The aim of cytochemical techniques is to localize specific biochemical components in particular tissue and cell compartments. However, since preparation tissues for structural observation results major alterations the properties their components, a problem retain an adequate degree as well preservation. In present study, we describe obtained using various colloidal gold on processed through different approaches. We found that any manipulation during its processing can result modifications...

Abstract The nature of cholesterol‐binding proteins acting upon human spermatozoa during in vitro capacitation was determined by measuring the efflux [ 3 H]cholesterol and H]cholesteryl sulfate from labeled spermatozoa. Efflux H]sterols stimulated when gametes were incubated Ham's F‐10 medium supplemented with female serum or follicular fluid. Upon centrifugation capacitated application supernatant to density‐gradient ultracentrifugation for lipoprotein analysis, both found be carried...

Abstract Exosomes and microvesicles are extracellular vesicles released from cells can contain lipids, miRNAs proteins that affect at distant sites. Recently, containing miRNA have been implicated in uterine microenvironment of pigs, a species with unique epitheliochorial (non-invasive) placentation. Here we report novel role conceptus-derived exosomes/microvesicles (hereafter referred to as vesicles; EVs) embryo-endometrial cross-talk. We also demonstrate the stimulatory effects EVs...

We introduce here a technique, "label-fracture," that allows the observation of distribution cytochemical label on cell surface. Cell surfaces labeled with an electron-dense marker (colloidal gold) are freeze-fractured and fracture faces replicated by plantinum/carbon evaporation. The exoplasmic halves membrane, apparently stabilized deposition Pt/C replica, washed in distilled water. new method reveals surface coincident replica face. Initial applications indicate high resolution (less than...

Fracture-label, surface-replica, and routine freeze-fracture techniques were used in combination with phospholipase A2-colloidal gold (PLA2-CG) filipin as probes to study changes the distribution of phospholipids cholesterol, respectively, morphologically defined plasma membrane domains mouse spermatozoa during vitro capacitation. In noncapacitated spermatozoa, quantitative analysis revealed that fractured overlying equatorial segment carried highest PLA2-CG labeling density. The next...

The zona pellucida (ZP), a glycoprotein layer that encloses the mammalian oocyte, is formed during follicular development in ovary, persists at time of fertilization within oviduct, and then surrounds embryo until implantation uterus. Although structure chemical properties ZP have been extensively studied, precise site origin remains matter controversy. Moreover, mechanism synthesis secretion constituents not fully elucidated. We recently developed monoclonal antibodies (MAbs) against...

We used fracture-label to establish ultrastructural localization of glycoproteins in cross-fractured nuclei duodenal columnar and exocrine pancreatic cells. Mannose residues were detected cell by labeling freeze-fractured tissues with concanavalin A-horseradish peroxidase X colloidal gold (Con A-HRP CG) or direct A CG); fucose Ulex Europaeus I (UEA markers. Areas the three main intranuclear compartments (euchromatin, heterochromatin, nucleolus) exposed freeze-fracture determined automated...

Studies carried out in several mammalian species suggest that oviductin, also known as oviduct-specific glycoprotein or OVGP1, plays a key role sperm capacitation, fertilization, and development of early embryos. In the present study, we used recombinant DNA technology to produce, for first time, hamster OVGP1 (rHamOVGP1) human embryonic kidney 293 (HEK293) cells. rHamOVGP1 secreted culture medium was purified by affinity chromatography. The resulting protein migrated poly-dispersed band...

The tight junction has been characterized as a domain of focal fusions the exoplasmic leaflets lipid bilayers from adjacent epithelial cells. Approximating membranes to within fusion distance is thermodynamically unfavorable process and requires participation membrane-bridging or -fusion proteins. No known protein exhibits such activities. Annexin A2 (A2), in particular its heterotetramer (A2t), form junctions between bilayer structures through molecular bridging their external leaflets. We...

Abstract We have previously localized an antigen of oviductal origin to the zona pellucida superovulated hamster oocytes (Kan et al.: Journal Histochemistry and Cytochemistry 36:1441—1447, 1988) described intracellular distribution this in epithelium Biology Reproduction 40:585—598, 1989). These results led our hypothesis that oviduct is a bona fide site certain components pellucida. In report, using high resolution lectin‐gold approach with Helix pomatia lectin (HPL)‐colloidal gold complex,...

