase-3, and p53, and upregulated Sirt-1 and Bcl-2 in the CA3 hippocampal region (p < 0.01). In conclusion, Stevia R. exhibits neuroprotective and antiepileptic actions in PTZ-induced epilepsy due to its antioxidant, anti-apoptotic, and anti-inflammatory effects. Additionally, the Sirt-1 pathway might be involved in the antiepileptic and neuroprotective effects of stevia in PTZ-kindled epileptic rat model.

The biomechanistic aspects of renal cortical injury induced by diesel exhaust particles in rats and the renoprotective contribution of quercetin pretreatment: Histological and biochemical study.

Although several studies have reported a toxic effect of diesel exhaust particles (DEP) exposure on the kidney tissues, the involvement of autophagy/NF-kB signaling as encountered mechanisms and the protective effects of a natural flavonoid, quercetin on DEP remains unclear. Thirty-two albino rats were divided as control, quercetin-treated (60 mg/kg, oral), DEP-exposed (0.5 mg/kg, intra-tracheal), and quercetin/DEP-exposed groups. Specimens of the renal cortex were subjected to histo-biochemical study and immunohistochemical analysis using anti-NF-kB, and anti-LC3ß antibodies followed by morphometric and statistical analyses. The expression level of autophagy genes was quantitatively evaluated using RT-PCR, as well. The DEP-exposed rats showed an elevation in the renal tissue levels of MDA and a decrease in the catalase and superoxide dismutase (p < .05). Histologically, there were cytoplasmic vacuolar changes in the lining cells of the renal tubules, glomerular atrophy, and vascular congestion. In addition, renal inflammation was evident as confirmed by the increased NF-kB immunoexpression. Moreover, the gene expression of Becn1, ATG5, and LC3ß increased (p <. 0) due to DEP exposure. Conversely, quercetin pretreatment improved these renal histo-biochemical alterations (p < .05) and regulated autophagy/NF-kB pathways. Overall, the study proved the renal toxicity mediated by DEP exposure via precipitating renal inflammation, autophagy activation, and oxidative stress. Quercetin pretreatment could antagonize such machinery to protect the kidney against DEP.

Medical Students' Feedback of Applying the Virtual Dissection Table (Anatomage) in Learning Anatomy: A Cross-sectional Descriptive Study.

OBJECTIVE: Conventional cadaver dissection is the main learning tool for studying human anatomy. Other tools have been developed to add another dimension of depth to anatomy teaching, such as Anatomage. 3D Anatomage is a touch interactive anatomy virtual dissection table used in anatomy education. The aim of this study was to address students' opinions on applying the virtual dissection table (3D Anatomage) as an additional tool to cadaver dissection in learning anatomy. METHODS: An electronic questionnaire consisted of 6 questions that included items regarding the effect of using 3D Anatomage on students' deep understanding of anatomy topics, the locations and relationships of the different internal body structures and the application of anatomical knowledge. The survey was completed by 78 medical students. RESULTS: The results of this study showed that the majority of students preferred using 3D Anatomage as an additional tool to cadaver dissection for learning anatomy since it enhances active learning, and approximately 89% of students agreed that 3D Anatomage helped them to better understand the relationships between internal structures and visualizing the body system. Additionally, approximately 72% of the students indicated that the imaging facility in 3D Anatomage was useful for enabling their understanding of anatomy as it is envisioned through medical imaging. CONCLUSION: In conclusion, using the 3D Anatomage virtual dissection table is effective in anatomy education, and its use is recommended as an anatomy learning resource in addition to cadavers.

Autophagy Promotes the Survival of Adipose Mesenchymal Stem/Stromal Cells and Enhances Their Therapeutic Effects in Cisplatin-Induced Liver Injury via Modulating TGF-ß1/Smad and PI3K/AKT Signaling Pathways.

