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PubMed2004

Investigations of the methanolic leaf extract of Costus afer. Ker for pharmacological activities in vitro and in vivo.

The methanolic leaf extract of Costus afer. Ker (family: Zingiberaceae) was investigated for some pharmacological effects in vivo and in vitro. Brine shrimp lethality test showed that the extract was significantly (p < 0.05) cytotoxic with LC50 of 21.3 ppm. The extract showed moderate local anesthetic property, about twice less than lignocaine of the same concentration, on guinea pig wheal test. The extract contracted the guinea pig ileum in a concentration-dependent manner, but had no effect on pleuripara and nullipara non-gravid uteri at progestogenic and estrogenic phases respectively. The contractile effect on the guinea pig ileum was partially inhibited by atropine but completely reversed by adrenaline. The extract induced expulsion of whole fetuses still enveloped within the placental membrane at the 3rd trimester of pregnancy. The extract exhibited a biphasic antihyperglycemic activity. At 200 mg/kg body wt., p.o., it decreased the blood glucose level by 50% in Streptozotocin-induced hyperglycemia in male rats in 60 minutes post dosing. However, doses above 200 mg/kg body wt., p.o., caused increase in blood glucose level, potentiating the action of STZ. At 10 microg/ml the extract induced about 98% glucose uptake in differentiated 3T3-L1 adipocytes when compared with insulin (340 nm).

Abortifacient Agents↗

PubMed1975

Immunological relationship between human placental and intestinal alkaline phosphatase.

Crystalline human alkaline phosphatase from placenta and intestine was isolated by butanol extraction, acetone precipitation, a heat step (for the placental enzyme), ammonium sulfate precipitation, anion exchange chromatography, gel filtration and crystallisation with ammonium sulfate. Rabbit antibodies showed a partial cross-reaction between both enzymes in double diffusion, quantitative precipitation experiments and in serial precipitin curves. There was no reaction with human alkaline phosphatases from liver, kidney and bone. Three phenotypes of placental alkaline phosphatase with different electrophoretical mobility exhibited identical immunological reactions with their respective antisera. Monospecific antisera were obtained by absorption with crystalline intestinal or placental alkaline phosphatase. These monospecific antisera against the placental or the intestinal alkaline phosphatase can be used for an immunological determination of these two alkaline phosphatases without contamination by other alkaline phosphatases in human serum.

Alkaline Phosphatase↗

PubMed1991

Phosphorus 31 magnetic resonance spectroscopy of human placenta and quantitation with perchloric acid extracts.

Phosphorus 31 magnetic resonance spectroscopic studies of fresh placental tissue are reported that indicate resonances for adenosine triphosphate, inorganic phosphate, sugar phosphates-phosphomonoesters, and phosphodiesters. Perchloric acid extract methods were used to further characterize and quantitate phosphorous metabolites in term human placentas by phosphorus 31 magnetic resonance spectroscopy. The perchloric acid extracts give enhanced resolution of phosphorus signals and allow identification of other phosphorus metabolites including small amounts of phosphocreatine. Emphasis was placed on quantitating adenosine triphosphate levels in the acid extracts with the use of the external reference standard hexachlorocyclotriphosphazene in a coaxial capillary system. Adenosine triphosphate levels measured in this way ranged from 0.404 to 0.709 mumol per gram wet weight. Comparison with an internal standard method with phosphocreatine is also reported. Contribution to the measured high-energy phosphate pool from blood in the highly vascularized tissue was found to be relatively large and could range from 30% to 50% of the total adenosine triphosphate measured.

2,3-Diphosphoglycerate↗

PubMed1986

[Studies on the immunohistochemical localization of coagulation fibrinolysis factors in the placenta, especially of placental plasminogen activator (PPA)].

We succeeded for the first time in extracting and purifying a coagulation fibrinolysis factor in the placenta, that is, a placental plasminogen activator (PPA). Using the enzyme-labeled antibody technique (indirect method), we investigated the localization of PPA, fibrinogen (FBG) and fibronectin (FN) in the placenta. And we tried to elucidate the physiological significance of the above three. The F(ab')2 fragments of their antibodies were produced by pepsin digestion, because there were various Fc receptors in the placenta. The results are summarized as follows: PPA was produced in the trophoblast, and was secreted out of the microvilli. FBG was located in the plasma. But, in the placenta, it was not in the cells and the connective tissue of which chorionic villi was composed. FN was located in the ground substances of connective tissue in the chorionic villi, and was associated with the cell structure. Because PPA, FBG and FN showed the same distribution on the fibrinoid material, it is expected that coagulation fibrinolysis activity proceeds in that fibrinoid material.

