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Our electron microscopy observations demonstrate for the first time that the number of microvilli on the mice oocyte membrane decreases when meiosis progresses from prophase I to metaphase II (MII) stage, and the morphology of the microvilli also changes. Microvilli are significantly shorter and larger on the ovulated oocyte membrane than at the previous stages. Although clathrin vesicles clearly disappear during oocyte maturation, exosome-like vesicles begin to be secreted at the metaphase I stage, more strongly at the MII stage. Multivesicular bodies are visible only at the MII stage. Since several oocyte tetraspanins are involved in the gamete interaction, Cd9 being congregated on the MII oocyte microvilli, we analyzed the effect of tetraspanin deletion on oocyte membrane morphology. The Cd9−/− and Cd9−/− Cd81−/− deletions are associated with a decreased microvilli density on the MII oocyte surface. Microvilli thickness is significantly increased whatever the deleted tetraspanin gene be. Only Cd9 deletion clearly disturbs the vesicular traffic, increasing the number of clathrin and exosome vesicles. Additional investigations are necessary to elucidate how tetraspanins modulate the microvilli morphology, likely in relation with cytoskeleton. The role of oocyte exosomes in gamete adhesion/fusion remains to be further studied. [ABSTRACT FROM AUTHOR]

Purpose: To assess the impact of peritoneal endometriosis on oocyte and embryo quality in a mouse model. Methods: Peritoneal endometriosis was surgically induced in 33 B6CBA/F1 female mice (endometriosis group, N = 17) and sham-operated were used as control (sham group, N = 16). Mice were superovulated 4 weeks after surgery and mated or not, to collect E0.5-embryos or MII-oocytes. Evaluation of oocyte and zygote quality was done by immunofluorescence under spinning disk confocal microscopy. Results: Endometriosis-like lesions were observed in all mice of endometriosis group. In both groups, a similar mean number of MII oocytes per mouse was observed in non-mated mice (30.2 vs 32.6), with a lower proportion of normal oocytes in the endometriosis group (61 vs 83 %, p < 0.0001). Abnormalities were incomplete extrusion or division of the first polar body and spindle abnormalities. The mean number of zygotes per mouse was lower in the endometriosis group (21 vs 35.5, p = 0.02) without difference in embryo quality. Conclusions: Our results support that induced peritoneal endometriosis in a mouse model is associated with a decrease in oocyte quality and embryo number. This experimental model allows further studies to understand mechanisms of endometriosis-associated infertility. [ABSTRACT FROM AUTHOR]

Context: Cactus pear ( Opuntia ficus-indica (L.) Mill. (Cactaceae)) is a medicinal plant widely used to treat diabetes. Objective: This work investigates the hypoglycemic and antihyperglycemic effect of cactus pear seed oil (CPSO), its mechanism of action, and any toxic effects. Materials and methods: The hypoglycemic effect of CPSO was evaluated in groups of six healthy Wistar rats given 1 or 2 ml kg−1 orally and compared with groups receiving glibenclamide (2 mg kg−1) or water. Glycemia was determined after 30, 60, 120, 240, and 360 min. The antihyperglycemic effect of CPSO was determined in healthy rats and in streptozotocin-induced diabetic rats (STZ); normal rats received 0.8 ml kg−1 CPSO, while diabetic rats received 1 ml kg−1 CPSO, their controls received water or 2 mg kg−1 glibenclamide. For the antihyperglycemic effect evaluation, all the animals were fasted for 16 h before treatment and received glucose orally at 1 g kg−1 30 min after treatment; blood was taken after 30, 90, 150, and 210 min. Intestinal glucose absorption was estimated in rat jejunum perfused with a solution containing 5.55 mmol l−1 glucose. Acute toxicity was determined in albino mice that received oral or intraperitoneal doses of 1, 3, or 5 ml kg−1 CPSO. Results: CPSO (p.o.) decreased postprandial hyperglycemia (60 min after glucose loading), 40.33% and 16.01%, in healthy and STZ-diabetic glucose-loaded rats, respectively. CPSO, also, significantly decreased intestinal glucose absorption by 25.42%. No adverse effects were seen in mice administered CPSO at up to 5 ml kg−1. Conclusion: CPSO is antihyperglycemic. The effect can be explained partly by inhibition of intestinal glucose absorption. [ABSTRACT FROM AUTHOR]

We have recently demonstrated the remarkable efficiency of self-complementary (sc) AAV9 vectors for central nervous system (CNS) gene transfer following intravenous delivery in mice and larger animals. Here, we investigated whether gene delivery to motor neurons (MNs) could also be achieved via intramuscular (i.m.) scAAV9 injection and subsequent retrograde transport along the MNs axons. Unexpectedly, we found that a single injection of scAAV9 into the adult mouse gastrocnemius (GA) mediated widespread MN transduction along the whole spinal cord, without limitation to the MNs connected to the injected muscle. Spinal cord astrocytes and peripheral organs were also transduced, indicating vector spread from the injected muscle to both the CNS and the periphery through release into the blood circulation. Moreover, we showed that i.m. injection of scAAV9 vectors expressing 'survival of motor neuron' (Smn) in spinal muscular atrophy (SMA) mice mediated high survival motor neuron (SMN) expression levels at both the CNS and the periphery, and increased the median lifespan from 12 days to 163 days. These findings represent to date the longest extent in survival obtained in SMA mice following i.m. viral vector gene delivery, and might generate a renewed interest in the use of i.m. adeno-associated viruses (AAV) delivery for the development of gene therapy strategies for MN diseases. [ABSTRACT FROM AUTHOR]

CD9 tetraspanin is the only egg membrane protein known to be essential for fertilization. To investigate its role, we have measured, on a unique acrosome reacted sperm brought in contact with an egg, the adhesion probability and strength with a sensitivity of a single molecule attachment. Probing the binding events at different locations of wild-type egg we described different modes of interaction. Here, we show that more gamete adhesion events occur on Cd9 null eggs but that the strongest interaction mode disappears. We propose that sperm-egg fusion is a direct consequence of CD9 controlled sperm-egg adhesion properties. CD9 generates adhesion sites responsible for the strongest of the observed gamete interaction. These strong adhesion sites impose, during the whole interaction lifetime, a tight proximity of the gamete membranes, which is a requirement for fusion to take place. The CD9-induced adhesion sites would be the actual location where fusion occurs. [ABSTRACT FROM AUTHOR]

Background: Following entry, uncoating, and reverse transcription, a number of cellular proteins become associated with the Human Immunodeficiency Virus type 1 (HIV-1) pre-integration complex (PIC). With the goal of obtaining reagents for the analysis of the HIV-1 PIC composition and localisation, we have constructed functional integrase (IN) and matrix (MA) proteins that can be biotinylated during virus production and captured using streptavidin-coated beads. Results: Although the labelled C-terminus allows for the sensitive detection of virion-associated IN, it becomes inaccessible in the presence of cellular proteins. This masking is not dependent on the nature of the tag and does not occur with the tagged MA. It was not observed either with an IN mutant unable to interact with LEDGF/p75, or when LEDGF/p75 was depleted from cells. Conclusion: Our observation suggests that a structural rearrangement or oligomerization of the IN protein occurs during the early steps of infection and that this process is related to the presence of LEDGF/p75. [ABSTRACT FROM AUTHOR]

Objective: To examine the relationship between seminal polymorphonuclear (PMN) granulocytes and sperm functions (particularly sperm motility) in infertile men. Design: A prospective clinical study. Setting: Assisted fertilization program in an academic research environment in France. Patient(s): Infertile men (n = 138) evaluated and classified as follows: group A with no detectable PMN in semen, group B with less than 0.5 × 106/mL, group C with 0.5-1 × 106/mL, and group D > 1 × 106/mL. Intervention(s): Seminal PMN, elastase concentrations, and sperm characteristics were analyzed. Seminal biochemical markers (free L-carnitine, citrate, zinc, acid phosphatase, and fructose) were measured. Main Outcome Measure(s): Relation between seminal markers and sperm motility. Result(s): Grade “b” motility and epididymal carnitine were statistically significantly increased in group C. Acid phosphatase was the only seminal marker to be statistically significantly impaired in group D. Elastase was proportional to the degree of leukocytospermia. Conclusion(s): These results suggest that sperm PMN originate from two regions: [1] the epididymis, where in small numbers and activated they may play a favorable role in sperm quality, and the [2] prostate, where their presence in large numbers, reflecting prostatitis, induces decreased secretion, especially of acid phosphatase. [Copyright &y& Elsevier]

