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Query: UNIPROT:P30536 (PBS)
 9,886 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

As an outgrowth of in-vitro fertilization and embryo transfer, detection of genetic and metabolic defects prior to implantation might be possible in the future. The objective for preimplantation diagnosis would be to sample a minimum of cell material of the conceptus for diagnosis prior to transfer. Different protocols for isolating individual blastomeres out of 2-cell mouse embryos were evaluated. 2-cell mouse embryos (from F1 hybrids C57B1 females x CBA males) were collected and the zona pellucida was removed by enzyme treatment (pronase), by exposure to Ca2+-Mg2+-free acid Tyrode (pH = 2.5) or by mechanical force. Individual blastomeres were obtained by exposure to an enzyme (pronase), to a chelating agent (EDTA-glycine mixture), to Ca2+-Mg2+-free PBS or after isolation by mechanical force. The biopsied blastomeres were then cultured in vitro as such or first replaced into a host zona pellucida. Evaluation was performed by culture in vitro up to the blastocyst stage and by transfer of embryos appearing morphologically normal into pseudopregnant foster mothers. A chromosomal study of the second mitotic division of the isolated blastomere was also performed. All isolation procedures had a negative impact on the in-vitro and in-vivo growth patterns of the isolated blastomeres. After culture in vitro to the blastocyst stage, different abnormalities could be observed: embryos lacking compaction, embryos with double blastocoelic cavities and embryos with no inner cell mass (trophoblastic vesicle). After replacement of the isolated blastomeres into a host zona pellucida, similar observations could be made. Chromosomal analysis did not reveal a clear influence of the different biopsy methods on the mitosis of the isolated blastomeres.
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PMID:Evaluation of different biopsy methods of blastomeres from 2-cell mouse embryos. 320 55

Eight day (8-d CEF) and 16 day old chick embryo fibroblasts (16-d CEF) obtained after a mild trypsin treatment (50 micrograms/ml in Ca2+ and Mg2+-free PBS, plus 10 mM EDTA) for 10 min at 37 degrees C present the same number of fibronectin (FN) binding sites at their surface (approximately 550,000 sites per cell) with a Kd approximately equal to 1.40 microM in both cases. Furthermore, FN interacted with high molecular weight plasma membrane proteins (150,000 and 125,000) insensitive to trypsin treatment. Both 8-d and 16-d CEF adhered and spread to the same extent on a fibronectin coated substratum (80% of the CEF adhered in 60 min). In contrast, 8-d and 16-d CEF behaved differently towards laminin (LM). 8-d CEF exhibited approximately 5500 binding sites per cell with a Kd of 1.5 nM (Codogno P., Doyennette, M.-A. and Aubery M., 1987, Experimental Cell Research, 169, 478-489.) and were highly sensitive to trypsin treatment, whereas 16-d CEF do not express cell surface binding sites for laminin. Differences were also observed in the adhesive capacities of 8-d and 16-d CEF on LM substrata: 8-d CEF adhered and spread on LM in a very specific manner (60% of the cells adhere in 60 min) and 16-d CEF did not adhere to LM even after long periods of incubation exceeding 360 min.
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PMID:Alterations of fibroblast interactions with fibronectin and laminin during chick embryo development. 344 Feb 93

As an extension of in-vitro fertilization and embryo transfer, detection of genetic and metabolic defects prior to implantation might be possible in the future. The objective of pre-implantation diagnosis would be to sample a minimal amount of cellular material of the conceptus for diagnosis prior to transfer. Different protocols for isolating individual blastomeres from two-cell mouse embryos were evaluated. Two-cell mouse embryos were collected and the zona pellucida was removed by enzyme treatment (pronase), by exposure to acid Tyrode (pH = 2.5) or by mechanical force (suction into a small pipette, removal with a microblade). Individual blastomeres were obtained by exposure to a chelating agent (EDTA-glycine mixture), to Ca2+-Mg2+-free PBS or after isolation by mechanical force (bisection with a microblade or suction in a small pipette). The isolated blastomeres were then cultured in vitro without zonae pellucidae. All isolation procedures had a negative impact on the growth patterns of the isolated blastomeres. Different abnormalities could be observed at the blastocyst stage including embryos lacking visible compaction features, embryos with double blastocoelic cavities and embryos with no inner cell mass (trophoblastic vesicles).
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PMID:Assessment of different isolation procedures for blastomeres from two-cell mouse embryos. 365 31

