OBJECTIVETo study the effects of advanced glycosylation end products (AGEs) on the production of fibronectin (FN) in human peritoneal mesothelial cells (HPMC) in vitro and the role of protein kinase C (PKC) in this course.

METHODSThe AGE-human serum albumin (HSA) (0, 100, 500, 1 000 microg/ml) was used in culture medium to stimulate the HPMC. The mRNA level of FN was measured with real-time PCR, moreover, the protein level of FN in HPMC was detected by ELISA. With the method of ELISA, the PKC activities were observed. Inhibitors or activators of PKC were used to observe the roles of PKC pathways on the AGE-HSA stimulated productions of FN in HPMC.

RESULTSAGE-HSA activated PKC in HPMC in a dose, time-dependent manner (P < 0.05). AGE-HSA up-regulated the expression of FN mRAN and protein in dose- and time-dependently (P < 0.01); PKC activator phorbol 12-myristate 13-acetate (PMA) induced FN expression, respectively depletion of PKC and calphostin C, a PKC inhibitor, effectively prevented both PMA and AGE-HSA-induced expression of the FN (P < 0.05).

CONCLUSIONAGEs can increase the activities of PKC. AGEs can directly increase FN expression in HPMC which may contribute to peritoneal fibrosis and this is regulated by PKC.

Cells, Cultured ; Epithelium ; secretion ; Fibronectins ; metabolism ; Glycation End Products, Advanced ; pharmacology ; Humans ; Peritoneum ; cytology ; Protein Kinase C ; metabolism

To study preliminarily the effect of Jiawei Bazhen decoction combined with oxytocin in promoting cervical ripening of full-term pregnancy women who were in the deficiency of qi and blood type through the syndrome differentiation of traditional Chinese medicine (TCM). 180 patients that met the inclusion criteria of the study were randomly divided into three groups: the control group(oxytocin group), the treatment group (Jiawei Bazhen decoction combined with oxytocin group), the blank control group (expected and observation group). Cervical maturity score (Bishop score), vaginal and cervical secretions fetal fibronectin (FFN), the result of induced labor, the result of mother and baby were observed in each group before and after treatment. The result comes out that the cervical Bishop score of pregnant women for treatment group were significantly higher than the control group and blank control group after treatment (P < 0.05). The FFN of pregnant women for the treatment group were significantly different from the control group and blank control group after treatment (P < 0.05). The pregnancy outcome of the three groups: the labor rate and rate of vaginal delivery of the treatment group were higher than the other two groups, and the difference was statistically significant (P < 0.05). The cesarean section rate of the treatment group was significantly lower than the other two groups, the difference was also statistically significant (P < 0.05). The three groups did not appear the phenomenon of neonatal asphyxia. Jiawei Bazhen decoction combined with oxytocin is effective in producing cervical ripening and induce labor. It is convenient, safe and reliable, for it is no obvious adverse effects on mother and fetus, but effective in reducing the rate of cesarean section, and playing a positive role in promoting natural delivery.
Adult ; Cervical Ripening ; drug effects ; metabolism ; Drug Therapy, Combination ; Drugs, Chinese Herbal ; administration & dosage ; Female ; Fibronectins ; secretion ; Humans ; Labor, Induced ; Oxytocin ; administration & dosage ; Pregnancy ; Pregnancy Complications ; drug therapy ; metabolism ; physiopathology ; Pregnancy Outcome ; Qi ; Vagina ; drug effects ; secretion ; Young Adult

