The present study is aimed to investigate the in vitro metabolic interconversion between baicalin (BG) and baicalein (B) in rat liver, kidney, intestine and bladder. BG and B were separately incubated with rat hepatic, renal, and intestinal microsomes, as well as bladder homogenates, for 30 min. The metabolites were identified and quantified by HPLC and metabolic kinetic parameters were obtained by fitting the data to the Michaelis-Menten equation. In hepatic microsomes, renal microsomes and bladder homogenates, but not in intestinal microsomes, BG was transformed into B, the hydrolysis metabolite of BG, with K(m) values being (44.65 +/- 6.01), (92.73 +/- 11.41), (74.60 +/- 3.68) micromol x L(-1), respectively, and V(max) values being (12.32 +/- 0.56), (3.30 +/- 0.18), (5.93 +/- 0.12) micromol x min(-1) x g(-1) (protein), respectively. In incubations with hepatic, renal, and intestinal microsomes and bladder homogenates, B was also transformed into BG, the glucuronidation metabolite of B, with K(m) values being (67.46 +/- 10.49), (226.7 +/- 71.59), (177.3 +/- 35.85), and (18.33 +/- 2.53) micromol x L(-1), respectively, and V(max) values being (14.74 +/- 0.97), (5.91 +/- 1.03), (38.14 +/- 3.60), and (1.22 +/- 0.05) micromol x min(-1) x g(-1) (protein), respectively. The results showed that the activity of UDP-glucuronosyltranferase (UGT) in intestinal microsomes was the highest among the four organs, and the activities of UGT were higher than that of glucuronidase (GUS) in hepatic, renal and intestinal microsomes, but the activity of GUS was higher than that of UGT in bladder homogenates.
Animals ; Anti-Infective Agents ; pharmacokinetics ; Antioxidants ; pharmacokinetics ; Biotransformation ; Flavanones ; pharmacokinetics ; Flavonoids ; pharmacokinetics ; Glucuronidase ; metabolism ; Glucuronosyltransferase ; metabolism ; Hydrolysis ; Intestines ; enzymology ; metabolism ; Kidney ; enzymology ; metabolism ; Liver ; enzymology ; metabolism ; Male ; Microsomes ; enzymology ; metabolism ; Rats ; Rats, Sprague-Dawley ; Urinary Bladder ; enzymology ; metabolism

OBJECTIVETo investigate the effect of hTERT antisense oligodeoxynucleotide on telomerase activity in bladder cancer cells.

METHODSAntisense phosphorothioate oligodeoxynucleotide (AS PS-ODN) was synthesized and purified. Telomerase activity was measured by telomerase PCR ELISA kit. hTERT mRNA expression was detected by reverse transcription-polymerase chain reaction (RT-PCR), and hTERT protein by immunohistochemistry and flow cytometry.

RESULTSTelomerase activity was decreased in T24 cells 48 h after treatment with AS PS-ODN, and was significantly inhibited at 72 h.

CONCLUSIONAS PS-ODN can significantly inhibit telomerase activity by down-regulating hTERT mRNA and protein expression in bladder cancer cells.

Cell Line, Tumor ; Gene Expression Regulation, Neoplastic ; Humans ; Oligodeoxyribonucleotides, Antisense ; pharmacology ; RNA, Messenger ; genetics ; metabolism ; Telomerase ; genetics ; metabolism ; Urinary Bladder Neoplasms ; enzymology ; pathology

Ischemia/reperfusion (I/R) injury-induced oxidative stress plays an important role in the functional impairment of the bladder following acute bladder outlet obstruction (BOO) via induction of apoptosis. The purpose of this study was to investigate the time course of the bladder apoptosis, and apoptosis related molecular changes in the early stage of acute BOO. Twelve-week-old male Sprague Dawley rats were divided into control, acute BOO only (I), and acute BOO plus subsequent emptying (I/R) for 30, 60, 120 min, 3 days and 2 weeks. We examined the extent of bladder apoptosis, expression of Mn-superoxide dismutase (Mn-SOD), Bcl-2, Bax, caspase 3 and poly (ADP-ribose) (PAR) in the bladder. Bladder apoptosis was significantly increased in the I/R group at 30, 60, and 120 min following bladder emptying. BOO plus subsequent emptying for 30, 60, 120 min showed significant decrease in MnSOD and Bcl-2 expression, and significant increase in caspase 3, Bax expression, and amounts of PAR. These results indicate that bladder apoptosis, induced by acute BOO and subsequent emptying, is associated with decreased MnSOD expression, increased PARP activity and imbalance in apoptosis pathways.
Animals ; *Apoptosis ; Caspase 3/metabolism ; Male ; Oxidative Stress ; Poly(ADP-ribose) Polymerases/metabolism ; Proto-Oncogene Proteins c-bcl-2/*metabolism ; Rats ; Rats, Sprague-Dawley ; Superoxide Dismutase/*metabolism ; Time Factors ; Urinary Bladder/enzymology/metabolism ; Urinary Bladder Neck Obstruction/enzymology/*metabolism ; bcl-2-Associated X Protein/*metabolism