We have previously localized an antigen of oviductal origin in the zona pellucida superovulated hamster ova. This is a high-molecular-weight glycoprotein that secreted by nonciliated secretory cells oviduct and later transferred to oocyte during transit. rich N-acetyl-D-galactosamine residues has been designated Hamster Oviductin-1. In present study, we examined intracellular localization oviductin blastomeres developing embryos golden hamster. Seventeen cycling females were used for...

The zona pellucida is an extracellular matrix of glycoproteins which surrounds the mammalian oocyte and preimplantation embryo. We have recently developed monoclonal antibodies against oviductal golden hamster. applied post-embedding immunocytochemical method using a antibody (IgGl,k) to determine precise location antigenic sites in cumulus oophorus complex superovulated By applying high-resolution protein A-gold technique, we demonstrated that immunoreactivity were exclusively encompassing...

Cysteinyl leukotrienes (CysLTs) are potent mediators of inflammation synthesized by the concerted actions 5-lipoxygenase (5-LO), 5-LO-activating protein (FLAP), leukotriene C(4) synthase, and additional downstream enzymes, starting with arachidonic acid substrate. CysLTs produced macrophages, eosinophils, mast cells, other inflammatory cells activate 3 different high-affinity CysLT receptors: CysLT(1)R, CysLT(2)R, GPR 17. We sought to investigate vascular sites CysLT(2)R expression role...

Implantation failure is a major hurdle to successful pregnancy. The high rate of postimplantation fetal loss in nonobese diabetic (NOD) mice believed be related an abnormal decidual production interferon (IFN)gamma. To address whether diabetes alters the natural events associated with implantation, certain morphological and molecular features uterine receptivity NOD (dNOD) were examined normally mated pregnancy concanavalin A (ConA)-induced pseudopregnancy. As opposed normoglycemic (cNOD)...

Oviductin or OVGP1, also known as oviduct-specific glycoprotein, has been shown to enhance sperm capacitation in addition its other beneficial effects on fertilization and early embryo development. We hypothesized that estrus stage-specific hamster oviductin (eHamOVGP1) can potentiate the enhancement of tyrosine phosphorylation proteins during capacitation. Immunofluorescent staining confocal microscopy well immunocytochemistry surface replica technique localized tyrosine-phosphorylated...

The mammalian oviduct synthesizes and secretes a major glycoprotein known as oviductin (OVGP1), which has been shown to interact with gametes early embryos. Here we report the use of recombinant DNA technology produce, for first time, secretory form human OVGP1 in HEK293 cells. colonies stably expressing (rHuOVGP1) were established by transfecting cells an expression vector pCMV6-Entry constructed cDNA. Large quantities rHuOVGP1 obtained from transfected using CELLSPIN cell cultivation...

Abstract The synthesis and secretion of proteins in the different regions human epididymis were studied vitro. Epididymal tissues obtained from patients undergoing castration for prostatic carcinoma or cadavers incubated presence [ 35 S]methionine, resulting radiolabeled analysed on SDS‐PAGE. corpus region was found to be most active segment total protein synthesis. Significant qualitative quantitative changes observed pattern secreted epididymal regions. To establish those that interact...

We have previously reported that the calcineurin inhibitor macrolide immunosuppressant Tacrolimus (TAC, FK506) can promote migration and invasion of human-derived extravillous trophoblast cells conducive to preventing implantation failure in immune-complicated gestations manifesting recurrent failure. Although exact mode action TAC promoting has yet be elucidated, integral association its binding protein FKBP12 with inositol triphosphate receptor (IP3R) regulated intracellular calcium [Ca

The zona pellucida (ZP) is an extracellular coat synthesized and secreted by the oocyte during follicular development surrounding plasma membrane of mammalian eggs. To date, mechanism synthesis secretion, mode assembly, intracellular trafficking ZP glycoproteins have not been fully elucidated. Using antibodies against mouse ZP1, ZP2, ZP3 in conjunction with protein A-gold technique, we shown association immunolabeling Golgi apparatus, secretory granules, a complex structure called vesicular...

Smooth endoplasmic reticulum assembly was studied in a cell-free system using thin-section and freeze-fracture electron microscopy. Incubation of rat hepatocyte rough smooth microsomes the presence ATP, GTP, cytosol (Xenopus egg) an ATP-regenerating led to membrane networks comprising central core interconnecting tubules continuous with peripherally located cisternae. Glucose-6-phosphatase cytochemistry confirmed origin reconstituted membranes. When both ATP GTP were omitted from incubation...