Abortifacient Agents, Nonsteroidal ; pharmacology ; Animals ; Drugs, Chinese Herbal ; pharmacology ; Female ; Pinellia ; chemistry ; Plant Proteins ; pharmacology ; Pregnancy

C-Terminal carboxyl methylation of a human placental 23 kDa protein catalyzed by membrane-associated methyltransferase has been investigated. The 23 kDa protein substrate methylated was partially purified by DEAE-Sephacel, hydroxyapatite and Sephadex G-100 gel filtration chromatographies. The substrate protein was eluted on Sephadex G-100 gel filtration chromatography as a protein of about 29 kDa. In the absence of Mg2+, the methylation was stimulated by guanine nucleotides (GTP, GDP and GTPgammaS), but in the presence of Mg2+, only GTPgammaS stimulated the methylation which was similar to the effect on the G25K/rhoGDI complex. AFC, an inhibitor of C-terminal carboxyl methylation, inhibited the methylation of human placental 23 kDa protein. These results suggests that the substrate is a small G protein different from the G25K and is methylated on C-terminal isoprenylated cysteine residue. This was also confirmed by vapor phase analysis. The methylated substrate protein was redistributed to membrane after in vitro methylation, suggesting that the methylation of this protein is important for the redistribution of the 23 kDa small G protein for its putative role in intracellular signaling.
Cysteine/metabolism* ; Female ; GTP-Binding Proteins/metabolism* ; Guanine Nucleotides/pharmacology ; Human ; Methylation ; Placenta/metabolism* ; Placenta/enzymology ; Pregnancy ; Pregnancy Proteins/metabolism* ; Protein Methyltransferases/metabolism*

OBJECTIVE: This study aims to investigate whether dexamethasone (DEX) can down-regulate the PAR complex and disrupt the cell polarity in the palatal epithelium during palatal fusion. METHODS: Pregnant rats were randomly divided into control and DEX groups, which were injected intraperitoneally with 0.9% sodium chloride (0.1 mL) and DEX (6 mg·kg ⁻¹), respectively, every day from E10 to E12. The palatal epithelial morphology was observed using hematoxylin and eosin staining and scanning electron microscopy. Immunofluorescence staining, Western Blot analysis, and real-time polymerase chain reaction were performed to detect the expression of PAR3, PAR6, and aPKC. RESULTS: The incidence of cleft palate in DEX group (46.15%) was significantly higher than that in control group (3.92%), and the difference was statistically significant (χ2=24.335, P=0.00). DEX can also retard the growth of the palatal shelves and the short palatal shelves. The morphology and arrangement of MEE cells changed from polarized bilayer cells to nonpolarized monolayer ones. Additionally, the spherical structure decreased, which caused the cleft palate. PAR3 and PAR6 were only detected in the palatal epithelium, and aPKC was expressed in the palatal epithelium and mesenchyme. DEX can reduce the expression levels of PAR3, PAR6, and aPKC in the protein and gene levels. CONCLUSIONS DEX can down-regulate the complex gene expression in the MEE cells, thereby destroying the cell polarity and causing cleft palate.
Animals ; Carrier Proteins ; physiology ; Cell Polarity ; drug effects ; Cleft Palate ; etiology ; Dexamethasone ; pharmacology ; Epithelial Cells ; drug effects ; Female ; Glucocorticoids ; pharmacology ; Palate ; Pregnancy ; Rats

This study was aimed to investigate the activity of matrix metalloproteinase-9 (MMP-9) in umbilical cord blood mononuclear cells, the effect of recombinant human tumor necrosis factor alpha (rhTNF-alpha) on the activity of MMP-9 and cell migration capability of umbilical cord blood mononuclear cells, as well as the relationship between them. The different concentrations of TNF-alpha was added into the mononuclear cells. The activity of MMP-9 in umbilical cord blood mononuclear cells were analyzed by gelatinase spectrum. The cell migration capability was observed through the millicell carbine. The results showed that after umbilical cord blood mononuclear cells were cultured in serum-free RPMI 1640 medium for 24 and 48 hours, the activity of MMP-9 could be detected, the cell migration capability at 24 and 48 hours were (1.371 +/- 0.011)% and (1.384 +/- 0.014)% respectively, there was no statistical difference between them. Large dose of TNF-alpha (500 U/ml) led to the death of the cells; small doses of TNF-alpha (< 200 U/ml) up-regulated the activity of MMP-9, and as the concentration of TNF-alpha increased, the activity of MMP-9 was enhanced. Small doses of TNF-alpha (< 200 U/ml) improved the migration capability of umbilical cord blood mononuclear cells in a dose-dependent manner. It is concluded that small doses of TNF-alpha can upregulate the activity, enhance the migration capability of umbilical cord blood mononuclear cells, which probably accelerates cell homing after umbilical cord blood stem cell transplant and improves the hematopoietic reconstruction after transplantation.
Adult ; Cell Movement ; drug effects ; Cells, Cultured ; Female ; Fetal Blood ; cytology ; Humans ; Matrix Metalloproteinase 9 ; metabolism ; Pregnancy ; Recombinant Proteins ; pharmacology ; Tumor Necrosis Factor-alpha ; administration & dosage ; pharmacology

