OBJECTIVETo observe the effects of two main isoflavones, daidzein and genistein on the bone-nodule formation in rat calvaria osteoblasts in vitro.

METHODSOsteoblasts obtained from newborn Sprague-dawley rat calvaria were cultured for several generations. The second generation cells were cultured in Minimum Essential Medium supplemented with ascorbic acid and Na-beta-glycerophosphate for several days, in the presence of daidzein and genistein, with or without the estrogen receptor antagonist ICI 182780. Number of nodules was counted at the end of the incubation period (day 20) by staining with Alizarin Red S calcium stain. The release of osteocalcin, as a marker of osteoblast activity, was also determined on day 7 and day 12 during the incubation period.

RESULTSCompared with the control, the numbers of nodules were both increased by incubation with daidzein and genistein. 17 beta-estradiol was used as a positive control and proved to be a more effective inducer of the increase in bone-nodules formation that daidzein and genistein. The release of osteocalcin into culture media was also increased in the presence of daidzein and genistein, as well as 17 beta-estradiol on day 7 and day 12 (day 12 were higher). The estrogen receptor antagonist ICI 182780 completely blocked the genistein- and 17 beta-estradiol-induced increase of nodule numbers and osteocalcin release in osteoblasts. However, the effects induced by daidzein could not be inhibited by ICI 182780.

CONCLUSIONThese findings suggest that geinistein can stimulate bone-nodule formation and increase the release of osteocalcin in rat osteoblasts. The effects, like those induced by 17 beta-estradiol, are mediated by the estrogen receptor dependent pathway. Daidzein also can stimulate bone-nodule formation and increase the release of osteocalcin in rat osteoblasts, but it is not, at least not merely, mediated by the estrogen receptor dependent pathway.

Animals ; Cells, Cultured ; Genistein ; pharmacology ; Isoflavones ; pharmacology ; Osteoblasts ; cytology ; drug effects ; metabolism ; Osteocalcin ; biosynthesis ; Osteogenesis ; drug effects ; Rats ; Rats, Sprague-Dawley ; Skull ; cytology ; drug effects ; metabolism

OBJECTIVETo study the tolerance of rats induced by 28 day pretreatment with low dosage of dimthoate and the toxic effects challenged by higher dosage of dimethoate, and to investigate the change of M receptor and the mechanism of tolerance formation.

METHODSSD rats were given 25 mg/kg dimethoate daily(sc) while control group was given saline daily instead for 28 days. The activity of whole blood acetylcholinesterase (AChE) was examined. On the 29th day three groups of administrated rats were challenged by saline solution, 50 mg/kg and 100 mg/kg dimethoate, respectively. The density and mRNA level of brain M(1), M(2) receptor were determined. Lymphocytes of peripheral blood were isolated, and basal, inducible M(3) gene expression were measured by RT-PCR.

RESULTSDuring pretreatment, blood AChE activity decreased continually, it reached the lowest on the 13th day. And it decreased more after exposed to higher dosage of dimethoate. Brain AChE activity in the pretreated groups was lower than that in control group and decreased with the increase in challenging dosage. The density of M(1) receptor in negative control, pretreated, and 50, 100 mg/kg challenging groups were 979.15, 856.54, 539.46, 539.14 fmol/mg pro respectively. The change in relative levels of mRNA of M(1) receptor (2.59, 2.47, 2.20, 1.81) were consistent with the density of receptor but the level declined continually as the challenging dosage increased. The density of M(2) receptor were 507.38, 611.11, 548.42, 337.47 fmol/mg pro respectively, which were not obviously affected by pretreatment but decreased as the challenging dosage increased. The change in levels of M(2) receptor mRNA was not obvious. The basal gene expression of M(3) receptor mRNA was not different among all experimental groups while the inducible gene expression decreased with the increase in challenging dosage.

CONCLUSIONLow level dosage of dimethoate could induce animals to tolerate dimethoate toxicity. Reduction of M(1) receptor density which may be induced by the decrease in its gene expression may be the mechanism of tolerance. The change of M(3) receptor mRNA inducible expression in lymphocyte accorded with M(1) receptor mRNA expression in the brain.

