In this study,primary serum-free cultured rat granulosa cells (rGCs) were used as a cellular model to investigate the effects of fenvalerate on progesterone production. Various concentrations (0,1,5,25,125and625μM) of fenvalerate were added to the cell cultures for 24h.rGCs were stimulated by compounds such as follicle-stimulating hormone (FSH), 8-bromo-cAMP or 22(R)-hydroxycholesterol (22R-HC) Progesterone production and intracellular cAMP content were measured in control and treated groups.Expression of P450 side chain cleavage enzyme (P450scc) and steroidogenic acute regulatory protein(StAR)were monitored by real-time PCR and Western blotting.Results showed that fenvalerate inhibited basal progesterone production in rGCs in the absence of stimulators.This inhibition was stronger in the presence of FSH and was not fully reversed by 8-bromo-cAMP or 22R-HC.The increase of cAMP content,stimulated by FSH, was inhibited by fenvalerate implicating that the intracellular cAMP-dependent signal pathway was involved.Fenvalerate reduced mRNA and protein expression of P450scc.These results suggested that multi-site inhibition of progesterone production by fenvalerate including a cAMP-dependent protein kinase pathway and reduction on P450scc gene expression and/or its enzymatic activity in rGCs.

Objective: Urinary bladder hyperplasia associated with terephthalic acid (TPA) treatment was examined with concomitant use of sodium bicarbonate (NaHCO3) or hydrochlorothiazide to allow assessment of the relationship among bladder stones,epithelial hyperplasia,and corresponding cell cycle checkpoint gene expression in Sprague–Dawley (SD)rat. Methods:A total of 112 weanling male SD rats that divided between six groups were given basal diet (control), diets containing 5% TPA or in combination with either 4% sodium NaHCO3 or 0.02% hydrochlorothiazide.After 90-day feeding,bladder samples were collected for histopathological diagnoses,and immunohistochemical method was used to characterize the expression of p16Ink4a,cyclin D1, CDK4, EGFr and cyclin E in relation to that of proliferating cell nuclear antigen (PCNA). Results:In TPA treatment groups, bladder stone incidence was 40% (21/52) with 14 cases of proliferative bladder.In control and other groups,neither stone nor epithelial cell proliferation was diagnosed.PCNA-positive focal hyperplasic lesions involved all epithelial layers.Overexpressions of cyclin D1, CDK4, EGFr are found in the corresponding lesion. p16Ink4a nuclear staining reduced in proliferative bladders especially with a great quantity of stone.In addition,no positive expression was detected on cyclin E. Conclusion: The present study provides a strong evidence of a link between induction of bladder hyperplasia, deregulation of the p16Ink4a-cyclin D1/CDK4 pathway,and abnormal EGFr mediated signal transduction pathway.

Carbaryl is known to impede female reproductive function,however,the mechanisms through which the adverse effects are mediated are not clearly elucidated.In order to get insight into the mechanisms,this study was conducted to raise fresh concerns about the potential effects of carbaryl on steroidogenesis by primary human granulosa-lutein cells(hGLCs)and explore the possible nature of this action.hGLCs were co-incubated with various concentrations of carbaryl at 0,1,5,25,125μmol/L for 24 h to examine effects of this carbamate pesticide on progesterone accumulation. We observed that the carbaryl inhibited basal and FSH-induced progesterone production in a dose-dependent manner.We also investigated the effects of carbaryl on 22(R)-hydroxycholesterol(22R-HC)-stimulated progesterone yield,basal and FSH-stimulated StAR gene expression and cyclic adenosine monophosphate(cAMP)production,as well as forskolin(non-specific activator of adenylyl cyclase)-induced progesterone and cAMP production of hGLCs.We found that the decreased progesterone biosynthesis was accompanied with a reduced cAMP abundance on both basal and FSH-induced condition.Furthermore, our results demonstrated that the 22R-HC could remove the carbaryl-induced restraint of progesterone biosynthesis,suggesting that carbaryl caused a disruption of cholesterol transport across mitochondrial membranes,which was further confirmed by the observation that carbaryl inhibited the gene expression of steroidogenic acute regulatory protein (StAR).In addition,the inhibitory effects of carbaryl on progesterone and cAMP production were completely reversed by addition of forskolin to the cell culture,which indicated a repaired site on the upstream components of adenylate cyclase or adenylate cyclase per se by carbaryl in the cAMP-mediated signal pathway.All the effects mentioned above were not due to a detrimental action of carbaryl on cell viability by MTS assay.In conclusion,carbaryl may inhibit steroidogenesis,at least in part,by obstructing the delivery of cholesterol over mitochondrial membranes and attenuating cAMP generation.