OBJECTIVETo investgate the metabolism of terephthalic acid (TPA) in rats and its mechanism. Methods Metabolism was evaluated by incubating sodium terephthalate (NaTPA) with rat normal liver microsomes, or with microsomes pretreated by phenobarbital sodium, or with 3-methycholanthrene, or with diet control following a NADPH-generating system. The determination was performed by high performance liquid chromatography (HPLC), and the mutagenic activation was analyzed by umu tester strain Salmonella typhimurium NM2009. Expression of CYP4B1 mRNA was detected by RT-PCR. Results The amount of NaTPA (12.5-200 micromol x L(-1)) detected by HPLC did not decrease in microsomes induced by NADPH-generating system. Incubation of TPA (0.025-0.1 mmol x L(-1)) with induced or noninduced liver microsomes in an NM2009 umu response system did not show any mutagenic activation. TPA exposure increased the expression of CYP4B 1 mRNA in rat liver, kidney, and bladder.

CONCLUSIONLack of metabolism of TPA in liver and negative genotoxic data from NM2009 study are consistent with other previous short-term tests, suggesting that the carcinogenesis in TPA feeding animals is not directly interfered with TPA itself and/or its metabolites.

Animals ; Aryl Hydrocarbon Hydroxylases ; genetics ; metabolism ; Gene Expression Regulation, Enzymologic ; drug effects ; Genes, Bacterial ; genetics ; Kidney ; enzymology ; Liver ; enzymology ; Male ; Microsomes, Liver ; drug effects ; enzymology ; Mutagenicity Tests ; Phthalic Acids ; pharmacokinetics ; toxicity ; RNA, Messenger ; metabolism ; Rats ; Rats, Sprague-Dawley ; Salmonella typhimurium ; genetics ; Urinary Bladder ; enzymology ; beta-Galactosidase ; metabolism

Animals ; Carcinoma, Squamous Cell ; genetics ; Colorectal Neoplasms ; genetics ; metabolism ; Enzyme Activation ; Esophageal Neoplasms ; genetics ; Genome-Wide Association Study ; Head and Neck Neoplasms ; genetics ; Humans ; Neoplasms ; chemically induced ; enzymology ; genetics ; Phosphoinositide Phospholipase C ; chemistry ; genetics ; metabolism ; physiology ; Signal Transduction ; Skin Neoplasms ; chemically induced ; enzymology ; Stomach Neoplasms ; genetics ; Urinary Bladder Neoplasms ; metabolism ; pathology ; ras Proteins ; metabolism

OBJECTIVETo investigate the relationship of protein kinase C-alpha (PKCalpha) expression/activation with tumor differentiation and resistance to chemotherapy drugs in superficial bladder carcinoma.

METHODSExpression of PKCalpha was measured by Western-blot analysis in 76 samples including tumor and normal tissues, respectively. A human RT4 bladder cancer cell line stably expressing green fluorescent protein (GFP)-PKCalpha (RT4/PKCalpha) was established. The sensitivity of the RT4/PKCalpha and parental cells to adriamycin (ADM) was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The change of sensitivity of the RT4/PKCalpha to ADM were observed under the conditions of PKC activation and inhibition, respectively.

RESULTSTotal level of PKCalpha expression and the ratio of the amount of PKCalpha expression or PKC activity in membrane to that in cytosol (M/C) were all more higher in cancerous tissues than in normal tissues (P < 0.01); With the increase of tumor grade, the relative level of PKCalpha expression significantly increased in membrane (P < 0.01) and decreased in cytosol (P < 0.01), M/C of PKCalpha was significantly elevated (P < 0.01), and total relative level of PKCalpha expression significantly increased (P < 0.01). Thirty-eight cases recurred during the follow-up period in total seventy cases. Multivariate analysis showed that high M/C of PKCalpha was independent prognostic factor for tumor recurrence after standard ADM treatment in the 2-year follow-up (RR = 3.98, 95% CI 1.22-5.68, P = 0.03). Transfection of PKCalpha increased resistance of RT4 cells to ADM [resistance index (RI): 6.97, t = 3.24, P < 0.01]. PKCalpha activation further greatly promoted the resistance (RI: 148.11, t = 5.18, P < 0.001) while inhibition of PKCalpha did conversely (RI: 1.6, t = 1.29, P > 0.05).

CONCLUSIONThe abnormal activation and expression level of PKCalpha closely correlate with both tumor grade and intrinsic resistance to ADM in patients with superficial bladder carcinoma.

Aged ; Antibiotics, Antineoplastic ; pharmacology ; Carcinoma, Transitional Cell ; enzymology ; pathology ; Doxorubicin ; pharmacology ; Drug Resistance, Neoplasm ; physiology ; Enzyme Activation ; Female ; Follow-Up Studies ; Humans ; Male ; Middle Aged ; Neoplasm Staging ; Protein Kinase C-alpha ; metabolism ; Transfection ; Tumor Cells, Cultured ; Urinary Bladder Neoplasms ; enzymology ; pathology