Autophagy is a key metabolic process where cells can recycle its proteins and organelles to regenerate its own cellular building blocks. Chemotherapy is indispensable for cancer treatment but associated with various side-effects, including organ damage. Stem cell-based therapy is a promising approach for reducing chemotherapeutic side effects, however, one of its main culprits is the poor survival of transplanted stem cells in damaged tissues. Here, we aimed to test the effects of activating autophagy in adipose-derived mesenchymal stem/stromal cells (ADSCs) on the survival of ADSCs, and their therapeutic value in cisplatin-induced liver injury model. Autophagy was activated in ADSCs by rapamycin (50 nM/L) for two hours before transplantation and were compared to non-preconditioned ADSCs. Rapamycin preconditioning resulted in activated autophagy and improved survival of ADSCs achieved by increased autophagosomes, upregulated autophagy-specific LC3-II gene, decreased protein degradation/ubiquitination by downregulated p62 gene, downregulated mTOR gene, and finally, upregulated antiapoptotic BCL-2 gene. In addition, autophagic ADSCs transplantation in the cisplatin liver injury model, liver biochemical parameters (AST, ALT and albumin), lipid peroxidation (MDA), antioxidant profile (SOD and GPX) and histopathological picture were improved, approaching near-normal conditions. These promising autophagic ADSCs effects were achieved by modulation of components in TGF-ß1/Smad and PI3K-AKT signaling pathways, besides reducing NF-κB gene expression (marker for inflammation), reducing TGF-ß1 levels (marker for fibrosis) and increasing SDF-1 levels (liver regeneration marker) in liver. Therefore, current results highlight the importance of autophagy in augmenting the therapeutic potential of stem cell therapy in alleviating cisplatin-associated liver damage and opens the path for improved cell-based therapies, in general, and with chemotherapeutics, in particular.

Anatomical study and clinical importance of the inferior epigastric artery / Estudio anatómico e importancia clínica de la arteria epigástrica inferior

SUMMARY: The inferior epigastric artery (IEA) is a major blood vessel that supplies the anterior abdominal wall. The aim of the current study was to provide clinicians, surgeons, and obstetricians with sufficient anatomical data on the inferior epigastric artery, such as its origin and branching pattern. The study included 20 embalmed cadavers, these cadavers were dissected, and the inferior epigastric artery and vena comitans/venae comitantes were identified and traced downwards to the external iliac vessels. The origins, caliber, course and pedicle length of both the artery and the vein(s) were studied. The inferior epigastric artery arose independently from the distal external iliac artery deep to the inguinal ligament in 19 (95 %) cadavers. The artery entered the rectus abdominis muscle at its middle third in 13 (65 %) cases and at its lower third in the remaining specimens. In this study, we found that the artery divided into two branches in 18 (90 %) of the cases; in the remaining two cases, it continued as one trunk. The average pedicle length was 7.2 cm. The mean caliber of the IEA was 3.7 mm. In 18 (90 %) dissections, the venous drainage consisted of a pair of venae comitantes that united to form a common vessel at their draining point on the external iliac vein. The average diameter was 3.9 mm. The current study focuses on the anatomical features of the inferior epigastric artery to increase the success rate of abdominal and pelvic operations in clinical practice.

RESUMEN: La arteria epigástrica inferior (AEI) es un vaso sanguíneo principal que irriga la pared abdominal anterior. El objetivo del presente estudio fue proporcionar a los médicos, cirujanos y obstetras suficientes datos anatómicos sobre la arteria epigástrica inferior, como su origen y patrón de ramificación. El estudio incluyó 20 cadáveres embalsamados, los que se disecaron y se identificó la arteria epigástrica inferior y la vena concomitante y se siguieron hasta los vasos ilíacos externos. Se estudiaron los orígenes, calibre, trayecto y longitud del pedículo tanto de la arteria como de la (s) vena (s). La arteria epigástrica inferior surgió independientemente de la arteria ilíaca externa profunda al ligamento inguinal en 19 (95 %) cadáveres. La arteria ingresó al músculo recto del abdomen en su tercio medio en 13 (65 %) casos y en su tercio inferior en las muestras restantes. En este estudio, encontramos que la arteria se dividió en dos ramas en 18 (90 %) de los casos; en los dos casos restantes, continuó como un tronco. La longitud media del pedículo fue de 7,2 cm. El calibre medio del AEI fue de 3,7 mm. En 18 (90 %) disecciones, el drenaje venoso consistió en un par de venas concomitantes las que formaron un vaso común en su punto de drenaje en la vena ilíaca externa. El diámetro medio fue de 3,9 mm. El estudio actual se centra en las características anatómicas de la arteria epigástrica inferior con el propósito de mejorar la tasa de éxito de las cirugías abdominales y pélvicas en la práctica clínica.

