Biomedical subjects

H C Huang

Publications and source records attributed to H C Huang.

At least 19 recordsLinked to original sources

PubMed2000

Optimization of separation and migration behavior of cephalosporins in capillary zone electrophoresis.

The influences of buffer pH, buffer concentration and buffer electrolyte on the migration behavior and separation of 12 cephalosporin antibiotics in capillary zone electrophoresis using three different types of buffer electrolyte, including phosphate, citrate, and 2-(N-morpholino)ethanesulfonate (MES), were investigated. The results indicate that, although buffer pH is a crucial parameter, buffer concentration also plays an important role in the separation of cephalosporins, particularly when cefuroxime and cefazolin, cephalexin and cefaclor, or cefotaxime and cephapirin are present as analytes at the same time. The electrophoretic mobility of cephalosporins and electroosmotic mobility measured in citrate and MES buffers are remarkably different from those measured in phosphate buffer. With citrate buffer, optimum buffer concentration is confined to a small range (35-40 mM), whereas buffer concentrations up to 300 mM can be used with MES buffer. Complete separations of 12 cephalosporins could be satisfactorily achieved with these three buffers under various optimum conditions. However, the separability of 12 cephalosporins with citrate or MES buffer is better than that with phosphate buffer. As a consequence of a greater electrophoretic mobility of cephalosporins than the electroosmotic mobility with citrate buffer at pH below about 5, some cephalosporins are not detectable. The cloudiness of the peak identification and of the magnitudes of the electrophoretic mobility of cefotaxime and cefuroxime reported previously are clarified. In addition, the pKa values of cephradine, cephalexin, cefaclor, and cephapirin attributed to the deprotonation of either an amino group or a pyridinium group are reported, and the migration behavior of these cephalosporins in the pH range studied is quantitatively described.

Alkanesulfonic Acids↗

PubMed2000

Transcriptional regulation of the antioxidant response element. Activation by Nrf2 and repression by MafK.

The antioxidant response element (ARE) mediates the transcriptional activation of many genes encoding phase II drug-metabolizing enzymes in response to oxidative stress. Recent studies using knockout mice suggest that NF-E2-related factor 2 (Nrf2), along with small Maf proteins, binds and activates the ARE. In this study, using in vitro binding assays, Nrf2/MafK heterodimers were found to interact with high affinity to the ARE. However, distinct differences were observed when this interaction was compared with that formed with nuclear proteins from H4II EC3 or HepG2 cells. Overexpression of Nrf2 activated ARE-mediated transcription in HepG2 cells, and this activation was further increased by tert-butylhydroquinone. In HeLa cells, overexpression of Nrf2 resulted in activation of the ARE, but this activation was no longer induced by tert-butylhydroquinone. Using ARE constructs with point mutations in the core sequence, we found that only mutations at the T or G nucleotides within the core (TGAC) render the ARE unresponsive to Nrf2. Overexpression of MafK led to dose-dependent repression of ARE activity. Activation of the ARE by Nrf2 was similarly antagonized by MafK. These data suggest that Nrf2 plays an important role mediating basal activity of the ARE but that small Maf proteins are repressors and not activators of ARE-mediated transcription.

Animals↗

PubMed2000

Capillary zone electrophoretic separation of neutral species of chloro-s-triazines in the presence of cationic surfactant monomers.

Chloro-s-triazines are difficult to separate by capillary zone electrophoresis (CZE), due to their low pKa values. However, these analytes can be effectively separated by CZE in the presence of cationic surfactant monomers, such as tetradecylammonium bromide (TTAB) and dodecyltrimethylammonium bromide (DTAB). The separation mechanism based on a 1:1 binding of analytes to cationic surfactant monomers is proposed. The binding constants of chloro-s-triazines to cationic surfactant monomers are estimated. The results show that the strength of the interactions of these analytes with TTAB monomers is considerably strong, whereas that of the corresponding analyte with DTAB monomers is about 12- to 14-fold weaker. A linear correlation of binding constants with log P(ow) (the logarithm of the partition coefficient of analytes between 1-octanol and aqueous phases) indicates that the migration order of these chloro-s-triazines depends primarily on their hydrophobicity. Moreover, the skewed peaks of chloro-s-triazines observed may reveal the occurrence of adsolubilization of these analytes in the adsorbed cationic surfactant layer on the capillary surface.