AIMTo study the effect of XW630 on expression of pro-oncogene c-myc in the long bones of fetal mice in vitro for postulating the mechanism by which XW630 exerts its effect on bone.

METHODSThe fetuses of pregnant mice were removed on day 16 of gestation, the long bones of the forelimbs of female fetal mice were freed of muscle and soft tissue and cultured in a specific device for 48 h in BGJb medium treated with 1 x 10(-7), 1 x 10(-8) and 1 x 10(-9) mol.L-1 XW630 in the final medium. After cultured for 48 h, the long bones were harvested and immunohistochemical analysis was performed for determination of c-Myc protein expression in epiphyseal plates. The areas of positive cells in the resting zone, proliferative zone and hypertrophic zone in epiphyseal plate were determined under image analytic system.

RESULTSWhen the concentration of XW630 in the medium was 1 x 10(-9) mol.L-1, the area of c-Myc positive cells increased in the proliferative zone compared with 1 x 10(-9) mol.L-1 in the estrone group, significant increase was also observed in the resting zone compared with the control group. When the concentration of XW630 in medium was 1 x 10(-8) or 1 x 10(-7) mol.L-1, stronger expression than that in the control group and the estrone group at the same concentration was observed in each of the three zones.

CONCLUSIONThe estrogenic effect of XW630 on bone was stronger than that of estrone. XW630 may promote proliferation and differentiation of chondroncytes by promoting c-Myc protein expression in chondroncytes. Thus, endochondral bone formation was enhanced.

Animals ; Chondrocytes ; metabolism ; Culture Techniques ; Estrone ; pharmacology ; Female ; Fetus ; Mice ; Piperazines ; pharmacology ; Pregnancy ; Proto-Oncogene Proteins c-myc ; metabolism ; Tetracyclines ; pharmacology ; Ulna ; drug effects ; metabolism ; Up-Regulation ; drug effects

OBJECTIVEMultiple signal transduction pathways, for example, Wnt signal transduction pathway, fibroblast growth factor (FGF), bone morphogenetic protein (BMP) etc, are involved in rat fetal lung development. Wnt signal has been shown to play important roles in regulating cell differentiation and proliferation. It is demonstrated that antenatal dexamethasone (DEX) use can induce lung dysplasia. A rat premature delivery model was developed in this study to compare the effects of DEX on antenatal rat fetal lung morphogenesis and the expressions of Wnt7b, beta-catenin and glycogen synthase kinase 3beta (GSK-3beta) genes in the lung of offspring on 19th day of embryo.

METHODSTwelve pregnant rats were divided into three groups randomly: small dose DEX group, large dose DEX group and control group with 4 rats in each group. The rats in the control group were injected with saline 0.5 ml/d; those in small dose DEX group were injected with DEX 0.4 mg/(kg.d), the rats in large dose DEX group were injected with DEX 0.8 mg/(kg.d), DEX was diluted to 0.5 ml by saline. On the 19th day of gestation, the fetuses were surgically taken out and the histologic structures of fetal rat lungs were observed with light microscope. The RT-PCR and Western-blot methods were used to detect the expressions of Wnt7b, GSK-3beta and beta-catenin genes mRNA and protein.