PURPOSE: Deposition of polymeric IgA1 in the kidney mesangium is the hallmark of IgA nephropathy, but the molecular mechanisms of IgA-mediated mesangial responses and inflammatory injuries remain poorly understood. We hypothesize that Toll-like receptor 4 (TLR4) is involved in IgA-induced mesangial cell activation. MATERIALS AND METHODS: Mouse mesangial cells were stimulated with lipopolysaccharide (LPS) (1 microg/mL), IgA (20 microg/mL), or both, and TLR4 expression was measured by real time RT-PCR and Western blot. Intracellular responses to LPS or IgA were assessed by Western blot for ERK1/2, JNK, p38 MAP kinases (MAPKs), Ikappa-Balpha degradation and fibronectin secretion. MCP-1 secretion was assessed by ELISA. Small interfering RNA (siRNA) of TLR4 was used to confirm that the effects were caused by TLR4 activity. RESULTS: LPS- or IgA-treatment upregulated the levels of TLR4 mRNA and protein in cultured MMC at 24 h. LPS and IgA induced rapid phosphorylation of MAPKs, but degradation of Ikappa-Balpha was observed only in LPS-treated MMC. LPS, but not IgA, induced increased secretion of MCP-1 and fibronectin at 24 h or 48 h. Combined LPS and IgA treatment did not cause additional increases in TLR4 mRNA and protein levels or Ikappa-Balpha degradation, and MCP-1 and fibronectin secretions were less than with LPS alone. LPS- or IgA-induced TLR4 protein levels and MAPK activation were inhibited by transfection with TLR4 siRNA. CONCLUSION: These results indicate that the activation of MAPKs and MCP-1 secretion are mediated by TLR4, at least in part, in IgA-treated mesangial cells. TLR4 is involved in mesangial cell injury by induction of pro-inflammatory cytokines in IgA nephropathy.
Animals ; Chemokine CCL2/secretion ; Enzyme-Linked Immunosorbent Assay ; Extracellular Signal-Regulated MAP Kinases/metabolism ; Fibronectins/secretion ; Glomerulonephritis, IGA/*metabolism ; I-kappa B Proteins/metabolism ; Mesangial Cells/*metabolism/secretion ; Mice ; Mice, Transgenic ; Phosphorylation ; RNA Interference ; RNA, Messenger/metabolism ; *Signal Transduction ; Toll-Like Receptor 4/antagonists & inhibitors/genetics/*metabolism

BACKGROUNDConnective tissue growth factor (CTGF) contributes greatly to renal tubulointerstitial fibrosis, which is the final event leading to end-stage renal failure. This study was designed to investigate the effects of CTGF antisense oligodeoxynucleotides (ODNs) on the expressions of plasminogen activator inhibitor-1 (PAI-1) and fibronectin in renal tubular cells induced by transforming growth factor beta1 (TGF-beta1) in addition to the role of CTGF in the accumulation and degradation of renal extracellular matrix (ECM).

METHODSA human proximal tubular epithelial cell line (HKC) was cultured in vitro. Cationic lipid-mediated CTGF antisense ODNs were transfected into HKC cells. After HKC cells were stimulated with TGF-beta1 (5 microg/L), the mRNA levels of PAI-1 and fibronectin were measured by RT-PCR. Intracellular PAI-1 protein synthesis was assessed by flow cytometry. The secreted PAI-1 and fibronectin in the medium were determined by Western blot and ELISA, respectively.

RESULTSTGF-beta1 was found to induce tubular CTGF, PAI-1, and fibronectin mRNA expression. PAI-1 and fibronectin mRNA expression induced by TGF-beta1 was significantly inhibited by CTGF antisense ODNs. CTGF antisense ODNs also inhibited intracellular PAI-1 protein synthesis and lowered the levels of PAI-1 and fibronectin protein secreted into the medium.

CONCLUSIONSCTGF may play a crucial role in the accumulation and degradation of excessive ECM during tubulointerstitial fibrosis, and transfecting CTGF antisense ODNs may be an effective way to prevent renal fibrosis.

Cells, Cultured ; Connective Tissue Growth Factor ; Extracellular Matrix ; metabolism ; Fibronectins ; genetics ; secretion ; Humans ; Immediate-Early Proteins ; genetics ; physiology ; Intercellular Signaling Peptides and Proteins ; genetics ; physiology ; Kidney Tubules ; metabolism ; Oligonucleotides, Antisense ; pharmacology ; Plasminogen Activator Inhibitor 1 ; genetics ; secretion ; RNA, Messenger ; analysis ; Transfection ; Transforming Growth Factor beta ; pharmacology ; Transforming Growth Factor beta1

OBJECTIVETo investigate the effect of Moutan Cortex on mesangial proliferation and basement membrane thickening induced by advanced glycation end products (AGEs).