Female↗

PubMed1997

Inhibition by green tea extract of diethylnitrosamine-initiated but not choline-deficient, L-amino acid-defined diet-associated development of putative preneoplastic, glutathione S-transferase placental form-positive lesions in rat liver.

The effects of green tea extract (GTE) on exogenous and endogenous models of rat liver carcinogenesis using diethylnitrosamine (DEN) and a choline-deficient, L-amino acid-defined (CDAA) diet were studied. For the exogenous carcinogenesis study, male Fischer 344 rats, 6 weeks old, were given a single intraperitoneal dose of 200 mg/kg body weight of DEN, partially hepatectomized at week 3, and administered GTE at doses of 0, 0.01 and 0.1% in the drinking water from week 2 for 10 weeks. For the endogenous carcinogenesis study, rats were fed the CDAA diet and simultaneously given GTE for 12 weeks. All rats were killed at the end of week 12. After DEN-initiation, the apparent numbers of glutathione S-transferase placental form-positive foci, assayed as putative preneoplastic lesions, were decreased by the administration of GTE, though their sizes were not altered. In contrast, GTE did not significantly reduce the numbers of the lesions induced by the CDAA diet or affect their sizes. While the levels of 8-hydroxyguanine, a parameter of oxidative DNA damage, were reduced by the GTE administration in both experimental models, GTE did not protect against the CDAA-diet-associated liver tissue damage in terms of either histology or plasma marker enzyme levels. We conclude that, while GTE may be a possible chemopreventive agent for nitrosamine-initiated hepatocarcinogenesis in the absence of chronic hepatocyte damage, it does not significantly inhibit lesion development in hepatocarcinogenesis associated with the CDAA diet, a cirrhosis-associated model.

Alanine Transaminase↗

PubMed2003

Human placental amnion is a novel substrate for detecting autoantibodies in autoimmune bullous diseases by immunoblotting.

BACKGROUND: Identification of antigens by immunoblotting techniques, using epidermal and dermal extracts, is regarded as essential for making a definitive diagnosis in autoimmune bullous diseases (AIBDs). These procedures involve epidermal-dermal separation for subsequent protein extraction, which may result in partial loss of some antigenic polypeptides and changes in the conformational epitopes targeted by autoantibodies in AIBDs. It may therefore be necessary to use different substrates for consistent results. Objectives To evaluate the usefulness of human placental amnion extract as a substrate for immunoblotting in the diagnosis of AIBDs. METHODS: We checked the structural components of the desmosomes and basement membrane zone (BMZ) of amnion by electron microscopy. Using immunofluorescence and immunoblotting techniques, we tested the amnion immunoreactivity with antibodies to desmosomal and BMZ proteins, and with sera from 76 patients with AIBDs including pemphigus vulgaris, pemphigus foliaceus, bullous pemphigoid (BP), pemphigoid gestationis, linear IgA bullous dermatosis, epidermolysis bullosa acquisita, paraneoplastic pemphigus and mucous membrane pemphigoid. RESULTS: The desmosomes and BMZ of the amnion tissue were ultrastructurally similar to those in skin. Antigen mapping confirmed that amnion contains all the proteins that were recognized by a panel of monoclonal and polyclonal antibodies. Immunoblotting showed that the antibodies clearly detected bands corresponding to desmogleins 1 and 3, desmocollins 1 and 2, desmoplakins 1 and 2, three subunits (alpha3, beta3 and gamma2) of laminin 5, BP antigens 1 and 2, the 97-kDa LAD antigen and type VII collagen. In addition, most of the patient sera (82%) reacted exclusively with their respective antigens. CONCLUSIONS: Harvesting proteins from amnion does not require epidermal-dermal separation, and a sufficient yield of desmosomal and hemidesmosomal proteins can be obtained. Therefore, amnion may be a more reliable source of substrate than skin samples for immunoblot analysis of AIBDs.

Amnion↗

PubMed1986

Human fallopian tube contains placental protein 14.

Radioimmunoassay and immunoperoxidase staining were used to study the tissue content and localization of an endometrial protein, placental protein 14, in the human fallopian tube. Placental protein 14 immunoreactivity was found in saline extracts from all fallopian tubes tested (n = 14). In the fimbrial part the placental protein 14 content was higher in the secretory than in the proliferative phase (p less than 0.01). No difference was found in the placental protein 14 content between the isthmic, ampullar, and fimbrial parts of the tube. Immunoperoxidase staining localized placental protein 14 to the ciliated and secretory epithelial cells of the mucosa in all parts of the tube regardless of the phase of menstrual cycle. The occurrence of the same protein in the endometrium and fallopian tube is compatible with the common embryonic origin from the müllerian duct of these tissues.