Abstract: Fertilization includes a series of cellular interactions culminating with the fusion of gamete membranes, creating a zygote. Two ADAM proteins present on sperm, fertilin β and cyritestin, drew much attention. However, gene deletion in mice showed that fusion can happen in their absence. The presence of the integrin α6β1 on egg, a putative fertilin β receptor, is also dispensable. In contrast, sperm lacking Izumo, a molecule with a single Ig domain, are unable to fuse. On the egg side, a role for GPI-anchored molecules has been shown, and in mice lacking both tetraspanins CD9 and CD81 fertilization is completely blocked. [Copyright &y& Elsevier]

Abstract: In somatic cells, the tetraspanins CD81 and CD9 associate with each other, with additional tetraspanins and with non-tetraspanin molecules to form proteolipidic complexes. Here we show that CD81 is expressed on the surface of oocytes where it associates with tetraspanin-enriched membrane structures. A major CD9 and CD81 partner, CD9P-1, is also expressed by oocytes. Deletion of CD81 gene in mice results in a 40% reduction of female fertility. In vitro insemination indicated that this infertility is due to a deficiency of oocytes to fuse with sperm. While the fertility of CD9−/− mice is severely but not completely impaired, double knock-out CD9−/− CD81−/− mice were completely infertile indicating that CD9 and CD81 play complementary roles in sperm–egg fusion. Finally, a fraction of CD9 was transferred from CD81−/− oocytes to sperm present in the perivitelline space indicating that the defect of fusion of CD81−/− oocytes does not result from an impaired initial gamete interaction. [Copyright &y& Elsevier]

BACKGROUND: α6β1 integrin has been proposed to act as a sperm receptor on the mouse oocyte by interacting with spermatozoon fertilin β. We investigated, in humans, whether oocyte integrins could act similarly in gamete fusion, using a cyclic peptide containing the putative disintegrin-binding domain of human fertilin β [cyclic FEE (cFEE)] and RGD peptide. METHODS: Zona-free eggs were inseminated in the absence or presence of peptides. To maintain the membrane protein pattern, the zona pellucida was removed by microdissection. Immunofluorescence and confocal microscopy were used to detect integrin subunits on the oocyte. RESULTS: Unexpectedly, cFEE alone increased human gamete fusion by 94% instead of inhibiting fertilization. Furthermore, cFEE together with RGD potentiated the RGD-induced inhibition of fertilization in a dose-dependent manner. The data suggested the hypothesis of integrin cross-talk, further supported by the co-localization of α6β1 and αvβ3 integrins, the putative receptors of cFEE and RGD peptides, respectively. CONCLUSIONS: RGD-sensitive and -insensitive integrins may be associated in a multimolecular complex working as a sperm receptor on the human oocyte membrane. Supplementation of human IVF culture medium with cFEE peptide might improve fertilization rates in ART. [ABSTRACT FROM AUTHOR]

Rubus ulmifolius Schott commonly known as wild blackberry, is a perennial shrub native to North Africa and Europe. It has been used for both medicinal and food purposes. Different parts of the plant have been used in herbal medicine for a long time to treat numerous diseases such as diabetes, hypertension, gastro-intestinal disorders, mouth ulcers, heart ailments, menstrual pain, hemorrhoids, tonsillitis, skin diseases, etc. Its fruits are consumed fresh or in jams, juices, and marmalades. The shoots are used in omelets and soups. The present review aims to summarize data on ethnomedicinal uses, phytochemistry, pharmacological effects, botanical description, geographical distribution, and toxicity studies of Rubus ulmifolius over a period covering the last 23 years using online scientific databases such as Science direct, Google Scholar, Scopus, PubMed and Springer. Various studies have shown that Rubus ulmifolius possesses different biological activities in vitro and in vivo, including antipyretic, anti-inflammatory, anthelmintic, anticoagulant, antiproliferative, antioxidant, wound healing, amoebicidal, antifungal, antiviral, antibacterial, antihyperglycemic, and antihyperlipidemic properties. Phytochemical studies of Rubus ulmifolius have revealed the presence of various constituents belonging to different classes, such as flavonoids, tannins, terpenoids, and phenolic acids. We hope that our review will serve as an important source of information that can trigger further scientific research on this species. [ABSTRACT FROM AUTHOR]

• P. granatum exerted a significant preventive antithrombotic action (80 % protection) in mice. • The extract inhibited platelet aggregation induced in vitro by all agonists in a dose-dependent manner, prolonged bleeding time, and coagulation times in rats. The results of the ex vivo study in rats confirmed these effects. • We also noted that the extract did not exhibit toxicity following administration, and HPLC analysis revealed a richness of polyphenols and flavonoids in the extract. • P. granatum exhibited an interesting antithrombotic activity, which could be attributed to its antiplatelet and anticoagulant actions. The phytochemical compounds identified in this extract could, at least in part, be responsible for these effects. • Therefore, the demonstrated activities of this plant on hemostasis could lead to the development of more effective remedies and represent a means of prevention against thrombotic events. Hemostasis is the physiological process that stops bleeding by forming a thrombus (or blood clot). However, the inappropriate formation of a thrombus can constitute a pathophysiological event involving the same steps as those of hemostasis. Despite some adverse effects, antithrombotic therapy remains an effective means in the prevention and treatment of thrombotic diseases. In this regard, scientific research aimed at discovering new remedies of natural origin is particularly active. This study aims to evaluate the antithrombotic activity of the aqueous extract of Punica granatum peels in vivo. To understand the mechanism of action of this activity, we will investigate primary and secondary hemostasis in vitro and ex vivo. Furthermore, we will examine its antioxidant activity, toxicity, and phytochemical composition to identify the molecules responsible for the observed effects. The antithrombotic activity is assessed in vivo using the acute pulmonary thromboembolism model in mouse. Thrombosis is induced by injecting the thrombogenic solution (epinephrine + collagen) into the lateral caudal vein. Platelet aggregation, both in vitro and ex vivo , is performed on washed platelets isolated by centrifugation from rat blood. The platelet suspension is pre-incubated with the extract for 1 min. Aggregation is triggered by adding the following agonists: ADP, thrombin, collagen, and arachidonic acid. The bleeding time is measured in the absence and presence of the extract following an apical section of the mouse tail. This time corresponds to the duration of flow between the first and last drops of blood. To evaluate plasma coagulation, activated partial thromboplastin time (aPTT) and prothrombin time (PT) were measured under conditions both with and without the extract, using appropriate reagents. Acute toxicity is conducted by administering a single oral dose of the extract to mice (5 and 10 g kg-1). Possible changes in animal behavior are observed for fifteen days. The identification of polyphenols and flavonoids is performed by High Performance Liquid Chromatography (HPLC). The results obtained show that P. granatum exerted a significant preventive antithrombotic action (80 % protection) in mice. Furthermore, the extract inhibited platelet aggregation induced in vitro by all agonists in a dose-dependent manner, prolonged bleeding time, and coagulation times in rats. The results of the ex vivo study in rats confirmed these effects. We also noted that the extract did not exhibit toxicity following administration, and HPLC analysis revealed a richness of polyphenols and flavonoids in the extract. P. granatum exhibited an interesting antithrombotic activity, which could be attributed to its antiplatelet and anticoagulant actions. The phytochemical compounds identified in this extract could, at least in part, be responsible for these effects. Therefore, the demonstrated activities of this plant on hemostasis could lead to the development of more effective remedies and represent a means of prevention against thrombotic events. [Display omitted] [ABSTRACT FROM AUTHOR]