METHODThe glomerular mesangial cells (MC) injury model was established by inducing by AGEs. The cell were divided into 6 groups: the blank group ( BSA, 200 mg L-1) , the model group (AGEs, 200 mg L-1), the positive control group (AG, 10 mmol L L-1), and drug administration groups, namely the Moutan Cortex-treated high-dose group (2 x 10(-4) g mL(- 1)), the Moutan Cortex-treated medium-dose group (1 x 10(-4) g mL-1 ), and the Moutan Cortex-treated low-dose group (0. 5 x 10(-4) g . mL(-1)). The MTT method was performed to observe the effect of Moutan Cortex on the proliferation of MC. The content of fibronectin (FN) and collagen secretion 1V (Col IV) in cell supernatant were detected by ELISA kits. The western blot analysis was carried out to observe the FN expression. The Real-time PCR analysis was applied to examine the Col IV mRNA expression.

RESULTAGEs significantly increased AGEs-induced MC proliferation and FN and Col 1V secretion. The western blot analysis showed that MC could down-regulate the FN expression of MC secretion. According to the results of the real-time PCR assay, MC could down-regulate AGEs-induced MC secretion Col IV mRNA expression.

CONCLUSIONMC had a certain protective effect on MC cultured under AGEs conditions. MC could remarkably inhibit the composition and secretion of Col IV and FN in matrix and the basement membrane thickening, and provide an experimental basis for the treatment of diabetic nephropathy.

Animals ; Basement Membrane ; drug effects ; metabolism ; Cell Line ; Cell Proliferation ; drug effects ; Collagen Type IV ; genetics ; secretion ; Drugs, Chinese Herbal ; pharmacology ; Fibronectins ; biosynthesis ; Gene Expression Regulation ; drug effects ; Glycation End Products, Advanced ; adverse effects ; Mesangial Cells ; cytology ; drug effects ; metabolism ; secretion ; Paeonia

AIMTo study the mechanisms of anti-diabetic nephropathy of rhein on cultured human mesangial cells (HMCs).

METHODSTo mimic the hyperglycemic (HG) environment of diabetic nephropathy, 30 mmol x L(-1) glucose were added to 10% FBS RPMI 1640. The HMCs were treated with rhein for 8, 24, 48 or 72 h, at these time, the bioactivity, total activity of transforming growth factor-beta1 (TGFbeta1), activity of p38MAPK (p38 mitogen-activated protein kinases, by using immunoprecipitate and Western blot), MMP-2 (matrix metalloproteinase-2), and MMP-9 (matrix metalloproteinase-9, by using gelatinase zymography) and the proliferation of HMCs in high glucose media were measured. Meanwhile the levels of secretion of FN in cultured HMCs were measured.

RESULTSThe results showed that rhein markedly inhibit the proliferation of HMCs, significantly reduce the bioactivity of TGFbeta1 and FN secretion in HMCs, and decrease the increased activity of p38MAPK, but showed no action on the activities of MMP-2 and MMP-9.

CONCLUSIONRhein reduced the secretion of FN and inhibited the proliferation of HMCs may through inhibiting the bioactivities of TGFbeta1 and p38MAPK.

Animals ; Anthraquinones ; pharmacology ; Cell Proliferation ; drug effects ; Cells, Cultured ; Culture Media ; Epithelial Cells ; cytology ; metabolism ; Fibronectins ; secretion ; Glomerular Mesangium ; cytology ; metabolism ; Glucose ; antagonists & inhibitors ; pharmacology ; Humans ; Lung ; cytology ; metabolism ; Matrix Metalloproteinase 2 ; metabolism ; Matrix Metalloproteinase 9 ; metabolism ; Mink ; Transforming Growth Factor beta ; metabolism ; Transforming Growth Factor beta1 ; p38 Mitogen-Activated Protein Kinases ; metabolism