RESULTS(1) Changes of histologic structure included alveolar numbers: small dose DEX group (15.6 +/- 2.1), large dose DEX group (13.2 +/- 1.6), control group (20.8 +/- 2.0); thickness of alveolar septum: small dose DEX group (11 +/- 5) microm, large dose DEX group (11 +/- 4) microm, control group (13 +/- 7) microm; alveolar space: small dose DEX group (2483 +/- 1336) microm2, large dose DEX group (2924 +/- 1705) microm(2), and control group (1913 +/- 764) microm(2). All the parameters showed significant difference between DEX groups and control group. (P < 0.01 for all comparisons). (2) The expressions of Wnt7b (0.55 +/- 0.19, 0.64 +/- 0.54) and beta-catenin (2.03 +/- 0.58, 2.40 +/- 0.89) genes mRNA of the study groups were significantly higher as compared with those of the control group [Wnt7b (0.18 +/- 0.10), beta-catenin (1.77 +/- 0.54)] (P < 0. 05 for all comparisons) while the expressions of GSK-3beta (1.0 +/- 0.5) were lower than those of the control group (1.1 +/- 0.6) (P < 0. 05). The expressions of GSK-3beta protein in cytoplasm of the study groups [(26.6 +/- 19.7) microg, (10.7 +/- 7.4)microg] reduced gradually while beta-catenin's [(79.5 +/- 1.2) microg, (148.3 +/- 30.4) microg] in the nucleus enhanced simultaneously compared with the control group [(50.0 +/- 00.0) microg ].

CONCLUSIONSSmall dose of antenatal DEX usage can improve the fetal lung development, larger dose of DEX may have negative effect on rat fetal lung morphogenesis. Antenatal DEX usage can change the expressions of Wnt7b, GSK-3beta and beta-catenin genes mRNA and protein, these changes may result in paramorphia during pregnancy.

Animals ; Dexamethasone ; pharmacology ; Female ; Lung ; drug effects ; embryology ; metabolism ; Male ; Pregnancy ; Rats ; Rats, Sprague-Dawley ; Signal Transduction ; drug effects ; Wnt Proteins ; metabolism

OBJECTIVETo observe the effect of recombinant human erythropoietin (rHuEPO) on immune function of premature rats.

METHODSRHuEPO of 250 IU/(kg.t) or 500 IU/(kg.t) was administered to premature rats every other day for nineteen days. The control premature rats were received normal saline. The changes of hemoglobin (Hb), serum erythropoietin (EPO), red blood cell (RBC) immune function, T lymphocyte proliferative responsiveness, and production of tumor necrosis factor alpha (TNF-alpha) were observed.

RESULTSPremature rats showed lower levels on Hb, RBC immune function, T cell responsiveness and production of TNF-alpha compared with mature rats at birth. The postnatal declines of Hb and RBC immune function were lessened in the treated groups of premature rats, the higher dosage group of 500 IU/(kg.t) was more significant than the lower dosage group of 250 IU/(kg.t). When experiments were over, Hb of control premature rats was (7.72 +/- 0.89) g/dl, Hb of premature rats received 500 IU/(kg.t) was (10.08 +/- 0.90) g/dl (P < 0.01). C3b-R% of control premature rats was (11.00 +/- 0.95)%, C3b-R% of premature rats received 500 IU/(kg.t) was (17.75 +/- 1.04)% (P < 0.01). IC-R% in control premature rats was (12.83 +/- 1.33)%, IC-R% of premature rats received 500 IU/(kg.t) was (10.50 +/- 1.67)% (P < 0.01). The postnatal rise of T cell responsiveness and the production of TNF-alpha in premature rats increased in the treated groups, which was more significant in the higher dosage group of 500 IU/(kg.t) than in the lower dosage group of 250 IU/(kg.t). The OD index of control premature rats was 0.159 +/- 0.014, the OD index of premature rats received 500 IU/(kg.t) was 0.354 +/- 0.050 (P < 0.01). TNF-alpha in control premature rats was (0.270 +/- 0.014) ng/ml, TNF-alpha of premature rats received 500 IU/(kg.t) was (0.415 +/- 0.010) ng/ml (P < 0.01).

CONCLUSIONS(1) Premature rats had lower RBC immune function and T cell responsiveness and underproduction of TNF-alpha at birth. (2) Premature rats had an improvement with the RBC immune function after rHuEPO administration. (3) Premature rats had improvements with T cell responsiveness and TNF-alpha after rHuEPO administration, and there was a positive correction between the RBC immune function and T cell responsiveness with the production of TNF-alpha.

Animals ; Erythrocyte Count ; Erythrocytes ; drug effects ; immunology ; Erythropoietin ; blood ; pharmacology ; Female ; Hemoglobins ; analysis ; Pregnancy ; Rats ; Recombinant Proteins ; T-Lymphocytes ; drug effects ; immunology ; Tumor Necrosis Factor-alpha ; analysis

OBJECTIVETo measure the expression of phosphatidylinositol 3-kinase (PI3K) and protein kinase B (PKB) in liver tissue among low-birth-weight newborn rats treated with L-arginine (L-Arg) in early life, and to investigate the effect of L-Arg on insulin resistance.