• P. granatum exerted a significant preventive antithrombotic action (80 % protection) in mice. • The extract inhibited platelet aggregation induced in vitro by all agonists in a dose-dependent manner, prolonged bleeding time, and coagulation times in rats. The results of the ex vivo study in rats confirmed these effects. • We also noted that the extract did not exhibit toxicity following administration, and HPLC analysis revealed a richness of polyphenols and flavonoids in the extract. • P. granatum exhibited an interesting antithrombotic activity, which could be attributed to its antiplatelet and anticoagulant actions. The phytochemical compounds identified in this extract could, at least in part, be responsible for these effects. • Therefore, the demonstrated activities of this plant on hemostasis could lead to the development of more effective remedies and represent a means of prevention against thrombotic events. Hemostasis is the physiological process that stops bleeding by forming a thrombus (or blood clot). However, the inappropriate formation of a thrombus can constitute a pathophysiological event involving the same steps as those of hemostasis. Despite some adverse effects, antithrombotic therapy remains an effective means in the prevention and treatment of thrombotic diseases. In this regard, scientific research aimed at discovering new remedies of natural origin is particularly active. This study aims to evaluate the antithrombotic activity of the aqueous extract of Punica granatum peels in vivo. To understand the mechanism of action of this activity, we will investigate primary and secondary hemostasis in vitro and ex vivo. Furthermore, we will examine its antioxidant activity, toxicity, and phytochemical composition to identify the molecules responsible for the observed effects. The antithrombotic activity is assessed in vivo using the acute pulmonary thromboembolism model in mouse. Thrombosis is induced by injecting the thrombogenic solution (epinephrine + collagen) into the lateral caudal vein. Platelet aggregation, both in vitro and ex vivo , is performed on washed platelets isolated by centrifugation from rat blood. The platelet suspension is pre-incubated with the extract for 1 min. Aggregation is triggered by adding the following agonists: ADP, thrombin, collagen, and arachidonic acid. The bleeding time is measured in the absence and presence of the extract following an apical section of the mouse tail. This time corresponds to the duration of flow between the first and last drops of blood. To evaluate plasma coagulation, activated partial thromboplastin time (aPTT) and prothrombin time (PT) were measured under conditions both with and without the extract, using appropriate reagents. Acute toxicity is conducted by administering a single oral dose of the extract to mice (5 and 10 g kg-1). Possible changes in animal behavior are observed for fifteen days. The identification of polyphenols and flavonoids is performed by High Performance Liquid Chromatography (HPLC). The results obtained show that P. granatum exerted a significant preventive antithrombotic action (80 % protection) in mice. Furthermore, the extract inhibited platelet aggregation induced in vitro by all agonists in a dose-dependent manner, prolonged bleeding time, and coagulation times in rats. The results of the ex vivo study in rats confirmed these effects. We also noted that the extract did not exhibit toxicity following administration, and HPLC analysis revealed a richness of polyphenols and flavonoids in the extract. P. granatum exhibited an interesting antithrombotic activity, which could be attributed to its antiplatelet and anticoagulant actions. The phytochemical compounds identified in this extract could, at least in part, be responsible for these effects. Therefore, the demonstrated activities of this plant on hemostasis could lead to the development of more effective remedies and represent a means of prevention against thrombotic events. [Display omitted] [ABSTRACT FROM AUTHOR]

The process of gamete fusion has been largely studied in the mouse and has revealed the crucial role of the tetraspanin CD9. By contrast, human gamete fusion remains largely unknown. We now show that an anti-α 6 integrin mAb (GoH3) strongly inhibited human sperm-egg fusion in human zona-free eggs. Furthermore, a mAb directed against CD151, a tetraspanin known to associate with α 6β1, partially inhibited sperm-egg fusion. By contrast, the addition of an anti-CD9 mAb to zona free eggs had no effect. The integrin α 6β1 CD151 and CD9 tetraspanins were evenly distributed on human zona-intact oocytes. On zona-free eggs, the integrin α 6β1 and tetraspanin CD151 patched and co-localized but the tetraspanin CD9 remained unchanged. CD9 mAb prevented α 6β1 integrin clustering and gamete fusion when added prior to, but not after, zona removal. Antibody-mediated aggregation of integrin α 6β1 yielded patches that were bigger and more heterogeneous in mouse oocytes lacking CD9. Moreover, a strong labelling of α 6β1 could be observed at the sperm entry point. Altogether, these data show that CD9 controls the redistribution of some membrane proteins including the α 6β1 integrin into clusters that may be necessary for gamete fusion. [ABSTRACT FROM AUTHOR]

Juniperus oxycedrus L. (J. oxycedrus) has a rich historical background in herbal remedies to treating digestive system abnormalities. However, no comprehensive evaluation of its potential toxic effects has been conducted. The current investigation aimed to evaluate the acute and subacute toxicity of an aqueous extract of J. oxycedrus (AEJO). AEJO was prepared by the conventional Moroccan methods by decoction the arial part of the plant. The acute and subacute toxicity tests were conducted in mice and rats, respectively. Acute toxicity tests showed that the extract was not toxic even at high doses of 5000 mg/kg. In the subacute study, no detectable indications of toxicity or mortality were observed and there were no notable deviations in food intake or water consumption among all rats. However, changes in body weight of animals treated with 1000 and 2000 mg/kg underwent a significant decrease. AEJO administration decreased platelet number, elevated levels of alanine aminotransferase and alkaline phosphatase, and reduced albumin levels. Histological examination revealed normal renal parenchyma despite increased creatinine. It also showed binucleation, and hepatocyte vacuolation. The results indicate that AEJO has considerable tolerance for consumption, but repeated use can affect hepatocytes and kidneys. Therefore, additional analyses, such as subchronic, chronic, and neurotoxic studies, are required before using this plant in clinical research. [Display omitted] [ABSTRACT FROM AUTHOR]

Thrombosis is the formation of a clot within a blood vessel. Antithrombotic drugs are used for treating thrombosis, which can be the cause of hemorrhage. Currently, there is a need to discover novel antithrombotic drugs. Walnut is widely used to treat a wide range of health complaints. In this study, walnut bark extract was tested in hemostasis parameters: platelets adhesion, aggregation, and plasmatic coagulation in human blood. The crude aqueous extract of walnut bark was prepared by infusion and tested in vitro on hemostasis. Through blood collection from healthy volunteer donors, we studied different parameters of the primary hemostasis: platelet adhesion on the collagen-coated surface under flow, ADP, collagen, thrombin, and arachidonic acid-induced platelet aggregation, and of the secondary hemostasis by measuring prothrombin time (PT) and activated partial thromboplastin (APTT) parameters. All experiments are realized in the absence and presence of the extract and repeated at least twice. The obtained data showed that the extract (1 and 2 mg/mL) significantly (p < 0.001) reduced the activated platelet adhesion on the collagen-coated surface. In the same way, the effect of the extract on platelet aggregation seems to depend on its concentration and on the nature of the agonist. The strongest inhibition of aggregation was observed in the case of collagen at 1 mg/mL, while there was no observed effect on arachidonic acid-induced aggregation. Moreover, the extract (1 mg/mL) affects the extrinsic, intrinsic, and common pathways of the human blood coagulation cascade by extending significantly (p < 0.001), both PT and APTT times. This study provides evidence that walnut bark extract, by its antiadhesive, antiaggregant, and anticoagulant activities, could be considered as a serious source of biological compounds for the prevention and treatment of thrombosis. [ABSTRACT FROM AUTHOR]

A rapid, easy and simple method for the isolation and purification of α-glucosidase inhibitors of the ethyl acetate extract of Thymelaea hirsuta (EaTh) by a combination of thin layer chromatography (TLC) and enzymatic test has been developed. EaTh was demonstrated previously a potent α-glucosidase inhibitory effect. In this study, we developed a simple TLC-enzymatic test (TLC/EZ) combination to isolate α-glucosidase inhibitors present in EaTh.EaTh was extracted by Soxhlet from Thymelaea hirsuta (T. hirsuta). The EaTh was separated on a silica gel column and then on a TLC plate. After TLC separation, the TLC/EZ combination method was applied. α-glucosidase inhibitors were detected directly in the TLC plate using the glucose oxidase peroxidase method (GOD-POD). A good detection of active compounds was obtained in the TLC favoring the TLC/EZ method. Active compounds were then characterized using high performance liquid chromatography-mass spectrometry (HPLC–MS) analysis. The main α-glucosidase inhibitors present in EaTh have a molecular ion [M + H]+ at m/z = 543. This proposed method is suitable for a reliable isolation and purification of α-glucosidase inhibitors present in EaTh. It could be proposed as an interesting alternative of the classical method for the isolation and purification of α-glucosidase inhibitors in plant extracts. [ABSTRACT FROM AUTHOR]

Fertilization is a complex process that requires successive stages and culminates in the adhesion/fusion of gamete membranes. If the question of the involvement of oocyte integrins has been swept away by deletion experiments, that of the involvement of sperm integrins remains to be further characterized. In the present study, we addressed the question of the feasibility of sperm–oocyte adhesion/fusion and early implantation in the absence of sperm β1 integrin. Males and females with β1 integrin-depleted sperm and oocytes were mated, and fertilization outcome was monitored by a gestational ultrasound analysis. Results suggest that although the sperm β1 integrin participates in gamete adhesion/fusion, it is dispensable for fertilization in mice. However, sperm- and/or oocyte-originated integrin β1 is essential for post-implantation development. Redundancy phenomena could be at the origin of a compensatory expression or alternative dimerization pattern. [ABSTRACT FROM AUTHOR]