Hypercalcemia is frequently observed in patients with multiple myeloma and renal failure. Whether Bence Jones protein (BJP) is directly nephrotoxic and how and whether hypercalcemia might contribute to this putative nephrotoxicity is currently unclear. To examine this issue, we studied the effect of modest hypercalcemia on the glomerular filtration rate (GFR) of rats exposed to a BJP that by itself had been found to be nonnephrotoxic. Three groups of rats were studied. All were anesthetized and underwent a baseline measurement of inulin clearance (Cin). After this, group 1 (n = 13) rats were given 2 ml of vehicle (phosphate-buffered saline solution [PBS]) and were then made hypercalcemic with an infusion containing 0.048 mol/L CaCl2. At the end of 2 hours a second Cin was measured. Group 2 rats (n = 8) were given 100 mg BJP in 2 ml PBS and a non-calcium-containing infusate. Group 3 (n = 11) rats were given 100 mg of the BJP in 2 ml PBS and then the calcium-containing infusate used in group 1 rats. Rats in groups 2 and 3 also had a second Cin measured at the end of 2 hours. Renal blood flow was measured with an electromagnetic flow probe. At the completion of the second clearance, kidneys were processed for renal histologic assessment. The serum calcium level measured during the second Cin period was 13.5 mg/dl for group 1, 7.9 mg/dl for group 2, and 13.7 mg/dl for group 3. No significant decrement in GFR was observed in group 1 or 2 rats. In contrast, group 3 rats had a 46% fall in GFR.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Hypercalcemia can potentiate the nephrotoxicity of Bence Jones proteins. 365 25

A high-voltage generating machine which could generate semi-rectangular pulses in PBS solution was constructed, and the effects of field strength and duration of the pulse on electric pulse-mediated transformation of mouse mammary carcinoma FM3A cells by a linear form of plasmid pSV2neo DNAs were examined. In parallel, cell survival and growth after pulsing were analyzed. When the field strength and duration of the pulse were increased, the transformation frequency increased, although the cell survival rate decreased. Under the best conditions, the transformation frequency was 2 X 10(-4), which was 80 times higher than that obtained by the calcium phosphate coprecipitation method.
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PMID:Electric pulse-mediated gene transfer in mammalian cells grown in suspension culture. 373 Dec 84

The steady-state transmembrane potentials of P815 mastocytoma cells were recorded when the cells were bathed in salines of different compositions. In the normal growth medium (RPMI 1640 with added fetal calf serum) the mean membrane potential was -8.7 mV (SEM +/- 0.4, n = 22). A family of Tris-buffered salines (TBS), modeled from Dulbecco's modified PBS (289 mosmol, 169 milliionic strength units, pH 7.5), having different K+ and different C1- concentrations, were designed and used to bathe the tumor cells. All of the TBS solutions had constant, but reduced levels of ionized Ca2+. In the absence of external C1-, an increase of external K+ from 2 to 20 mM results in a 5.7 mV depolarization. In the presence of external C1- the same increase in external K+ results in a 2.1 mV depolarization. The presence of 145 mM C1- resulted in a steady-state depolarization (for either level of K) of about 50%. One explanation for these results would be the presence of an inward-going active C1- transport.
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PMID:Effects of potassium and chloride on the membrane potentials of P815 mastocytoma tumor cells. 681 35

Osteoclasts were isolated mechanically from the medullary bone of laying hens kept 7 days on a low calcium diet. Osteoclast enrichment was achieved with 3-4 sedimentations of the cell suspension in test-tubes prepared by layering on the bottom with BSA 10% in MEM-HEPES or PBS, above which the cells were suspended in MEM-HEPES or PBS. The final suspension of osteoclasts was cultivated in MEM with 10% FCS for 3 weeks. The cultures were observed by phase-contrast and scanning electron microscopy (SEM). By the third day, the osteoclasts were completely spread onto the plastic dishes and a variety of morphologies and of intercellular contacts was established. Osteoclasts in culture do not lose their morphology; they survive for long periods and can be used in many experimental systems.
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PMID:Isolated osteoclasts in primary culture: first observations on structure and survival in culture media. 715 21