Animals ; Brain ; metabolism ; Dimethoate ; toxicity ; Gene Expression ; drug effects ; Insecticides ; toxicity ; Lymphocytes ; metabolism ; Male ; Maximum Tolerated Dose ; RNA, Messenger ; genetics ; metabolism ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Receptors, Muscarinic ; genetics ; Reverse Transcriptase Polymerase Chain Reaction ; Time Factors

OBJECTIVETo study the expression of muscarinic receptor M(3) gene in peripheral blood lymphocytes of workers exposed to organophosphorus pesticides (OPPs) and to explore its role in the adverse effects of OPPs.

METHODSThe lymphocytes of peripheral blood from 33 workers exposed to dimethoate and 15 control people were isolated and treated with saline and dimethoate respectively in vitro. RT-PCR technique was used to determine M(3) gene expression. Basal and inducible gene expression levels were measured.

RESULTSThere was no significant difference in basal gene expression level between exposed group and control group, while the inducible gene expression level was significantly higher in exposure group (1.92 +/- 1.07) than in control group (1.22 +/- 0.19) and basal level (1.49 +/- 0.45, P < 0.05). No differences in basal and inducible gene expression level were found between male and female people in both exposed and control group. The level of inducible M(3) gene expression increased with the increase in length of exposure time [< 5 a: (1.69 +/- 0.95), 5 - 25 a: (1.91 +/- 1.03), > 25 a: (2.09 +/- 1.25), the latter was significantly different from that of < 5 a (P < 0.05)].

CONCLUSIONAfter long-term exposure to OPPs, the basal M(3) receptor gene expression level in the exposed workers did not show any difference from the control group, but the inducible gene expression level (treated with dimethoate in vitro) was increased and related to the extent of exposure to dimethoate.

Dimethoate ; blood ; poisoning ; Female ; Gene Expression ; Humans ; Insecticides ; blood ; poisoning ; Lymphocytes ; cytology ; metabolism ; Male ; Occupational Exposure ; analysis ; RNA, Messenger ; genetics ; metabolism ; Receptor, Muscarinic M3 ; genetics ; Reverse Transcriptase Polymerase Chain Reaction

Mesial temporal lobe epilepsy (MTLE) is associated with severe neuronal death and reactive gliosis in hippocampus. However, the molecular mechanisms underlying these pathological changes remain unanswered. ERK has been reported chronically activated in reactive glia of human epileptic hippocampus. In the present study, we investigated which of the downstream signaling molecules of ERK would be involved in MTLE. Western blot analysis demonstrated that CREB and p90RSK were strongly activated in MTLE patients. Increase in the active forms of CREB and p90RSK resulted not only from the increase in their phosphorylation levels but also from the increase in the protein levels. Activation of CREB and p90RSK was noted in the whole subfields of hippocampus with Ammon's horn sclerosis (AHS) representing a distinctive cellular distribution. However, the common major change was present in proliferating reactive astrocytes. In contrast, their activation was not significant in adjacent temporal lobes despite the presence of a number of astrocytes expressing high levels of GFAP. Our results demonstrate that chronic activation CREB and p90RSK in the epileptic hippocampus may be closely associated with the histopathological changes of AHS.
Blotting, Western ; DNA-Binding Protein, Cyclic AMP-Responsive/*metabolism ; Enzyme Activation ; Epilepsy/enzymology/*metabolism ; Epilepsy, Temporal Lobe/enzymology/metabolism ; Hippocampus/enzymology/*metabolism ; Human ; Immunohistochemistry ; Mitogen-Activated Protein Kinases/metabolism ; Ribosomal Protein S6 Kinases, 90kD/*metabolism ; Signal Transduction ; Support, Non-U.S. Gov't ; Temporal Lobe/enzymology/metabolism