SPACA6 is a sperm-expressed surface protein that is critical for gamete fusion during mammalian sexual reproduction. Despite this fundamental role, little is known about how SPACA6 specifically functions. We elucidated the crystal structure of the SPACA6 ectodomain at 2.2-Å resolution, revealing a two-domain protein containing a four-helix bundle and Ig-like β-sandwich connected via a quasi-flexible linker. This structure is reminiscent of IZUMO1, another gamete fusion-associated protein, making SPACA6 and IZUMO1 founding members of a superfamily of fertilization-associated proteins, herein dubbed the IST superfamily. The IST superfamily is defined structurally by its distorted four-helix bundle and a pair of disulfide-bonded CXXC motifs. A structure-based search of the AlphaFold human proteome identified more protein members to this superfamily; remarkably, many of these proteins are linked to gamete fusion. The SPACA6 structure and its connection to other IST-superfamily members provide a missing link in our knowledge of mammalian gamete fusion. The crystal structure of sperm-expressed surface protein SPACA6 shows similarities to IZUMO1 though their functions are not redundant. Structural homology search in the AlphaFold human proteome and sequence search suggest a superfamily of mammalian gamete fusion-associated proteins. [ABSTRACT FROM AUTHOR]

The article focuses on how the actin-spectrin submembrane scaffold regulates endocytosis along the proximal axon by restricting clathrin-coated pit (CCP) formation. Topics include the nanoscale architecture of CCPs at the axon initial segment (AIS), the role of the dense spectrin mesh in limiting endocytosis, and how endocytosis can be triggered by physiological stimuli such as N-Methyl-D-Aspartate (NMDA).

Duchenne muscular dystrophy is a severe neuromuscular disease causing a progressive muscle wasting due to mutations in the DMD gene that lead to the absence of dystrophin protein. Adeno-associated virus (AAV)-based therapies aiming to restore dystrophin in muscles, by either exon skipping or microdystrophin expression, are very promising. However, the absence of dystrophin induces cellular perturbations that hinder AAV therapy efficiency. We focused here on the impact of the necrosis-regeneration process leading to nuclear centralization in myofiber, a common feature of human myopathies, on AAV transduction efficiency. We generated centronucleated myofibers by cardiotoxin injection in wild-type muscles prior to AAV injection. Intramuscular injections of AAV1 vectors show that transgene expression was drastically reduced in regenerated muscles, even when the AAV injection occurred 10 months post-regeneration. We show also that AAV genomes were not lost from cardiotoxin regenerated muscle and were properly localised in the myofiber nuclei but were less transcribed leading to muscle transduction defect. A similar defect was observed in muscles of the DMD mouse model mdx. Therefore, the regeneration process per se could participate to the AAV-mediated transduction defect observed in dystrophic muscles which may limit AAV-based therapies. [ABSTRACT FROM AUTHOR]

Abstract: Objective: Argan oil has been shown to inhibit in vitro and ex vivo platelet aggregation without extending bleeding time. In this report, we examined in vivo the antithrombotic activity of argan oil in an experimental thrombosis model in mice: acute pulmonary thromboembolism and in vitro its effect in a coagulation assay. Methods: Acute pulmonary thromboembolism was induced, after argan oil treatment, by an intravenous injection of a collagen and epinephrine mixture. The paralyzed and dead mice in each group were numbered and the percentage of protection against acute pulmonary thromboembolism was calculated. The histologic study was conducted in lung tissue to estimate the percentage of opened and occluded vessels by platelet thrombi. The coagulation assay was monitored in platelet-poor plasma from normal rats by measuring the clotting parameters (activated partial thromboplastin time, prothrombin time, and thrombin time) in the presence and absence of argan oil. Results: Argan oil (1 mL/100 g/day), administered orally, showed an antithrombotic activity preventing the paralysis or death (50%) induced by the collagen–epinephrine intravenous injection. This observation was confirmed by the lung histologic examination, in which the density of occluded blood vessels was significantly decreased (62.16 ± 3.95%). However, the argan oil remained inactive for the coagulation parameters of activated partial thromboplastin time, prothrombin time, and thrombin time at variance with heparin, an anticoagulant reference drug. The antithrombotic activity of argan oil seemed unrelated to the anticoagulant activity. Conclusion: We suggest that argan oil might be an interesting natural dietary source for the nutritional prevention of hemostasis and cardiovascular disorders. Clinical trials would be necessary and relevant to confirm this hypothesis. [Copyright &y& Elsevier]

Abstract: Objective: Argan oil has been shown to inhibit in vitro and ex vivo platelet aggregation without extending bleeding time. In this report, we examined in vivo the antithrombotic activity of argan oil in an experimental thrombosis model in mice: acute pulmonary thromboembolism and in vitro its effect in a coagulation assay. Methods: Acute pulmonary thromboembolism was induced, after argan oil treatment, by an intravenous injection of a collagen and epinephrine mixture. The paralyzed and dead mice in each group were numbered and the percentage of protection against acute pulmonary thromboembolism was calculated. The histologic study was conducted in lung tissue to estimate the percentage of opened and occluded vessels by platelet thrombi. The coagulation assay was monitored in platelet-poor plasma from normal rats by measuring the clotting parameters (activated partial thromboplastin time, prothrombin time, and thrombin time) in the presence and absence of argan oil. Results: Argan oil (1 mL/100 g/day), administered orally, showed an antithrombotic activity preventing the paralysis or death (50%) induced by the collagen–epinephrine intravenous injection. This observation was confirmed by the lung histologic examination, in which the density of occluded blood vessels was significantly decreased (62.16 ± 3.95%). However, the argan oil remained inactive for the coagulation parameters of activated partial thromboplastin time, prothrombin time, and thrombin time at variance with heparin, an anticoagulant reference drug. The antithrombotic activity of argan oil seemed unrelated to the anticoagulant activity. Conclusion: We suggest that argan oil might be an interesting natural dietary source for the nutritional prevention of hemostasis and cardiovascular disorders. Clinical trials would be necessary and relevant to confirm this hypothesis. [ABSTRACT FROM AUTHOR]

Objective: To assess the effect of leukocytospermia on assisted reproductive technology outcomes. Design: Retrospective analysis. Setting: University laboratory. Patient(s): Couples attending the infertiliy clinic and involved in ART program for IVF or ICSI. Intervention(s): During a 7-year follow-up in an assisted reproductive technology program, leukocytospermia was routinely determined using the peroxidase technique. Donor sperm were excluded from the study. Main Outcome Measure(s): Egg retrievals (N = 3,508) were distributed in 3 groups according to the leukocyte levels in semen from which fertilizing sperm were extracted: group 1, absence of leukocytes (n = 3,026); group 2, moderate leukocytospermia (<106/mL) (n = 344); or group 3, high leukocytospermia (≥106/mL) (n = 138). They resulted in 1,463 IVF and 2,045 intracytoplasmic sperm injection procedures that gave 802 clinical pregnancies. Result(s): Surprisingly, the fertilization rate, cleavage rate, clinical pregnancy rate, gestational age, and mean infant weight were significantly improved when seminal leukocytes were present, regardless of the technique used. The only negative side effects associated with a high level of seminal leukocytes (group 3) were an elevated rate of early pregnancy loss (from 26.6% to 40.5%) and a 3-fold increase in the percentage of ectopic pregnancies. Conclusion(s): At moderate levels (<106/mL), leukocytospermia appears to be physiologic. It is associated with improved sperm fertilization ability and pregnancy outcome. At higher concentrations, leukocytospermia alters neither sperm fertilization ability nor the probability of clinical pregnancy when compared with nonleukocytic patients with infertility. However, the pregnancy outcome is reduced. [Copyright &y& Elsevier]

BACKGROUND We previously demonstrated in a small pilot study that oral medroxyprogesterone acetate and percutaneous testosterone (OMP/PT) induce reversible spermatogenesis suppression. The aims of this study were to determine the rate of spermatogenetic inhibition and recovery and to obtain preliminary data on efficacy for a larger population under OMP/PT. METHODS A total of 35 healthy men with normal spermiograms requesting male hormonal contraception were treated with OMP (20 mg/day) and PT (50–125 mg/day) for periods up to 18 months. Couples were included in a contraceptive efficacy phase after a value of ≤1 million/ml spermatozoa was reached between 1 and 3 months of treatment. RESULTS Sperm counts decreased by 47% at 1 month, reaching 90% at 2 months and 98–100% between 4 and 8 months. At 3 months, 80% of men had ≤1 million/ml spermatozoa. Follicle-stimulating hormone and luteinizing hormone decreased to 35% of pretreatment levels after 1 month of treatment and to 75–80% at 2 and 6 months, respectively. Plasma testosterone and estradiol levels were in the eugonadal range at 3, 6, 9 and 12 months of treatment. Dihydrotestosterone concentrations were 2–4 times higher than pretreatment values. The rate of spermatogenetic recovery was rapid (73 ± 29.5 days). During the efficacy phase (211 months for 25 couples), one pregnancy attributable to poor compliance of the male partner was observed. CONCLUSIONS OMP/PT efficiently inhibits spermatogenesis in 80% of men, maintains testosterone at physiological levels and avoids the need for parenteral administration, which is poorly accepted by French men. These results justify larger studies to define a more adequate dosage of OMP/PT and to confirm its efficacy and safety. [ABSTRACT FROM PUBLISHER]