Chromatography, Gel↗

PubMed1981

Growth and development of sympathetic neurons in tissue culture.

Adrenergic neurons from the superior cervical ganglion of the neonatal rat, when studied under certain culture conditions, develop cholinergic properties including hexamethonium-sensitive synaptic interactions, choline acetyltransferase activity and synaptic endings containing clear vesicles. Evidence from correlative biochemical, physiological and morphological studies on populations of neurons indicates that cholinergic function is acquired by the majority of neurons and not by subpopulation. The factors that influence the development of cholinergic function in culture include the presence of non-neuronal cells, the addition of human placental serum and chick embryo extract to the culture medium as well as the stage of development at which the neurons are placed in culture. Neurons from mature rats, maintained as explants in culture, develop low choline acetyltransferase activity and the synaptic endings containing dense-cored vesicles. In contrast, if dissociated, these adult neurons develop several cholinergic characteristics. Studies to determine which adrenergic properties are retained in neurons expressing cholinergic characteristics have shown an increase in the activities of tyrosine hydroxylase and dopamine beta-hydroxylase in both explanted and dissociated perinatal neurons. In addition, tyrosine hydroxylase has been localized immunocytochemically in neurons identified as cholinergic by electrophysiological methods.

Animals↗

PubMed1993

Hepatocellular carcinoma producing universal type of alkaline phosphatase.

We report on a 54-year-old man with hepatocellular carcinoma (HCC) associated with a marked elevation of serum alkaline phosphatase (ALP) levels. Serum ALP was biochemically similar to that of universal (liver/bone/kidney) type. The noncarcinomatous area revealed typical micronodular cirrhosis due to excessive alcohol consumption. By histochemical staining, ALP activity was demonstrated diffusely within the cytoplasm of carcinoma cells. Immunohistochemical observation of the carcinoma cells excluded the intestinal or placental type of ALP. Tissue extracts from the carcinomatous area had much higher ALP activities than those from a noncarcinomatous area, which also showed characteristics of the universal type. The present HCC is the first reported to produce and excrete the universal type of ALP.

Alkaline Phosphatase↗

PubMed1977

In vitro metabolism of prednisolone, dexamethasone, betamethasone, and cortisol by the human placenta.

Many of the factors influencing the pharmacologic action of corticosteroids on the maternal-placental-fetal unit remains obscure. To clarify the effect of the placenta on corticoid transfer, tritiated steroids were incubated in saline for 2 hours with minces of midgestational or term placental tissue. The media were extracted and chromatographed on Sephadex-LH-20. Large peaks corresponding to the inactive 11-keto metabolites were observed for cortisol (67 per cent) and prednisolone (51 per cent) while conversion of dexamethasone (1.8 per cent) and betamethasone (7.1 per cent) was low or negligible.

Betamethasone↗

PubMed1989

Trans-placental transport and metabolism of carbacyclin by perfused human placental in vitro.

When carbacyclin (5E-6a-carba-prostaglandin I2) was added to the maternal afferent circulation of in vitro perfused placentae from normal term pregnancies, relatively little carbacyclin was found in either the maternal or fetal efferent circulations. When carbacyclin was added to the perfusate at 1.0 microM, the peak level in the maternal effluent was only 0.06 microM and in the fetal effluent, 0.026 microM. When infused at 10 microM, 0.77 microM carbacyclin was measured in the maternal effluent and 0.13 in the fetal effluent. These findings demonstrate that carbacyclin is transferred across the placenta from the maternal side to the fetal, but that the net transfer is small. The assay procedure employed HPLC resolution, followed by capillary gas chromatography and selected ion monitoring using PGB as an internal standard. The low levels of carbacyclin detected in the effluents did not result from poor recovery in the analyses. When carbacyclin was added to maternal or fetal effluents at 1 microM, the recovery averaged 85.4 +/- 14.1% (SD); at 10 microM recovery averaged 97.3 +/- 4.2%. Much of the loss of carbacyclin on passage through placental circulation resulted from metabolism. Extracts of both fetal and maternal effluents from placenta perfused with carbacyclin contained a component which on reverse phase HPLC appeared less polar than carbacyclin. When analyzed by GC/MS as the methyl ester-trimethylsilyl ether, this component had a mass spectrum expected for 15-dehydro-carbacyclin. When the presumed metabolite was further converted to the methoxime, the mass spectrum was identical to published spectra for that derivative of 15-dehydro-carbacyclin. When extracts of fetal effluents were analyzed for 15-dehydro-carbacyclin metabolite as well as carbacyclin, it appeared that the metabolite accounted for the majority of the carbacyclin recovered. Most of the metabolite was apparently not formed in the fetal circulation, since when carbacyclin was added to the fetal afferent circulation, little 15-dehydro-carbacyclin was observed in either efferent fluid, and most of the perfused carbacyclin was recovered unaltered in the fetal effluent.