In order to clarify the presence and localization of crystal matrix protein (CMP) upon calcium oxalate crystals, scanning electron microscopy (SEM) and backscattered electron imaging (BEI) techniques were used. This protein exhibits a remarkable affinity with calcium oxalate crystals and may be important in stone pathogenesis. In this paper, rabbit anti-human CMP polyclonal antibody was used as first antibody, and for the second antibody, goat anti-rabbit IgG conjugated with 20 nm immunogold was used. Freshly prepared crystals from male urine were fixed in SEM fixative, then blocked and washed with phosphate-buffered saline and bovine serum albumin (PBS/BSA). First and second antibodies were reacted in PBS/BSA. Crystals were then dehydrated and finally coated for SEM study. The SEM technique showed bipyramidal shaped dihydrate calcium oxalate crystals in every sample and even at high magnification, colloidal gold could barely be seen. BEI clearly demonstrated the presence and localization of the gold on the surface of the crystals as well as on the macromolecules eluted from the crystals by dissolving them in ethylenediamminetetraacetic acid solution.
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PMID:Backscattered electron imaging of crystal matrix protein on the surface of calcium oxalate crystals using colloidal gold. 755 94

Dietary factors have been shown to affect excretion of fecapentaenes, potent mutagens present in human feces. Apart from effects of the diet on the bacterial synthesis of fecapentaenes in the bowel, fecapentaene excretion is likely to be indirectly influenced by the composition of the bowel contents, in particular fecapentaene-binding or -solubilizing factors. In the present study, interactions between dietary fiber and fecapentaene-12 (FP-12), as well as the effects of bile acids and calcium on the solubility of FP-12 in aqueous solutions, have been investigated in vitro. The results demonstrated that FP-12 may strongly adsorb to fiber, as indicated by reduced concentrations in the aqueous PBS phase when increasing amounts of fiber are added. This fecapentaene-binding capacity of fiber may explain the positive correlations that have previously been found between excreted fecapentaene concentrations and fiber consumption in human population studies. Further, it was found that at concentrations physiologically occurring in feces, both cholic and deoxycholic acid as well as mixtures of bile acids may increase the aqueous solubility of FP-12. This solubilizing effect of bile acids can be reduced by adding calcium at physiological concentrations of 2.5 mg/ml. It is hypothesized that high dietary fiber intake may increase fecapentaene excretion as a result of this fecapentaene fiber adsorption, which in turn may result in diminished exposure of the human bowel epithelium to these putative initiators of colorectal cancer. In contrast, high concentrations of fecal bile acids may act as fecapentaene-solubilizing factors which increase fecapentaene bioavailability, thereby possibly resulting in increased risk for colorectal cancer.
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PMID:In vitro study on the effects of fecal composition on fecapentaene kinetics in the large bowel. 768 4

Immunostaining of human neutrophils incubated with endothelin-1 (ET-1) showed intense and spreading pattern of anti human granulocyte elastase within the cytosol. That reflected neutrophil activation followed by the release of granule contents by ET-1. In contrast, PBS (phosphate buffered saline) treated neutrophils showed localized and faintly stained granules. Intracellular calcium in fura-2 loaded neutrophils was measured at 340/380 nm. A dose and time dependent increase in intracellular calcium by ET-1 occurred in human single neutrophil. Elastase activity assay was done with chromogenic substrate S2484. ET-1 induces dose and time dependent increase in elastase activity in neutrophil suspensions like ionophore A23187. A similar time dependent increase in elastase activity was retained even after repeated wash and ET-1 treatment. That confirmed the viability of most of the neutrophils after each treatment. In umbilical cord preparations, ET-1 treated neutrophils could migrate from the venous lumen into the tissue matrix of the umbilical cords. Hematoxylin and eosin staining revealed a massive tissue destruction in ET-1 activated neutrophil treated cords when compared to sham control and untreated neutrophil injected cords. Immunostaining with monoclonal anti human elastase revealed an intense staining in former sections when compared to the others. We suggest that ET-1 activated neutrophil might play a major role in endothelial injury and tissue damage in conditions with high blood level of endothelin.
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PMID:Activated neutrophil by endothelin-1 caused tissue damage in human umbilical cord. 774 May 23


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