The goal of the present study is to test the effect of water extract (WE) of four medicinal plants used as antidiabetics in Eastern Morocco (Arbutus unedo: Au, Ammoïdes pusilla: Ap, Thymelaea hirsuta: Th, and Urtica dioïca: Ud). These plants are used in cooking to bring out the flavor in a dish or to complement it. The first experiment was realized in order to determine the antidiabetic effect of the WE of these plants during 5 weeks' treatment. Seven groups of Wistar rats were used: Healthy controls, neonatal streptozotocin (n-stz) induced-diabetic rats (90 mg/kg; intraperitoneally [i.p.]), n-stz + tolbutamide (400 mg/l), and 4 groups n-stz + WE of plants (400 mg/l, drink water). The percentages of Plasma glucose lowering effect were, respectively for Au, Ap, Th, Ud and tolbutamide: 31.6 % p<0.01, 27.4 % p<0.05, 38.2 % p<0.01, 13 % and 33.9 % p<0.05 when compared with untreated diabetic controls. In a second experiment, oral glucose tolerance tests were carried out in n-stz induced-diabetic rats. The i.p. administration of the water extract (WE) of Ap and Ud (150 mg/kg) 30 minutes before the glucose overload (2 g/kg) showed a significant reduction glycemia, respectively of 36 % at 60 min (p<0.05) and 50 % at 180 min (p<0.05) after glucose overload compared with controls. In contrast, the effect of WE of Au and Th (150 mg/kg, i.p.) was not significant. The in vitro study of glucose utilization by isolated rat hemidiaphragm suggests that these extracts in combination with insulin potentiate its activity and enhance the utilization of glucose. In conclusion, it seems that these plants possess antidiabetic activity. [ABSTRACT FROM AUTHOR]

The article presents a study that analyzed the link of CD9 tetraspanin and glycosyl-phosphatidylinositol (GPI)-anchored proteins on the sperm-egg interaction on mice. It discusses the structures, functions, and the roles of the proteins in the oocyte membrane. Two hypotheses were proposed, the first one is that the proteins are involved in signaling cascade and the other one is that they are involved in the organization of plasma membrane. It concludes that their absence affects gamete fusion.

Abstract: Cistus ladaniferus L. (Cistaceae) is a medicinal plant originated from the Mediterranean region which exerts different pharmacological effects. In the present study, our goal was to examine whether the plant possessed antihypertensive properties. Aqueous extract of Cistus leaves (AEC, 500mg/kg/day) reduced systemic blood pressure (SBP) in two animal models of hypertension, the l-NAME and renovascular two kidney-one clip (2K-1C) hypertensive rats. In the former, AEC prevented the increase in SBP when co-administered with l-NAME during four weeks (164±3mm Hg in l-NAME vs. 146±1mm Hg in l-NAME+AEC, p <0.001). In the latter, AEC reversed the increase in SBP when administered during four weeks after installation of the hypertension (146±5mm Hg with AEC vs. 179±6mm Hg without, p <0.05). AEC treatment also reversed the endothelial dysfunction observed in both animal models of hypertension. A direct effect on cardiac and vascular tissue was also tested by examining the contractile effects of AEC in rat isolated aortic rings and Langendorff perfused hearts. AEC (10mg/L) had no effect on left ventricular developed pressure and heart rate in isolated perfused heart. However, AEC produced a strong relaxation of pre-contracted rat aortic rings (80±2% relaxation, n =25). When the rings were denuded from endothelium or were incubated with 1mM Nω-nitro-l-arginine (l-NNA), the relaxant effect of AEC was lost. We conclude that C. ladaniferus possesses antihypertensive properties which are mainly due to an endothelium-dependent vasodilatory action. [Copyright &y& Elsevier]

When administered 30 min before glucose loading, the aqueous extract of Urtica dioica (nettle) (250 mg/kg) showed a strong glucose lowering effect. The decrease of glycemia has reached to 33±3.4% of the control value 1 h after glucose loading. This effect was persistent during 3 h. In contrast, nettle did not show hypoglycemic effect in alloxan-induced diabetic rats. The amount of glucose absorbed in a segment jejunum in situ was 8.05±0.68 mg in presence of nettle extract vs. 11.11±0.75 mg in control rats during 2 h (P<0.05). The results indicate that nettle has a significant antihyperglycemic effect in OGTT model. This effect may be caused in part by the reduction of intestinal glucose absorption. LD50 is 3.5 g/kg (i.p.). [Copyright &y& Elsevier]

The synthesis of new functionalized tetrazole–pyrazole compounds is reported. They were fully characterized by spectroscopy and spectrometry methods. The vasorelaxant potency of these molecules was also investigated. All compounds showed a good vasorelaxant activity but the Compound 6 "ethyl 1‐((2‐(3‐bromopropyl)‐2H‐tetrazol‐5‐yl)methyl)‐5‐methyl‐1H‐pyrazole‐3‐carboxylate" seems to have the highest effect, which reached 70% at 10−4 M concentration. This effect was partially endothelium‐dependent; also, the vasorelaxant pattern of this compound was quite similar to that of the verapamil. [ABSTRACT FROM AUTHOR]

Opuntia ficus-indica (L.) Mill is frequently observed in the Moroccan traditional medicinal system, where these approaches are employed to mitigate the onset of diabetes and the subsequent complications it may entail. The aim of this research was to examine the effectiveness of Opuntia ficus-indica seed oil in preventing diabetic complications. Specifically, the study assessed its ability to counteract glycation at various stages, protected red blood cells from the harmful effects of glycated albumin, and inhibited pancreatic lipase digestive enzymes to understand its potential antihyperglycemic properties. Additionally, the study aimed to identify the chemical components responsible for these effects, evaluate antioxidant and anti-inflammatory properties, and conduct computational investigations such as molecular docking. The assessement of Opuntia ficus-indica seed oil antiglycation properties involved co-incubating the extract oil with a bovine serum albumin-glucose glycation model. The study investigated various stages of glycation, incorporating fructosamine (inceptive stage), protein carbonyls (intermediate stage), and AGEs (late stage). Additionally, measurement of β-amyloid aggregation of albumin was performed using Congo red, which is specific to amyloid structures. Additionally, the evaluation of oil's safeguarding effect on erythrocytes against toxicity induced by glycated albumin included the measurement of erythrocyte hemolysis, lipid peroxidation, reduced glutathione. The fatty acid of Opuntia ficus-indica seed oil were analyzed using Gas Chromatography-Mass Spectrometry (GC–MS). The in vitro evaluation of antihyperglycemic activity involved the use of pancreatic lipase enzyme, while the assessement of antioxidant capability was carried out through the utilization of the ABTS and FRAP methods. The in vitro assessement of the denaturation of albumin activity was also conducted. In conjunction with the experimental outcomes, computational investigations were undertaken, specifically employing ADMET (absorption, distribution, metabolism, excretion, and toxicity) analysis. Furthermore, molecular docking was utilized to predict antioxidant and antiglycation mechanisms based on protein targets. In vitro glycation assays, Opuntia ficus-indica seed oil displayed targeted inhibitory effects at multiple distinct stages. Within erythrocytes, in addition to mitigating hemolysis and lipid peroxidation induced by glycated albumin. GC-MS investigation revealed a richness of fatty acids and the most abundant compounds are Linoleic acid (36.59%), Palmitic acid (20.84%) and Oleic acid (19.33%) respectively. The findings of antioxidant ability showed a remarkable activity on FRAP and ABTS radicals. This oil showed a pronounced inhibitory impact (p < 0.001) on pancreatic lipase enzyme. It also exerted a notibale inhibition of albumin denaturation, in vitro. The identified results were supported by the abundant compounds of fatty acids unveiled through GC-MS analysis, along with the computational investigation and molecular docking. [Display omitted] • The fatty acids derived from Opuntia ficus-indica (L.) Mill oil play diverse pharmacological roles. • Fatty acids compounds were responsible for the antiglycation activity observed in proteins, including albumin and hemoglobin. • The glycation of albumin induces erythrocyte toxicity, which is inhibited by the oil of Opuntia ficus-indica (L.) Mill. • The predominant components of Opuntia ficus-indica (L.) Mill oil were palmitic acid, oleic acid, and linoleic acid. [ABSTRACT FROM AUTHOR]