Cations↗

PubMed2000

The decline of porcine sperm motility by geldanamycin, a specific inhibitor of heat-shock protein 90 (HSP90).

Sperm motility is an important parameter for fertility. The molecular mechanisms of mammalian sperm motility are still largely undefined. Our previous observations suggested that heat shock protein 90 (HSP90) may be associated with porcine sperm motility. The aim of the present study was to further characterize the plausible novel function of HSP90 on sperm motility. Semen from normal, sexually mature boars with sperm motility higher than 80% was used. An HSP90-specific inhibitor, geldanamycin (GA), was added to diluted semen at 0.5, 1.0, 2.5 or 5.0 microg/mL and the semen was then incubated at 37 degrees C for 15, 30, 45 or 60 min. Sperm motility was determined by using computer-assisted semen analyzer at the end of incubation. The results indicated that GA significantly reduced sperm motility in a dose and time dependent manner. Moreover, incubation of semen with 5.0 microg/mL GA for 15 min completely stopped sperm motility. To test the reversibility of the GA effect on sperm motility, GA was removed after 30 min incubation and was replaced with fresh extender alone or with extender plus 5 mM caffeine, then incubated for another 15, 30, 45 or 60 min. The results showed that simply removing GA did not reverse the inhibitory effect on sperm motility, while adding caffeine partially reversed this inhibitory effect. However, the effect of 2.5 or 5.0 microg/mL GA was not reversed by caffeine. Considering the specificity of GA targeting to HSP90, the above observations suggested that HSP90 may play a crucial role in regulating porcine sperm motility.

Animals↗

PubMed2000

The gene coding for the Hrp pilus structural protein is required for type III secretion of Hrp and Avr proteins in Pseudomonas syringae pv. tomato.

Bacterial surface appendages called pili often are associated with DNA and/or protein transfer between cells. The exact function of pili in the transfer process is not understood and is a matter of considerable debate. The Hrp pilus is assembled by the Hrp type III protein secretion system of Pseudomonas syringae pv. tomato (Pst) strain DC3000. In this study, we show that the hrpA gene, which encodes the major subunit of the Hrp pilus, is required for secretion of putative virulence proteins, such as HrpW and AvrPto. In addition, the hrpA gene is required for full expression of genes that encode regulatory, secretion, and effector proteins of the type III secretion system. hrpA-mediated gene regulation apparently is through effect on the mRNA level of two previously characterized regulatory genes, hrpR and hrpS. Ectopic expression of the hrpRS gene operon restored gene expression, but not protein secretion, in the hrpA mutant. Three single amino acid mutations at the HrpA carboxyl terminus were identified that affect the secretion or regulatory function of the HrpA protein. These results define an essential role of the Hrp pilus structural gene in protein secretion and coordinate regulation of the type III secretion system in Pst DC3000.

Amino Acids↗

PubMed2000

A novel class of apical sodium co-dependent bile acid transporter inhibitors: the 2,3-disubstituted-4-phenylquinolines.

A series of 2,3-disubstituted-4-phenylquinolines were prepared and were found to inhibit the apical sodium co-dependent bile acid transporter (ASBT). Alkyl and ester substitution at the 3-position showed comparable activities while substitution at the 2-position was much more sensitive to the nature of the substituent. The synthesis and in vitro potency data are presented for this novel class of compounds.

Carrier Proteins↗

PubMed2000

G1 kinases and transforming growth factor-beta signaling are associated with a growth pattern switch in diabetes-induced renal growth.