Biological Transport↗

PubMed1979

Qualitative study of prostaglandins in pregnant uterine tissues: effects of estradiol- 17 beta and betamethasone.

The presence of prostaglandins (PGs) E1 and F2 alpha in extracts of endometrial, myometrial and placental samples from 19 pregnant women at term, was investigated. The tissue samples were obtained from the following group of patients: untreated, A) with onset of labor, B) without onset of labor; pre-treated (without onset of labor); C) with betamethasone, D) with estradiol-17 beta. Thin-layer chromatography determinations demonstrated PGE1 presence in all tissue samples from the four groups. On the other hand, PGF2 alpha was consistently found in placenta and endometrial samples from groups A, C, and D, but it was absent in those from group B. The results suggest a physiological role of PGF2 alpha in human parturition, and a possible influence of estrogens and corticosteroids on the appearance of this prostaglandin.

Betamethasone↗

PubMed1981

Heparin facilitates the extraction of tissue fibronectin.

Extraction of fibronectin from two human tissues, lung parenchyma and placental villi, was facilitated by the incorporation of heparin into extraction media. The effect of heparin was additive to the effect of urea which is known to extract fibronectin. These experiments provide further evidence that fibronectin and glycosaminoglycans are associated in connective tissues and the use of heparin forms the basis for a simple method for extraction and quantitation of tissue fibronectin.

Dermatan Sulfate↗

PubMed1984

Mouse mammary tumor virus: specific methylation patterns of proviral DNA in normal mouse tissues.

The methylation state of endogenous mouse mammary tumor virus (MuMTV) proviral DNA was examined in normal mouse tissues. DNAs from various tissues were cleaved with the methylation-sensitive enzymes HhaI and HpaII and analyzed by Southern blotting. Tissue-specific MuMTV proviral DNA methylation patterns were found in the BALB/c, C3H, C57BL, GR/A, and GR-Mtv-2- mouse strains. MuMTV proviral DNA was hypomethylated in DNAs from the spleens and testes of all strains examined. The GR/A mouse strain, which was most thoroughly studied, also showed hypomethylation of MuMTV proviral DNA in bone marrow and placental tissues. Analysis of RNAs extracted from GR/A liver, mammary tumor, testes, placenta, and spleen tissues demonstrated that MuMTV proviral hypomethylation need not reflect significant proviral transcription.

Cell Transformation, Viral↗

PubMed1977

Presence of corticotropin in human placenta: demonstration of in vitro synthesis.

Immunoassayable (I) and bioassayable (B) ACTH activity is present in extracts of extensively washed human placental tissue and dispersed viable trophoblasts (Tr). Similar concentrations (I) (1.7-2.7 ng/g) are present in term placentas from elective cesarian section or vaginal delivery and from plancentas obtained by prostaglandin-induced abortion at 12, 15 and 18 weeks of gestation. Oral dexamethasone (8-12 mg) administered over an 8-48 h period prior to delivery does not significantly alter placental ACTH content. B-ACTH concentrations are 31-40% of I-ACTH. Sephadex G-50 filtration of term placental homogenates demonstrates two I-ACTH peaks, one eluting in the void volume and the other eluting in the region of synthetic hACTH1-39. When Tr are incubated in tissue culture medium, ACTH content of cells and of medium is significantly greater than pre-incubation levels.

Adrenocorticotropic Hormone↗

PubMed1984

Human placental steroid-sulfatase solubilized with a cholic-acid derivative: molecular mass, kinetic properties and susceptibility to glycosidases.