METHODSEighteen pregnant rats were randomly divided into three groups: control, model and intervention (n=6 each). The control group was fed with normal protein feed (protein content=21%) during pregnancy to establish a normal-birth-weight newborn rat model, and the model and intervention groups were fed with low-protein feed (protein content=10%) during pregnancy to establish a low-birth-weight newborn rat model. Newborn rats from the three pregnant rat groups were also assigned to control, model and intervention groups. During 21 days of lactation, maternal rats in the control and model groups were fed with normal protein feed and normal drinking water, while maternal rats in the intervention group were fed with normal protein feed and drinking water rich in L-Arg (200 mg/kg·d). After ablactation, the three groups of newborn rats were fed with normal protein feed and normal drinking water. Liver tissue samples were collected from these newborn rats at 1, 3 and 8 weeks after birth. Protein expression of PI3K and PKB in liver tissue was measured by Western blot.

RESULTSAt 1 week after birth, the newborn rats in the intervention group had significantly higher protein expression of PI3K than in the model group (P=0.045), but there was no significant difference when compared with the control group (P=0.503). At 8 weeks after birth, the newborn rats in the intervention group had significantly higher protein expression of PKB than the model group (P=0.039), but there was no significant difference when compared with the control group (P>0.05).

CONCLUSIONSA supplement of L-Arg in early life can boost protein synthesis, increase protein expression of PI3K and PKB in liver tissue, promote insulin signaling and reduce insulin resistance.

Animals ; Animals, Newborn ; Arginine ; pharmacology ; Birth Weight ; Female ; Liver ; metabolism ; Male ; Phosphatidylinositol 3-Kinases ; genetics ; Phosphorylation ; Pregnancy ; Proto-Oncogene Proteins c-akt ; genetics ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo detect aberrant methylation in the promoter region of fetal endometriosis susceptibility gene homeobox-10 (HOXA10) in women with and without folic acid supplementation and explore the effect of folic acid in optimizing intrauterine environment.

METHODSThirty-six cord blood specimens were collected between January, 2010 and December, 2012 from pregnant women with endometriosis, including 22 with folic acid treatment and 15 without. Methylation-specific polymerase chain reaction (MSP) and bisulfite salt modified sequencing (BSP) were employed to detect aberrant methylation of HOXA10 gene in these specimens.

RESULTSThe methylation rate of HOXA10 gene differed significantly between pregnant women with endometriosis taking folic acid and those who did (P<0.05).

CONCLUSIONFolic acid treatment can significantly reduce the methylation rate of fetal endometriosis susceptibility gene HOXA10.

DNA Methylation ; drug effects ; Endometriosis ; genetics ; metabolism ; Female ; Fetus ; metabolism ; Folic Acid ; pharmacology ; therapeutic use ; Homeodomain Proteins ; genetics ; metabolism ; Humans ; Pregnancy ; Promoter Regions, Genetic

This study was aimed to investigate the expressions of genes hoxb2 and hoxb4 after interference of the proliferation and differentiation of hematopoietic stem cells (HSC) to the erythroid progenitors (CFU-E) in vitro by using all-trans retinoic acid (ATRA). The cord blood was collected from 12 cases of fetal placenta umbilical vein and cultured by using culture technique of HSC in vitro. The proliferation and differentiation of HSC to CFU-E were interfered with 6 x 10(-8) mol/L of ATRA. The expression levels of genes hoxb2 and hoxb4 in blank control and ATRA groups were detected by FQ-RT-PCR on day 3, 7 and 10 of culture. The results showed that the expressions of genes Hoxb2 and hoxb4 were a little on day 3, obviously increased on day 7 and reached highest level on day 10 in 2 groups. The expression level of hoxb4 on day 3, 7 and 10 in blank control group was obviously higher than expression level of hoxb2. As compared with blank control group, the expressions of genes hoxb2 and hoxb4 in the ATRA group were significantly up-regulated. It is concluded that the genes hoxb2 and hoxb4 all expressed in process of proliferation and differentiation to erythroid progenitors, which suggests that hoxb2 and hoxb4 relate to erythroid hematopoiesis, and the hoxb4 has more great relevance to erythroid hematopoiesis as compared with hoxb2. The ATRA (6 x 10(-8) mol/L) can up-regulate the expression of hoxb2 and hoxb4 significantly.
Cells, Cultured ; Female ; Fetal Blood ; cytology ; Gene Expression ; drug effects ; Hematopoietic Stem Cells ; drug effects ; Homeodomain Proteins ; genetics ; Humans ; Pregnancy ; Transcription Factors ; genetics ; Tretinoin ; pharmacology