Structural and chemical role of mesenchymal stem cells and resveratrol in regulation of apoptotic -induced genes in Bisphenol-A induced uterine damage in adult female albino rats.

The probable beneficial effects of mesenchymal stem cells (MSCs) and resveratrol were assessed in an experimental model of Bisphenol-A (BPA)-evident uterine damage in rats. Thirty-five albino rats were involved and equally divided into five groups: Group I: negative control rats received usual diet, Group II: positive control rats received BPA by oral gavage for 15 days, Group III: BPA-treated rats received single oral gavage of resveratrol daily for two weeks, Group IV: BPA-treated rats received a single intravenous dose of MSCs and Group V: BPA-treated rats received combined treatment of resveratrol and MSCs. Oxidative stress markers, apoptosis-related genes, and gonadal hormones were assessed. Histological and immunohistochemical examination of uterine tissue was conducted for TGF-ß 1. Caspases-3, 8, and 9 (Casp3, Casp8, Casp9) genes were assessed in uterine tissues by quantitative real-time PCR. Results revealed that BPA induced significant changes in the endometrial tissue, inflammatory cell infiltration, focal blood extravasation, increase in collagen fibers, decrease in PAS staining, and increase in TGF-ß 1 immunoreactivity. BPA also induced a significant increase in oxidative stress markers; malondialdehyde (MDA), SOD, CAT, and apoptosis-related genes. BPA induced a significant change in blood levels of gonadal hormones; a significant increase in FSH and a significant decrease in estradiol (E2) and progesterone (P). Treatment with either resveratrol, MSCs, or a combination of them resulted in significant enhancement of histological findings, restoration of gonadal hormones to near-normal levels, and a significant decrease in oxidative stress markers and apoptosis genes. Combined treatment with resveratrol and MSCs demonstrated more significant therapeutic effects as regard to the studied parameters in association with rat groups treated with either MSCs or resveratrol separately.

Extracellular vesicles derived from bone marrow mesenchymal stem cells repair functional and structural rat adrenal gland damage induced by fluoride.

The adrenal glands have striking morpho-biochemical features that render them vulnerable to the effects of toxins. AIMS: This study was conducted to explore the therapeutic utility of extracellular vesicles derived from bone marrow mesenchymal stem cells (BMSC-EVs) against fluoride-induced adrenal toxicity. MATERIALS AND METHODS: The work included isolation and further identification of BMSC-EVs by transmission electron microscopy and flow cytometric analysis. Adrenal toxicity in rats was induced by oral administration of 300 ppm of sodium fluoride (NaF) in drinking water for 60 days followed by a single dose injection of BMSC-EVs. The effects of BMSC-EVs against NaF was evaluated by adrenal oxidant/antioxidant biomarkers, hormonal assay of plasma adrenocorticotrophic hormone (ACTH) and corticosterone (CORT) and mRNA gene expression quantitation for adrenal cortical steroidogenic pathway-encoding genes. Histopathological examination of the adrenal tissue was performed. KEY FINDINGS: BMSC-EVs were effectively isolated and characterized. NaF exposure decreased adrenal superoxide dismutase and catalase activities, increased adrenal malondialdehyde levels, elevated plasma ACTH, diminished CORT concentrations and downregulated the adrenal cortical steroidogenic pathway-encoding genes. In addition, NaF-induced marked adrenal histopathological lesions. SIGNIFICANCE: BMSC-EVs treatment repaired damaged adrenal tissue and recovered its function greatly following NaF consumption. BMSC-EVs reversed the toxic effects of NaF and reprogramed injured adrenal cells by activating regenerative processes.