The effect of terephthalic acid (TPA) on urinary bladder carcinogenesis was examined.Male Wistar rats were initiated by injection of N-Methyl-N-Nitrosourea (MNU)(20mg/kg b.w.ip) twice a week for 4 weeks,then given basal diet containing 5% TPA,5% TPA plus 4% Sodium bicarbonate (NaHCO3) and 1%TPA for the next22weeks, and then euthanized. 5%TPA treatment induced a high incidence of urinary bladder calculi and a large amount of precipitate.Though 5% TPA plus 4% Sodium bicarbonate(NaHCO3)and 1% TPA treatment did not induce urinary bladder calculi formation,they resulted in a moderate increase in urinary precipitate.Histological examination of urinary bladder revealed that MNU-5% TPA treatment resulted in a higher incidence of simple hyperplasia,papillary or nodular hyperplasia (PN hyperplasia), papilloma and cancer than MNU control.MNU-5% TPA plus 4% Sodium bicarbonate (NaHCO3) and 1% TPA treatment increased slightly the incidence of simple hyperplasia and PN hyperplasia(not statistically significant). The major elements of the precipitate are phosphorus, potassium, sulfur, chloride, calcium and TPA. The present study indicated that the calculi induced by TPA had a strong promoting activity on urinary bladder carcinogenesis and the precipitate containing calcium terephthalate (Ca TPA) may also have weak promoting activity on urinary bladder carcinogenesis.

Purpose:Folate deficiency and reduced DNA repair capacity are established risk factors for squamous cell carcinoma of the head and neck(SCCHN). We hypothesized that polymorphisms of the thymidylate synthase(TYMS)gene,which regulates a key enzyme in folate metabolism required for DNA synthesis and repair, are associated with SCCHN risk. Experimental Design:In a hospital-based case-control study of 704 SCCHN cases and 1,085 controls, frequency matched by age,sex, nd ethnicity,we genotyped the TSER(thymidylate synthase in the 5'-untranslated enhanced region)and TS3ˊUTR(thymidylate synthase in the 3'-untrans-lated region) polymorphisms. Results: The TS3ˊUTR 0bp/0bp genotype was associated with a significantly decreased risk of SCCHN[adjusted odd ratio(OR)=0.67,95%confidence interval (CI)=0.47-0.94] compared with the 6bp/6bp genotype, but the TSER polymorphism had no main effect on risk of SCCHN. When we evaluated the two polymorphisms together by the number of protective alleles(the TSER 3R and TS3'UTR 0bp alleles), we found that the combined genotypes with four protective alleles(the TSER 3R3R and TS3'UTR 0bp/0bp) was associated with significantly decreased SCCHN risk (OR=0.60, 95%CI=0.37-0.98). In addition,the TS3ˊUTR 0bp genotypes were associated in an allele dose-dependent manner with a decreased risk of overall stage IV oral cancer (OR=0.84, 95%CI=0.52-1.34 for the 6bp/0bp genotype and OR=0.26, 95%CI=0.08-0.87 for the 0bp/0bp genotype; Ptrend=0.035). Conclusion: The TSER and TS3'UTR polymorphisms are associated with SCCHN risk. The TSER 3R and TS3ˊUTR 0bp alleles seemed to jointly protect against SCCHN. In particular, the 0bp allele seemed to protect against oral cancer progression.