Diabetes is a significant metabolic disease impacting many of the world's population. In Morocco, a wide range of medicinal plants has taken great importance in the treatment of diabetes, among these plants; we find Argania spinosa (L.) Skeels. The objective of our work is based on the evaluation of the effect of roasted (Roil) and unroasted (UnRoil) Argan seed oil on diabetic nephropathy. Roasted and unroasted oils from Argania spinosa (L.) Skeels seeds were examined for their effects on diabetic nephropathy using an experimental streptozotocin-induced model. Biochemical and histopathological analyses were conducted on blood and kidney samples to assess renal function and tissue damage. Both oils ameliorated significantly diabetic nephropathy symptoms. They limited the renal damage caused by streptozotocin and improved diabetes symptoms, including blood glucose levels, body weight, water intake, urinary volume, and kidney parameters. This activity could be elucidated by the antioxidant effect of Argan oil, enabling to neutralize free radicals and undertake a fundamental role in preventing the onset of these complications. Based on our findings, Argan oil could be used as dietary supplement for people with diabetes as a preventive measure against the emergence of diabetic complications. [Display omitted] [ABSTRACT FROM AUTHOR]

Origanum genus is one of the most widely used herbs in folk medicine for its biological properties. The present investigation aims to characterize, for the first time, endemic Origanum grosii collected from the Taounate region, Morocco. This characterization was realized by determining the concentration of metals in different parts of the plant (flowers, leaves, and stems) by ICP-MS, and the results were studied statistically by Principal Component Analysis (PCA). Phytochemical screening with the dosage of polyphenols and flavonoids has been conducted. To know more about this species, antihyperglycemic tests have been performed to highlight the presence or absence of the antidiabetic effect for this plant. An Oral Glucose Tolerance Test (OGTT) has been performed on normal mice which were divided into two groups of six mice each. Group 1 (control group) was treated with distilled water, and group 2 was treated with an aqueous extract of O. grosii by gavage at 150 mg/kg. Digestive enzyme α-amylase inhibition assay has also been evaluated to study the inhibition effect of the studied extract using acarbose as a control. The results showed that the leaves exhibited a high concentration of trace elements (Ca, Mg, and K) and total absence of heavy metals, which were found in small quantities (Cr, Ni, and B) on the stems, and this makes the plant safe to use. On the other hand, tannins, flavonoids, triterpenes, and steroids were the major families strongly present in this species. The antidiabetic results showed that O. grosii have significantly reduced postprandial hyperglycemia after glucose loading in normal rats. It showed also that this species has a significant antihyperglycemic activity reflected by the inhibition of α-amylase. The one responsible for this property could be the synergy between the trace elements and the nature of the chemical families of O. grosii species, which can make this plant useful in the management of postprandial hyperglycemia. [ABSTRACT FROM AUTHOR]

The present study aims to evaluate the hepatoprotective activity of stem aqueous extract of Caralluma europaea (AECe) on carbon tetrachloride- (CCl4-) induced hepatic damage in Wistar rats. The animals were daily treated with the aqueous extract of C. europaea at a dose of 250 mg/kg body weight for 14 days. CCl4 was injected (1 ml/kg, i.p.) two times, on the 7th and 14th days. At the end of the experimental period, all rats were anesthetized to collect blood for the assessment of biochemical parameters and then sacrificed to collect the liver for weighing. Hepatotoxicity was evaluated by measuring the serum levels of aspartate aminotransferase (AST), alanine aminotransferase (ALT), alkaline phosphatase (ALP), bilirubin (total and direct), malondialdehyde (MDA), total protein (TP), triglycerides (TG), total cholesterol, very low-density lipoprotein (VLDL-c), low-density lipoprotein (LDL-c), high-density lipoprotein (HDL-c), urea, creatinine, and uric acid. Based on the results obtained in this study, the administration of C. europaea before exposure to the administration of CCl4 conferred favorable hepatoprotective effect in rats. The treatment with AECe (250 mg/kg) exhibits a significant hepatoprotective effect by ameliorating CCl4-induced alterations of these biochemical parameters. Hence, C. europaea could be a potential medicinal herb that can be used in the future to prevent liver intoxication. [ABSTRACT FROM AUTHOR]

• Argania spinosa oils showed a significant inhibition of pancreatic ɑ-amylase. • Argania spinosa oils inhibit non-competitively intestinal ɑ-glucosidase. • Argania spinosa oils reduce intestinal glucose absorption. • The roasting process reduce the anti-hyperglycemic effect of the oils. Diabetes mellitus is a serious metabolic disease, which affects a large number of populations in the world. Phytotherapy is considered as the most remedies used by Moroccan people to treat this pathology. Argania spinosa L. (SKEELS) seeds oils is among the products that are used traditionally in Morocco for the anti-diabetic purposes. The aim of this study is to investigate the effect of roasted (Roil) and unroasted (UnRoil) Argan seeds oils on α-glucosidase, α-amylase and intestinal glucose absorption activities in vitro and in vivo. The antihyperglycemic effect of the roasted and unroasted Argania spinosa L. seeds oils was tested against α-glucosidase and α-amylase activities, in vitro, at the concentrations (82, 165, 328 and 656 µg/mL) and (0.9, 2.25, 4.5 and 9 mg/mL) respectively. In addition, the inhibitory effect of the oils against these enzymes was confirmed, in vivo , in normal and STZ- diabetic rats. The antihyperglycemic effect of these oils was also tested against intestinal d -glucose absorption activity at the dose of 1 mL/Kg using the jejunum segment perfusion technique, in situ. The results of this study were showed that the Roil and the UnRoil were significantly (p < 0.001) inhibited the pancreatic α-amylase, in vitro , with (IC 50 = 23, 98 mg/mL) and (IC 50 = 5, 87 mg/mL) respectively. The same effect of these oils was confirmed against the intestinal α-glucosidase activity, with (IC 50 = 0.081 mg/mL) and (IC 50 = 0.117 mg/mL) for UnRoil and Roil respectively. Besides, the inhibitory effect of these oils against these enzymes, in vitro , was confirmed, in vivo. The oral intake of these oils at a dose of (2 mL/Kg) was significantly attenuated the hyperglycemia induced by the sucrose (substrate of intestinal α-glucosidase) and the starch (substrate of pancreatic α-amylase), in the normal and STZ-diabetic rats. Although, the Roil and the UnRoil at the dose 1 mL/Kg were found that they significantly (p < 0.001) reduce the intestinal glucose absorption, in situ , with 50, 99% and 57, 97% respectively. The Argan oils showed a very high anti-hyperglycemic activity. This effect could be achieved by the inhibiting of the activity of pancreatic α-amylase, intestinal α-glucosidase and the intestinal absorption of D-glucose. Moreover, the UnRoil has shown a better anti-hyperglycemic activity than the Roil, which means that the torrefaction of the Argan seeds before the oil extraction, influence on the bioactive constituents responsible for this beneficial effect. [ABSTRACT FROM AUTHOR]

Three genes are known to be essential for gamete adhesion/fusion (Cd9, Izumo1 and Juno). Here, we confirmed that Spaca6 null males are infertile and showed that their sperm accumulate in the perivitelline space but are unable to fuse with oocyte. Like IZUMO1, SPACA6 which is expressed by human sperm, is remained on the equatorial segment after acrosomal reaction and is involved in human fertilization since an anti-SPACA6 antibody inhibited it. Despite the similarity of the phenotypes caused by Spaca6 and Izumo1 knockouts, these are not redundant and the essential relocation of IZUMO1 is not affected by the lack of SPACA6. We propose a model in which IZUMO1 and SPACA6 would be part of a molecular complex necessary for gamete fusion and that their concomitant presence would be required for the recruitment of another essential molecular actor, such as a fusogen, for the fusion to take place. [ABSTRACT FROM AUTHOR]