A possible role of endoneurial fibroblast-like cells in the resolution of endoneurial oedema following nerve crush injury / El posible rol de los fibroblastos endoneurales en la resolución del edema endoneural después de una lesión por aplastamiento nervioso

SUMMARY: Endoneurial oedema is a salient feature of all types of neuropathy. Its elimination is crucial during the complications of nerve recovery. The objective was to study a possible role of the endoneurial fibroblasts in the resolution of nerve edema. Forty-two albino male rats aged between 30 and 40 days (weight 200 g to 250 g) were used in this study. The left sural nerves of 36 rats were subjected to crush injury at one to three-week intervals with six animals per interval. The right and left sural nerves of the remaining six rats were used as controls. At the end of the second week after crush injury, the endoneurium showed channel-like spaces that were lined by fibroblast-like cells and collagen bundles that contained degenerated myelin, and were connected to the subperineurial spaces. Flattened fibroblast-like cells were arranged in several layers in the subperineurial, forming barrier-like cellular sheets localizing to the endoneurial oedema in the space. Fibroblast-like cells also wrapped around the regenerating nerve fibres with their branching cytoplasmic processes. During the third week, the flattened fibroblast-like cells formed nearly continuous cellular sheets in the subperineurial spaces. Macrophages were frequently observed between these cellular barrier-like sheets and in the subperineurial. The endoneurial fibroblast-like cells form barrier-like cellular sheets that probably localise the endoneurial oedema in the subperineurial space. It also appear to create endoneurial channel-like spaces containing degenerated myelin and endoneurial oedema, which may be helpful in localizing and resolving such oedema.

RESUMEN: El edema endoneural es una característica destacada de todos los tipos de neuropatía. Su eliminación es importante durante las complicaciones de la recuperación nerviosa. El objetivo fue estudiar un posible papel de los fibroblastos endoneurales en la resolución del edema nervioso. En este estudio se utilizaron 42 ratas macho albinas con edades entre los 30 y 40 días (peso 200 a 250 g). Los nervios surales izquierdos de 36 ratas se sometieron a lesiones por aplastamiento en intervalos de una a tres semanas con seis animales por intervalo. Se usaron los nervios surales derecho e izquierdo de las seis ratas restantes como controles. Al final de la segunda semana después de la lesión por aplastamiento, el endoneuro mostró espacios en forma de canal que estaban revestidos por células similares a fibroblastos y haces de colágeno que contenían mielina degenerada y se conectaron a los espacios subperineurales. Las células aplanadas de fibroblastos se dispusieron en varias capas en el subperineuro, formando láminas celulares de tipo barrera que se localizaban en el espacio del edema endoneural. Las células similares a fibroblastos también envolvían las fibras nerviosas regeneradoras con sus procesos citoplásmicos ramificados. Durante la tercera semana, las células aplanadas de fibroblastos formaron láminas celulares casi continuas en los espacios subperineurales. Los macrófagos se observaron con frecuencia entre estas láminas similares a barreras celulares y en el subperineuro. Las células de tipo fibroblasto endoneural formaban láminas celulares de tipo barrera que probablemente localizan el edema endoneural en el espacio subperineural. También parece que crea espacios en forma de canal endoneural que contienen mielina degenerada y edema endoneural, que pueden ser útiles para localizar y resolver este edema.

Does oregano protect against testicular toxicity produced by ethylene glycol in adult male albino rat? / ¿El orégano protege contra la toxicidad testicular producida por etilenglicol en ratas macho albinas adultas?

SUMMARY: Origanum vulgare Linn has traditionally been used as a diuretic and antispasmodic. Therefore, we investigated the active extract of Origanum vulgare for possible andrological effect and preventive effects against testicular damage using ethylene glycol rat model of testicular damage, to rationalize its medicinal use. Male Wistar rats received lithogenic treatment comprising of 0.75 % ethylene glycol injection twice with one day interval, then in drinking water, active extract of Origanum vulgare treatment (20 mg/kg) was given for 3 weeks to prevent toxic damage including loss of body weight gain and appetite, Following oral administration of EGME, a rapid decrease in testis weight associated with testicular cell damage was observed. Origanum vulgare treatment (20 mg/kg) prevented as well as reversed toxic changes including loss of body weight gain.