Catecholamines are among the first molecules that displayed a kind of response to prolonged or repeated stress. It is well established that long-term stress leads to the induction of catecholamine biosynthetic enzymes such as tyrosine hydroxylase (TH) and dopamine beta-hydroxylase (DBH) in adrenal medulla. The aim of the present study was to evaluate the effects of ginseng on TH and DBH mRNA expression. Repeated (2 h daily, 14 days) immobilization stress resulted in a significant increase of TH and DBH mRNA levels in rat adrenal medulla. However, ginseng treatment reversed the stress-induced increase of TH and DBH mRNA expression in the immobilization-stressed rats. Nicotine as a ligand of the nicotinic acetylcholine receptor (nAChR) in adrenal medulla stimulates catecholamine secretion and activates TH and DBH gene expression. Nicotine treatment increased mRNA levels of TH and DBH by 3.3- and 3.1-fold in PC12 cells. The ginseng total saponin exhibited a significant reversal in the nicotine-induced increase of TH and DBH mRNA expression, decreasing the mRNA levels of TH and DBH by 57.2% and 48.9%, respectively in PC12 cells. In conclusion, immobilization stress induced catecholamine biosynthetic enzymes gene expression, while ginseng appeared to restore homeostasis via suppression of TH and DBH gene expression. In part, the regulatory activity in the TH and DBH gene expression of ginseng may account for the anti-stress action produced by ginseng.
Adrenal Medulla ; Animals ; Catecholamines ; Dopamine ; Dopamine beta-Hydroxylase ; Down-Regulation ; Gene Expression ; Homeostasis ; Immobilization ; Nicotine ; Panax ; PC12 Cells ; Rats ; Receptors, Nicotinic ; RNA, Messenger ; Saponins ; Tyrosine ; Tyrosine 3-Monooxygenase

OBJECTIVE: To explore the efficacy of serum CA19-9 determination for the detection of endometriosis. DESIGNS: A retrospective analysis. MATERIALS AND METHODS: Preoperative serum CA19-9 and CA125 concentrations were measured in 53 patients with surgically and/or histologically diagnosed endometriosis. The extent of disease was staged according to the revised American Fertility Society (1985) classification system. RESULTS: Serum CA19-9 levels were elevated (>or=37 U/ml) in 13 (24.5%) out of 53 patients with endometriosis and serum CA125 levels were elevated (>or=35 U/ml) in 13 (24.5%). Either CA19-9 or CA125 levels in serum were elevated in 18 (34.0%) patients. In patients with stage III or IV endometriosis, elevated CA19-9 levels were noted in 11 (31.4%) of 35 patients, while CA125 in 12 (34.3%). Either CA19-9 or CA125 levels in serum were elevated in 15 (42.9%) of 35 patients with stage III or IV. CONCLUSION: These results suggest that serum CA19-9, in addition to CA125, measurement could help in identifying patients with moderate to severe endometriosis.
Classification ; Endometriosis* ; Female ; Fertility ; Humans ; Retrospective Studies

The protein encoded by the Bcl-2 proto-oncogene has been shown to prolong cell su-rvival by preventing cell death(Apoptosis) induced by many insults including cancer therap-eutic drugs. Recently many researches have elucidated the bcl-2 expression in several hu-man solid cancers. However, there is still a few avaiable data to determine the role of Bcl-2 expression in the ovarian carcinogenesis and its prognostic significance in ovarian can-cers. Hence, we examined the expression of Bcl-2 in 68 ovarian epithelial cancers using immunohistochemistry and determined whether Bcl-2 expression has prognostic significance in the ovarian epithelial cancers. We found Bcl-2 expression(>5% positive cell) in 31 patients(40%). Bcl-2 expression were exclusively negative in the mucinous type of the ovarian epithelial cancer. Bcl-2 exp- ression was not correlated with tumor stage(stage I & II vs III & IV, p=0.63). The patients with Bcl-2 positivity had lower recurrence rate than the patients with negativity at the se- cond look operation(p<0.01). Although there was a trend that the patients with Bcl-2 posi- tivity had better acturial survival than the patient with negativity, the stastical significance was not present(3 years acturial survival;+vs-;63%, 29%;p>0.05). These results sug- gest the Bcl-2 expression appears an early event in the ovarian carcinogenesis and has an inhibiting role in progression of ovarian tumor.
Carcinogenesis ; Humans ; Immunohistochemistry ; Mucins ; Ovarian Neoplasms* ; Proto-Oncogenes ; Recurrence