The adeno-associated virus (AAV) vector is an efficient tool for gene delivery in skeletal muscle. AAV-based therapies show promising results for treatment of various genetic disorders, including muscular dystrophy. These dystrophies represent a heterogeneous group of diseases affecting muscles and typically characterized by progressive skeletal muscle wasting and weakness and the development of fibrosis. The tropism of each AAV serotype has been extensively studied using systemic delivery routes, but very few studies have compared their transduction efficiency through direct intramuscular injection. Yet, in some muscular dystrophies, where only a few muscles are primarily affected, a local intramuscular injection to target these muscles would be the most appropriate route. A comprehensive comparison between different recombinant AAV (rAAV) serotypes is therefore needed. In this study, we investigated the transduction efficiency of rAAV serotypes 1–10 by local injection in skeletal muscle of control C57BL/6 mice. We used a CMV-nls-LacZ reporter cassette allowing nuclear expression of LacZ to easily localize targeted cells. Detection of β-galactosidase activity on muscle cryosections demonstrated that rAAV serotypes 1, 7, 8, 9, and 10 were more efficient than the others, with rAAV9 being the most efficient in mice. Furthermore, using a model of human muscle xenograft in immunodeficient mice, we observed that in human muscle, rAAV8 and rAAV9 had similar transduction efficiency. These findings demonstrate for the first time that the human muscle xenograft can be used to evaluate AAV-based therapeutical approaches in a human context. [ABSTRACT FROM AUTHOR]

A key aspect to consider for vaccinal protection is the induction of a local line of defense consisting of nonrecirculating tissue-resident memory T cells (TRM), in parallel to the generation of systemic memory CD8+ T cell responses. The potential to induce TRM has now been demonstrated for a number of pathogens and viral vectors. This potential, however, has never been tested for recombinant adeno-associated virus (rAAV) vectors, which are weakly inflammatory and poor transducer of dendritic cells. Using a model rAAV2/1-based vaccine, we determined that a single intradermal immunization with rAAV2/1 vectors in mice induces fully functional TRM at the local site of immunization. The optimal differentiation of rAAVinduced transgene-specific skin TRM was dependent on local transgene expression and additional CD4+ T cell help. Transgene expression in dendritic cells, however, appeared to be dispensable for the priming of transgene-specific skin TRM, suggesting that this process solely depends on the cross-presentation of transgene products. Overall, this study provides needed information to properly assess rAAV vectors as T cell-inducing vaccine carriers. [ABSTRACT FROM AUTHOR]

Abstract Ethnopharmacological relevance Origanum majorana L. (Lamiaceae) was usually used in Moroccan folk medicine to treat infantile colic and abdominal discomfort. Materials and methods The essential oil from the aerial part of the dry Origanum majorana L. (EOOM) was obtained through hydro distillation and analyzed by gas chromatography/mass spectrometry (GC/MS). The effect of EOOM on muscle relaxation was measured on rabbit and rat intestinal smooth muscle mounted in an isotonic transducer. Results 1) The main compounds obtained from the aqueous extract of this plant were alpha Terpineol, L-terpinen-4-ol and Beta.-Linalool. 2) EOOM inhibited in a concentration-dependent manner spontaneous contraction of rabbit jejunum, with an IC 50 = 64.08 ± 2.42 μg/mL. 3) In rat intestine, EOOM induced the relaxation of the tissue in concentration-dependent manner with an IC 50 = 39.70 ± 2.29 μg/mL when the tissue was pre-contracted with CCh 10-6 M, and 48.70 ± 2.26 μg/mL when the tissue was pre-contracted with 25 mM KCl. 4) The relaxation effect induced by EOOM was more important than that obtained in the presence of atropine, hexamethonium, Nifedipine, L-NAME and Blue of methylene. Conclusion the present result indicates that essential oil of Origanum majorana L. exhibit an effect on intestinal relaxation in vitro. This effect further validates the traditional use of Origanum majorana L. to treat infantile colic and abdominal discomfort. Graphical abstract fx1 [ABSTRACT FROM AUTHOR]

Study Question: Is JUNO protein present at the surface membrane of human oocytes and involved in the fertilisation process?Summary Answer: JUNO protein is expressed on the plasma membrane of human oocytes and its inhibition by a monoclonal antibody completely blocks gamete fusion.What Is Known Already: Fusion of gamete membranes is the culminating event of the fertilisation process, but its molecular mechanisms are poorly understood. Until now, three molecules have been shown to be essential: CD9 tetraspanin in the oocyte, Izumo1 protein on the sperm and Juno, its corresponding receptor on the oocyte. Oocyte CD9 and sperm IZUMO1 have been identified in human gametes and their interaction is also well-conserved among several mammalian species. The presence of JUNO on human oocytes, however, has not yet been reported, nor has its role in fertilisation been investigated.Study Design, Size, Duration: We selected an anti-human JUNO antibody in order to investigate the presence of JUNO on the oocyte membrane surface and studied its potential involvement in gamete membrane interaction during fertilisation.Participants/materials, Setting, Methods: Monoclonal antibodies against human JUNO (anti-hJUNO mAb) were produced by immunisation of mice with HEK cells transfected with the putative human JUNO sequence (HEK-hJUNO). These antibodies were used for immunostaining experiments and in vitro fertilisation assays with human gametes (GERMETHEQUE Biobank).Main Results and the Role Of Chance: Three hybridoma supernatants, verified by immunostaining, revealed specifically HEK-hJUNO cells. The three purified monoclonal antibodies, FJ2E4 (IgG1), FJ8E8 (IgG1) and FJ4F5 (IgG2a), recognised the soluble recombinant human JUNO protein and, in a western blot of HEK-hJUNO extracts, a protein with an expected MW of 25 kDa. In addition, soluble recombinant human IZUMO protein inhibited the binding of anti-hJUNO mAbs to cells expressing hJUNO. Using these anti-hJUNO mAbs in immunostaining, we identified the presence of JUNO protein at the plasma membrane of human oocytes. Furthermore, we revealed a progressive expression of JUNO according to oocyte maturity. Finally, we showed that human zona-free oocytes, inseminated in the presence of anti-hJUNO mAb, were not fertilised by human sperm. These results suggest that, as seen in the mouse, JUNO is indeed involved in human gamete membrane fusion during fertilisation.Large-scale Data: N/A.Limitations, Reasons For Caution: In accordance with French bioethics laws, functional tests were performed using zona-free oocytes, which of course does not fully encompass all normal in vivo physiological conditions. However, these in vitro tests do provide direct information regarding sperm-oocyte membrane interactions.Wider Implications Of the Findings: Mechanisms of gamete fusion appear to be homologous between mice and humans. However, some differences do exist and analysing the human mechanisms is essential. In fact, this is the first report describing the presence of JUNO on human oocytes and its involvement in human fertilisation. This discovery allows further examination of the understanding of molecular mechanisms that drive gamete fusion: a crucial challenge at a time when infertility affects 16% of reproductively active couples.Study Funding/competing Interests: This work was supported by the Agence Nationale pour la Recherche, Grant no. ANR-13-BVS5-0004, and by Association Institut du Cancer et d'Immunogénétique (ICIG). There are no competing interests. [ABSTRACT FROM AUTHOR]

Many medicinal plants around the world are used for therapeutic purposes against several diseases, including diabetes mellitus. Due to their composition of natural substances that are effective and do not represent side effects for users, unlike synthetic drugs, in this study, we investigated the inhibitory effect of Caralluma europaea (CE) on α-glucosidase activity in vitro; then the kinetics of the enzyme were studied with increasing concentrations of sucrose in order to determine the inhibition type of the enzyme. In addition, this effect of Caralluma europaea (CE) was confirmed in vivo using rats as an experimental animal model. Among the five fractions of CE, only the ethyl acetate fraction of C. europaea (EACe) induced a significant inhibition of α-glucosidase and its inhibition mode was competitive. The in vivo studies were conducted on mice and rats using glucose and sucrose as a substrate, respectively, to determine the oral glucose tolerance test (OGTT). The results obtained showed that the EACe and the aqueous extract of C. europaea (AECe) have significantly reduced the postprandial hyperglycemia after sucrose and glucose loading in normal and diabetic rats. AECe, also, significantly decreased intestinal glucose absorption, in situ. The results obtained showed that Caralluma europaea has a significant antihyperglycemic activity, which could be due to the inhibition of α-glucosidase activity and enteric absorption of glucose. [ABSTRACT FROM AUTHOR]

Platelets have an important role in thrombosis and haemostasis. Hyperactivity of the platelets has been associated with thromboembolic diseases and represents the main cause of complications of cardiovascular diseases. Crude aqueous extract (CAE) of Juglans regia root bark was evaluated for bleeding time, antiaggregant activity by using agonists, thrombin, ADP, collagen, or arachidonic acid (in vitro and ex vivo), and anticoagulant activity by measuring the clotting parameters: activated partial thromboplastin time, prothrombin time, thrombin time, and fibrinogen dosage (in vitro and ex vivo). The result of this study reported that the strongest antiaggregant effect of CAE in vitro was observed on the ADP-induced aggregation with inhibitions up to 90 %, while, in ex vivo experiments, the inhibition (more than 80 %) was observed with all agonists. Anticoagulant effect of CAE significantly prolonged the TT and decreased the fibrinogen level in vitro and ex vivo without interfering with APTT and PT. The bleeding time in mice and rats was significantly increased by CAE. The antiplatelet and anticoagulant effect observed in this study suggest that Juglans regia could have antithrombotic and/or thrombolytic activities and provide an alternative therapy against thrombotic complications related to cardiovascular diseases. [ABSTRACT FROM AUTHOR]