RESUMEN: Origanum vulgare Linn se ha usado tradicionalmente como diurético y antiespasmódico. Por lo tanto, investigamos el extracto activo de Origanum vulgare por su posible efecto andrológico y efectos preventivos contra el daño testicular utilizando el modelo de rata de etilenglicol de daño testicular. El objetivo del estudio fue racionalizar su uso medicinal. Su utilizaron ratas Wistar macho que recibieron un tratamiento litogénico de una inyección de etilenglicol al 0,75 %, dos veces con un intervalo de un día, y luego se administró en agua potable. Se administró el extracto activo del tratamiento con Origanum vulgare (20 mg / kg) durante 3 semanas con el objetivo de prevenir el daño tóxico, la pérdida de peso corporal y el apetito. Tras la administración oral de EGME, se observó una rápida disminución del peso de los testículos asociada al daño de las células testiculares. El tratamiento con Origanum vulgare (20 mg / kg) logró prevenir y revertir las alteraciones tóxicas, incluyendo la pérdida de peso corporal.

The anatomical basis of the vascularized fibularis brevis grafts / Base anatómica de los injertos vascularizados del músculo fibularis brevis

Fibularis brevis grafts have been extensively used, especially as distally-based grafts, to cover defects in the lower leg and foot. The study has contributed to analyze the blood supply of the fibularis brevis muscle and the vascular basis of its possible different grafts. Both legs of twelve preserved cadavers, without congenital vascular anomalies, were utilized in the study. Fibularis brevis was exposed in all selected legs along with the verification of different arterial pedicles. The total means and standard deviations of the length and width of the muscle together with diameters of major vessels were calculated. Total length and width of middle portion of fibularis brevis were at means of 28.7±0.4 cm and 3±0.02 cm respectively. The upper and middle portions of the muscle were supplied by the fibular and the anterior tibial artery. The middle portion was supplemented by the upper perforating branch of the posterior tibial artery. The lower portion of the muscle was supplied by the lower perforating branch of the posterior tibial and the periosteal arteries. The muscle could be used as a proximally or distally based flap, free vascularized muscle graft, free vascularized osteo-muscular flap, and distally-based split flap. It can be split completely into two flaps; each of which can be used as a proximally or distally.

Los injertos de músculo fibularis brevis son usados ampliamente, especialmente como injertos de base distal, para cubrir defectos en la parte inferior de la pierna y el pie. El objetivo de este estudio fue analizar el suministro de sangre del músculo fibularis brevis y la base vascular de los posibles diferentes injertos. Para el estudio se utilizaron ambas piernas de 12 cadáveres preservados, sin anomalías vasculares congénitas. El músculo fibularis brevis fue encontrado en todas las piernas junto con los diferentes pedículos arteriales. Se calcularon las medias totales y las desviaciones estándar de la longitud y el ancho del músculo, junto con los diámetros de los vasos principales. La longitud y el ancho totales de la porción media del músculo fibularis brevis fueron de 28,7±0,4 cm y 3±0,02 cm, respectivamente. Las partes superior y media del músculo estaban suplidas vascularmente por la arteria fibular y la arteria tibial anterior. La parte media se complementó con la rama perforante superior de la arteria tibial posterior. La parte inferior del músculo fue suplida por una rama perforante inferior de la arteria tibial posterior y las arterias periósticas. El músculo podría usarse como un colgajo de base proximal o distal, injerto de músculo vascularizado libre, colgajo osteomuscular vascularizado libre y colgajo dividido distal. Se puede dividir por completo en dos colgajos; cada uno de estos puede ser utilizado como proximal o distal.

Specific loss of CatSper function is sufficient to compromise fertilizing capacity of human spermatozoa.