Background: Recently, mesenchymal stem cells therapy has been performed in dogs, although the outcome is not always favorable. Objectives: To investigate the therapeutic efficacy of mesenchymal stem cells (MSCs) using dog leukocyte antigen (DLA) matching between the donor and recipient in vitro. Methods: Canine adipose-derived MSCs (cA-MSCs) isolated from the subcutaneous tissue of Dog 1 underwent characterization. For major DLA genotyping (DQA1, DQB1, and DRB1), peripheral blood mononuclear cells (PBMCs) from two dogs (Dogs 1 and 2) were analyzed by direct sequencing of polymerase chain reaction (PCR) products. The cA-MSCs were co-cultured at a 1:10 ratio with activated PBMCs (DLA matching or mismatching) for 3 days and analyzed for immunosuppressive ( IDO, PTGS2, and PTGES ), inflammatory (IL6 and IL10 ), and apoptotic genes (CASP8, BAX, TP53, and BCL2) by quantitative real-time reverse transcriptase-PCR. Results: cA-MSCs were expressed cell surface markers such as CD90+/44+/29+/45- and differentiated into osteocytes, chondrocytes, and adipocytes in vitro. According to the Immuno Polymorphism Database, DLA genotyping comparisons of Dogs 1 and 2 revealed complete differences in genes DQA1, DQB1, and DRB1. In the co-culturing of cA-MSCs and PBMCs, DLA mismatch between the two cell types induced a significant increase in the expression of immunosuppressive (IDO/PTGS2) and apoptotic (CASP8/BAX) genes. Conclusions The administration of cA-MSCs matching the recipient DLA type can alleviate the need to regulate excessive immunosuppressive responses associated with genes, such as IDO and PTGES. Furthermore, easy and reliable DLA genotyping technology is required because of the high degree of genetic polymorphisms of DQA1, DQB1, and DRB1 and the low readability of DLA 88.

Background: Recently, mesenchymal stem cells therapy has been performed in dogs, although the outcome is not always favorable. Objectives: To investigate the therapeutic efficacy of mesenchymal stem cells (MSCs) using dog leukocyte antigen (DLA) matching between the donor and recipient in vitro. Methods: Canine adipose-derived MSCs (cA-MSCs) isolated from the subcutaneous tissue of Dog 1 underwent characterization. For major DLA genotyping (DQA1, DQB1, and DRB1), peripheral blood mononuclear cells (PBMCs) from two dogs (Dogs 1 and 2) were analyzed by direct sequencing of polymerase chain reaction (PCR) products. The cA-MSCs were co-cultured at a 1:10 ratio with activated PBMCs (DLA matching or mismatching) for 3 days and analyzed for immunosuppressive ( IDO, PTGS2, and PTGES ), inflammatory (IL6 and IL10 ), and apoptotic genes (CASP8, BAX, TP53, and BCL2) by quantitative real-time reverse transcriptase-PCR. Results: cA-MSCs were expressed cell surface markers such as CD90+/44+/29+/45- and differentiated into osteocytes, chondrocytes, and adipocytes in vitro. According to the Immuno Polymorphism Database, DLA genotyping comparisons of Dogs 1 and 2 revealed complete differences in genes DQA1, DQB1, and DRB1. In the co-culturing of cA-MSCs and PBMCs, DLA mismatch between the two cell types induced a significant increase in the expression of immunosuppressive (IDO/PTGS2) and apoptotic (CASP8/BAX) genes. Conclusions The administration of cA-MSCs matching the recipient DLA type can alleviate the need to regulate excessive immunosuppressive responses associated with genes, such as IDO and PTGES. Furthermore, easy and reliable DLA genotyping technology is required because of the high degree of genetic polymorphisms of DQA1, DQB1, and DRB1 and the low readability of DLA 88.

No abstract available.
Advanced Cardiac Life Support* ; Cardiopulmonary Resuscitation*