Human placental steroid-sulfatase was extracted nearly quantitatively from microsomes as well as from acetone dry powder of placenta homogenates using CHAPS as detergent. The solubilized enzyme was enriched 10-fold by ammonium sulfate precipitation and gel chromatography. The sulfatase extracted from both microsomes and acetone dry powder eluted as a single fraction on Sepharose 6B, but with different apparent molecular masses (390 and 270 kDa, respectively). Kinetic experiments with the sulfate esters of dehydroepiandrosterone, 16 alpha-hydroxydehydroepiandrosterone, estrone, and estriol as substrates or inhibitors indicated that the solubilized sulfatase was fully active. Both the particulate and the extracted enzyme showed higher affinities for the 16-unsubstituted than for the 16 alpha-hydroxylated substrates. Whereas a competitive inhibition was observed in mixed substrate incubations with dehydroepiandrosterone sulfate/16 alpha-hydroxydehydroepiandrosterone sulfate and estrone sulfate/estriol sulfate, diverse patterns of inhibition were obtained with dehydroepiandrosterone sulfate/estrone sulfate, depending on the sulfatase preparation used. However, evidence for the distinct nature of the steroid-sulfatase and the estrogen-sulfatase was not obtained. The membrane-bound, but not the solubilized enzyme was to a certain degree sensitive to lipase and acetone. The solubilized sulfatase strongly bound to ConA-Sepharose. This observation together with the elution by alpha-methyl mannoside were indicative of the presence of carbohydrates on the sulfatase. Since its enzymatic activity was markedly decreased by the effects of alpha-mannosidase and N-acetylglucosaminidase, a possible involvement of the carbohydrate moiety in the catalytic activity of the sulfatase might be considered.

Carbohydrates↗

PubMed2005

Nonhematopoietic stem cells of fetal origin--how much of today's enthusiasm will pass the time test?

Stem cells originating at fetal age are for many reasons superior as a material for the regenerative medicine purposes, when compared to their adult counterparts. While hematopoietic cells, isolated from fetal liver or cord blood, have been well known for a long time and have passed practical tests as clinical transplantation material, the non-hematopoietic cells are newly recognized, and the knowledge of their phenotype and differentiation potential is rather insufficient. We, and the others, have identified a subpopulation of cord blood cells phenotypically different from hematopoietic cells (CD34-, CD45-, CD29+, CD44+, CD51+, CD105+, SH-2, SH-3), in vitro plastic adherent, and capable of multilineage differentiation. The other candidates for multipotential stem cells are cells extracted from umbilical cord or placental tissue. The preliminary observations suggest, that these cells, phenotypically similar to the nonhematopoietic cord blood cells, are capable of extensive replication in vitro and of multilineage differentiation into a variety of tissues including cardiac muscle, bone and cartilage, adipocytes, and nerve cells. The other possible medical applications include "rejuvenation" of selected tissues and systems in senile patients, and therapeutical cloning - for both purposes, cells at the fetal stage of genetic regulation may be more useful than cells collected from adult donors. There is still, however, a high level of uncertainty concerning future medical applications of fetal stem cells. Their numbers and characteristics may differ from the preliminary observations, and their behavior in vivo may not fulfill the expectations originating from the in vitro studies. Finally, the autologous applications of stem cells collected at the stage of birth may need the involvement of technical and financial resources for the storage of frozen cell samples throughout the period of life of their potential user. Such procedure seems possible from technical point of view, but may be inadequately substantiated by the eventual advantages.

Cell Differentiation↗

PubMed1987

Purification and characterization of human placental aromatase cytochrome P-450.

Aromatase cytochrome P-450, which catalyzes the conversion of androgens to estrogens, was purified from human placental microsomes. The enzyme was extracted with sodium cholate, fractionated by ammonium sulfate precipitation, and subjected to column chromatography in the presence of its substrate, androstenedione, and the nonionic detergent, Nonidet P-40. The preparation exhibits a single major band when analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and has a specific content of 11.5 nmol of P-450/mg of protein. The purified enzyme displays spectroscopic properties typical of the ferric and ferrous forms of cytochrome P-450. Full enzymatic activity can be reconstituted with rabbit liver microsomal cytochrome P-450 reductase and Nonidet P-40. Purified aromatase cytochrome P-450 displays catalytic characteristics similar to the enzyme in intact microsomes in the aromatization of androstenedione, 19-hydroxyandrostenedione and 19-oxoandrostenedione. Testosterone and 16 alpha-hydroxytestosterone are aromatized at maximal rates similar to androstenedione, and all substrates exhibit relative affinities corresponding to those observed in microsomes. We have raised rabbit antibodies to the purified enzyme which show considerable specificity and sensitivity on immunoblots.

Androgens↗