STUDY QUESTION: Are significant abnormalities of CatSper function present in IVF patients with normal sperm concentration and motility and if so what is their functional significance for fertilization success? SUMMARY ANSWER: Sperm with a near absence of CatSper current failed to respond to activation of CatSper by progesterone and there was fertilization failure at IVF. WHAT IS KNOWN ALREADY: In human spermatozoa, Ca(2+) influx induced by progesterone is mediated by CatSper, a sperm-specific Ca(2+) channel. A suboptimal Ca(2+) influx is significantly associated with, and more prevalent in, men with abnormal semen parameters, and is associated with reduced fertilizing capacity. However, abnormalities in CatSper current can only be assessed directly using electrophysiology. There is only one report of a CatSper-deficient man who showed no progesterone potentiated CatSper current. A CatSper 2 genetic abnormality was present but there was no information on the [Ca(2+)]i response to CatSper activation by progesterone. Additionally, the semen samples had indicating significant abnormalities (oligoasthenoteratozoospermia) multiple suboptimal functional responses in the spermatozoon. As such it cannot be concluded that impaired CatSper function alone causes infertility or that CatSper blockade is a potential safe target for contraception. STUDY DESIGN, SIZE, DURATION: Spermatozoa were obtained from donors and subfertile IVF patients attending a hospital assisted reproductive techniques clinic between January 2013 and December 2014. In total 134 IVF patients, 28 normozoospermic donors and 10 patients recalled due to a history of failed/low fertilization at IVF took part in the study. PARTICIPANTS/MATERIALS, SETTING, METHODS: Samples were primarily screened using the Ca(2+) influx induced by progesterone and, if cell number was sufficient, samples were also assessed by hyperactivation and penetration into viscous media. A defective Ca(2+) response to progesterone was defined using the 99% confidence interval from the distribution of response amplitudes in normozoospermic donors. Samples showing a defective Ca(2+) response were further examined in order to characterize the potential CatSper abnormalities. In men where there was a consistent and robust failure of calcium signalling, a direct assessment of CatSper function was performed using electrophysiology (patch clamping), and a blood sample was obtained for genetic analysis. MAIN RESULTS AND THE ROLE OF CHANCE: A total of 101/102 (99%) IVF patients and 22/23 (96%) donors exhibited a normal Ca(2+) response. The mean (± SD) normalized peak response did not differ between donors and IVF patients (2.57 ± 0.68 [n = 34 ejaculates from 23 different donors] versus 2.66 ± 0.68 [n = 102 IVF patients], P = 0.63). In recall patients, 9/10 (90%) showed a normal Ca(2+) response. Three men were initially identified with a defective Ca(2+) influx. However, only one (Patient 1) had a defective response in repeat semen samples. Electrophysiology experiments on sperm from Patient 1 showed a near absence of CatSper current and exon screening demonstrated no mutations in the coding regions of the CatSper complex. There was no increase in penetration of viscous media when the spermatozoa were stimulated with progesterone and importantly there was failed fertilization at IVF. LIMITATIONS, REASONS FOR CAUTION: A key limitation relates to working with a specific functional parameter (Ca(2+) influx induced by progesterone) in fresh sperm samples from donors and patients that have limited viability. Therefore, for practical, technical and logistical reasons, some men (∼ 22% of IVF patients) could not be screened. As such the incidence of significant Ca(2+) abnormalities induced by progesterone may be higher than the ∼ 1% observed here. Additionally, we used a strict definition of a defective Ca(2+) influx such that only substantial abnormalities were selected for further study. Furthermore, electrophysiology was only performed on one patient with a robust and repeatable defective calcium response. This man had negligible CatSper current but more subtle abnormalities (e.g. currents present but significantly smaller) may have been present in men with either normal or below normal Ca(2+) influx. WIDER IMPLICATIONS OF THE FINDINGS: These data add significantly to the understanding of the role of CatSper in human sperm function and its impact on male fertility. Remarkably, these findings provide the first direct evidence that CatSper is a suitable and specific target for human male contraception.

The clinical significance of calcium-signalling pathways mediating human sperm hyperactivation.