Background: Inula viscosa L. (Asteraceae) is a medicinal plant widely used as a folk medicine in oriental Morocco, to treat hypertension. The antihypertensive effect of the methanolic extract obtained from I. viscosa leaves was evaluated in hypertensive L-NAME rats. Systolic blood pressure (SBP) was measured using a non-invasive indirect tail-cuff plethysmographic method. Four groups of rats were used: a control group; a hypertensive group treated with L-NAME (32 mg/kg/day); a positive control group treated with L-NAME plus enalapril (15 mg/kg/day) as a reference antihypertensive agent; and a group treated with L-NAME plus MeOH-extract (40 mg/kg/day). Methods: Treatment with L-NAME alone caused a progressive increase in SBP. After 4 weeks, the value of SBP reached 160 ± 2 mmHg which shows the installation of hypertension. Enalapril prevented the increase in SBP, which remained normal at 123 ± 1 mmHg after 4 weeks of treatment. The administration of MeOH-extract along with L-NAME prevented the increase in SBP as well, which remained constant at 115 ± 1 mmHg after 4 weeks of treatment. In ex-vivo models, MeOH-extract produced a relaxation of pre-contracted ring aorta (54 ± 2% of relaxation at 3 g/L). But, when the rings were denuded, MeOH-extract failed to relax pre-contracted rings of aorta. Phytochemical study of I. viscosa MeOH-extract revealed the presence of phenolic compounds, such as cynarin and chlorogenic acid. Results: The present results suggest that I. viscosa MeOH-extract has an antihypertensive, predominantly mediated by an endothelium-dependent vasodilatory effect. Cynarin and chlorogenic acid, which have a strong vasorelaxant effect may be involved in the antihypertensive effect of the plant extract. The bioinformatic POM analysis confirms the therapeutic potential of cynarin and chlorogenic acids as inhibitors of various biotargets. Based on the results, the coordination of these phytochemicals with calcium and transition metals should be studied, for better scope at antihypertensive drug development. [ABSTRACT FROM AUTHOR]

Ethnopharmacological relevance Artemisia campestris L. (Asteraceae) has many traditional uses, among which treatment of diabetes and hypertension. Aim of the study This study was conducted in order to confirm the antihypertensive and hypotensive effects of A. campestris L. aqueous extract (AcAE) and to explore the underlying mechanism of action of its vasorelaxant effect, besides the acute toxicity. Also, the chemical composition of AcAE was investigated. Material and methods the chemical content of AcAE was determined by using HPLC and NMR techniques. The antihypertensive effect was assessed indirectly by tail-cuff method on L-NAME induced hypertensive rats, while the hypotensive action was monitored intravenously by invasive method on normotensive rats. The vasorelaxant effect and vascular mechanism of action were studied in the presence of antagonists and blockers on aorta isolated from normotensive rats. On the other side, the acute toxicity was studied by oral feeding of extract to the mice. Results The global phytochemical profile of AcAE reveals the presence of several polyphenols as main components. A. campestris L. infusion was characterized by mono- and di-cinnamoyl compounds, with 3,5-dicaffeoylquinic (isochlorogenic A) acid being the main compound, followed by 5-caffeoylquinic (chlorogenic) acid. Vicenin-2 (apigenin 6,8-di-C-glucoside) appeared to be the most abundant compound among flavonoids. The daily treatment with AcAE at 150 mg/kg/day prevented the installation of hypertension on L-NAME hypertensive rats, and reduced SBP from 172 mmHg up to 144 mmHg. At the dose 40 mg/kg, AcAE provoked reduction of systolic (SBP), diastolic (DBP) and mean arterial pressure (MAP), without affecting the heart rate. Also, AcAE (10 −2 −2 mg/ml) relaxed the precontracted aorta by 95.8±1.3%. The denudation and preincubation of aorta with atropine, calmidazolium, L-NAME, hydroxycobalamin, ODQ, 8-RP-Br-PET-cGMP, thapsigargin and verapamil attenuated the vasorelaxant response, while the pre-treatment with 4-AP, TEA, glibenclamide and BaCl 2 did not alter this effect. The oral administration of AcAE (0–6 g/kg) reveals no mortality or toxicity. Conclusions our study proved that AcAE possess an important antihypertensive, hypotensive and vasorelaxant effect, which is mediated via calmodulin-NO-cGC-PKG pathway, and via inhibition of calcium influx through voltage-operated calcium channels and activation of intracellular calcium mobilization into sarcoplasmic reticulum. Therefore, our findings give first evidence about the traditional use of A. campestris L. as antihypertensive plant. [ABSTRACT FROM AUTHOR]

Recently, Juno, the oocyte receptor for Izumo1, a male immunoglobulin, was discovered. Juno is an essential glycosylphosphatidylinositol (GIP)-anchored protein. This result did not exclude the participation of other GIP-anchored proteins in this process. After bibliographic and database searches we selected five GIP-anchored proteins (Cpm, Ephrin-A4, Gas1, Gfra1 and Rgmb) as potential oocyte candidates participating in fertilisation. Western blot and immunofluorescence analyses showed that only three were present on the mouse ovulated oocyte membrane and, of these, only two were clearly involved in the fertilisation process, namely growth arrest specific 1 (Gas1) and glial cell line-derived neurotrophic factor receptor α1 (Gfrα1). This was demonstrated by evaluating oocyte fertilisability after treatment of oocytes with antibodies against the selected proteins, with their respective short interference RNA or both. Gfrα1 and Gas1 seem to be neither redundant nor synergistic. In conclusion, oocyte Gas1 and Gfrα1 are both clearly involved in fertilisation. [ABSTRACT FROM AUTHOR]

At present, the clinically most advanced strategy to treat Duchenne muscular dystrophy (DMD) is the exon-skipping strategy. Whereas antisense oligonucleotide-based clinical trials are underway for DMD, it is essential to determine the dystrophin restoration threshold needed to ensure improvement of muscle physiology at the molecular level. A preclinical trial has been conducted in golden retriever muscular dystrophy (GRMD) dogs treated in a forelimb by locoregional delivery of rAAV8-U7snRNA to promote exon skipping on the canine dystrophin messenger. Here, we exploited rAAV8-U7snRNA-transduced GRMD muscle samples, well characterized for their percentage of dystrophin-positive fibers, with the aim of defining the threshold of dystrophin rescue necessary for normalization of the status of neuronal nitric oxide synthase mu (nNOSµ), inducible nitric oxide synthase (iNOS), and ryanodine receptor-calcium release channel type 1 (RyR1), crucial actors for efficient contractile function. Results showed that restoration of dystrophin in 40% of muscle fibers is needed to decrease abnormal cytosolic nNOSµ expression and to reduce overexpression of iNOS, these two parameters leading to a reduction in the NO level in the muscle fibers. Furthermore, the same percentage of dystrophin-positive fibers of 40% was associated with the normalization of RyR1 nitrosylation status and with stabilization of the RyR1-calstabin1 complex that is required to facilitate coupled gating. We concluded that a minimal threshold of 40% of dystrophin-positive fibers is necessary for the reinstatement of central proteins needed for proper muscle contractile function, and thus identified a rate of dystrophin expression significantly improving, at the molecular level, the dystrophic muscle physiology. [ABSTRACT FROM AUTHOR]

Little is known about the molecular mechanisms that induce gamete fusion during mammalian fertilization. After initial contact, adhesion between gametes only leads to fusion in the presence of three membrane proteins that are necessary, but insufficient, for fusion: Izumo1 on sperm, its receptor Juno on egg and Cd9 on egg. What happens during this adhesion phase is a crucial issue. Here, we demonstrate that the intercellular adhesion that Izumo1 creates with Juno is conserved in mouse and human eggs. We show that, along with Izumo1, egg Cd9 concomitantly accumulates in the adhesion area. Without egg Cd9, the recruitment kinetics of Izumo1 are accelerated. Our results suggest that this process is conserved across species, as the adhesion partners, Izumo1 and its receptor, are interchangeable between mouse and human. Our findings suggest that Cd9 is a partner of Juno, and these discoveries allow us to propose a new model of the molecular mechanisms leading to gamete fusion, in which the adhesion-induced membrane organization assembles all key players of the fusion machinery. [ABSTRACT FROM AUTHOR]