STUDY QUESTION: What is the prevalence of defects in the Ca(2+)-signalling pathways mediating hyperactivation (calcium influx and store mobilization) among donors and sub-fertile patients and are they functionally significant, i.e. related to fertilization success at IVF? SUMMARY ANSWER: This study identifies, for the first time, the prevalence of Ca(2+) store defects in sperm from research donors, IVF and ICSI patients. It highlights the biological role and importance of Ca(2+) signalling (Ca(2+) store mobilization) for fertilization at IVF. WHAT IS KNOWN ALREADY: Sperm motility and hyperactivation (HA) are important for fertility, mice with sperm incapable of HA are sterile. Recently, there has been significant progress in our knowledge of the factors controlling these events, in particular the generation and regulation of calcium signals. Both pH-regulated membrane Ca(2+) channels (CatSper) and Ca(2+) stores (potentially activating store-operated Ca(2+) channels) have been implicated in controlling HA. STUDY DESIGN, SIZE, AND DURATION: This was a prospective study examining a panel of 68 donors and 181 sub-fertile patients attending the Assisted Conception Unit, Ninewells Hospital Dundee for IVF and ICSI. Twenty-five of the donors gave a second sample (∼4 weeks later) to confirm consistency/reliability of the recorded responses. Ca(2+) signalling was manipulated using three agonists, NH4Cl (activates CatSper via pH), progesterone (direct activation of CatSper channels, potentially enhancing mobilization of stored Ca(2+) by CICR) and 4-aminopyridine (4-AP) (effect on pH equivalent to NH4Cl and mobilizes stored Ca(2+)). The broad-spectrum phosphodiesterase inhibitor 3-isobutyl-1-methyxanthine (IBMX), a potent activator of HA was also used for comparison. For patient samples, an aliquot surplus to requirements for IVF/ICSI treatment was examined, allowing direct comparison of Ca(2+) signalling and motility data with functional competence of the sperm. MATERIALS, SETTING, METHODS: The donors and sub-fertile patients were screened for HA (using CASA) and changes in intracellular Ca(2+) were assessed by loading with Fura-2 and measuring fluorescence using a plate reader (FluoStar). MAIN RESULTS AND THE ROLE OF CHANCE: The relative efficacy of the stimuli in inducing HA was 4-AP >> IBMX > progesterone. NH4Cl increased [Ca(2+)]i similarly to 4-AP and progesterone but did not induce a significant increase in HA. Failure of samples to generate HA (no significant increase in response to stimulation with 4-AP) was seen in just 2% of research donors but occurred in 10% of IVF patients (P = 0.025). All donor samples generated a significant [Ca(2+)]i increase when stimulated with 4-AP but 3.3% of IVF and 28.6% of ICSI patients failed to respond. Amplitudes of HA and [Ca(2+)]i responses to 4-AP were correlated with fertilization rate at IVF (P= 0.029; P = 0.031, respectively). Progesterone reliably induced [Ca(2+)]i responses (97% of donors, 100% of IVF patients) but was significantly less effective than 4-AP in inducing HA. Twenty seven per cent of ICSI patients failed to generate a [Ca(2+)]i response to progesterone (P= 0.035). Progesterone-induced [Ca(2+)]i responses were correlated with fertilization rate at IVF (P= 0.037) but induction of HA was not. In donor samples examined on more than one occasion consistent responses for 4-AP-induced [Ca(2+)]i (R(2) = 0.97) and HA (R(2) = 0.579) were obtained. In summary, the data indicate that defects in Ca(2+) signalling leading to poor HA do occur and that ability to undergo Ca(2+) -induced HA affects IVF fertilizing capacity. The data also confirm that release of stored Ca(2+) is the crucial component of Ca(2+) signals leading to HA and that Ca(2+) store defects may therefore underlie HA failure. LIMITATIONS, REASONS FOR CAUTION: This is an in vitro study of sperm function. While the repeatability of the [Ca(2+)]i and HA responses in samples from the same donor were confirmed, data for patients were from 1 assessment and thus the robustness of the failed responses in patients' needs to be established. The focus of this study was on using 4AP, which mobilizes stored Ca(2+) and is a potent inducer of HA. The n values for other agonists, especially calcium assessments, are smaller. WIDER IMPLICATIONS OF THE FINDINGS: Previous studies have shown a significant relationship between basal levels of HA, calcium responses to progesterone and IVF fertilization rates. Here, we have systematically investigated the ability/failure of human sperm to generate Ca(2+) signals and HA in response to targeted pharmacological challenge and, related defects in these responses to IVF success. [Ca(2+)]i signalling is fundamental for sperm motility and data from this study will lead to assessment of the nature of these defects using techniques such as single-cell imaging and patch clamping. STUDY FUNDING/COMPETING INTEREST(S): Resources from a Wellcome Trust Project Grant (#086470, Publicover and Barratt PI) primarily funded the study. The authors have no competing interests.