BACKGROUND: Diabetes mellitus-induced nephromegaly is thought to involve both hyperplastic and hypertrophic proximal tubule cell growth. The temporal relationship between these growth patterns and the mechanisms that mediate them are unknown. METHODS: Renal growth was assayed in isolated renal proximal tubules harvested from diabetic rats. Diabetes mellitus was induced by streptozotocin. RESULTS: Following the induction of a diabetic state, there was a progressive increase in the kidney:body weight ratio. This was associated with an increase in 5-bromo-2-deoxyuridine incorporation (marker for hyperplastic cell growth) at day 2, which returned to baselines levels by day 4, and an increase in the protein:DNA ratio (marker for hypertrophic cell growth), which was clearly evident by day 10. Thus, diabetes-induced proximal tubule growth involved an initial hyperplastic, followed by a hypertrophic, growth period. During the hyperplastic growth period, both cdk4/cyclin D (cyclin D) and cdk2/cyclin E (cyclin E) kinase activities were increased. The switch between the growth periods was associated with continued activation of cyclin D, but inhibition of cyclin E kinase. The reduction in cyclin E kinase activity correlated with a reduction in cdk2/cyclin E complex abundance and an increased abundance of cyclin kinase inhibitors in cdk2/cyclin E complexes that did form. Also associated with the switch in growth patterns was a change in transforming growth factor-beta (TGF-beta) receptor expression. During the hyperplastic growth period, TGF-beta receptor II expression was decreased, while during the hypertrophic growth period, there was both a return of receptor II expression to baseline levels and increased expression of receptor I. Consistent with an increase in TGF-beta signaling during hypertrophy, there was an increase in Smad 2/3 protein expression and an increase in the abundance of Smad 2/4 complexes. CONCLUSIONS: Diabetes-induced proximal tubule growth involves an initial hyperplastic growth period that switches to a hypertrophic growth period within a couple of days. The pattern of G1 kinase activity associated with the growth pattern switch demonstrates that the hypertrophy is mediated by a cell cycle-dependent mechanism. Regulation of TGF-beta receptor expression and signaling activity through the Smad protein cascade possibly plays a role in the growth pattern switch.

Animals↗

PubMed2000

Profiles of inflammatory cytokines in bronchoalveolar lavage fluid from premature infants with respiratory distress disease.

In view of cytokine's effects in promoting or inhibiting inflammation, the objective of this study was to explore the characteristics of the proinflammatory cytokine, interleukin-8 (IL-8), and the inhibitory cytokine, interleukin-10 (IL-10), in the bronchoalveolar lavage (BAL) fluid of premature infants suffering from respiratory distress disease. Eighteen premature neonates with respiratory distress disease with gestational age (GA) ranging from 24 to 37 weeks were recruited for study. BAL fluids were collected following endotracheal intubation during an episode of hypoxemia or dyspnea. A series of BAL samples were obtained on day 1, 2, 4 and 7 after intubation for measuring IL-8 and IL-10 levels. The results indicate that premature infants with GA ranging from 24 to 32 weeks had a higher level of IL-8 (p = 0.029), but not level of IL-10 (p = 0.109), in the BAL obtained during the first intubation compared to premature infants with GA ranging from 33 to 37 weeks. The administration of exogenous surfactant did not influence the profiles of IL-8 and IL-10, as compared to those in-patients without treatment. Levels of IL-8 were correlated with IL-10 levels (r = 0.613, p = 0.007) in BAL fluid samples obtained on the day of intubation. The level of IL-8, but not IL-10, was significantly correlated with the duration of intubation. IL-8 and IL-10 levels in BAL fluid samples collected on the day of intubation were correlated with the development of chronic lung disease (CLD). The results suggest that extreme prematurity tends to have increased IL-8 and IL-10 levels in BAL fluid compared to premature infants with older GA, and that these increased levels are associated with the development of CLD.

Bronchoalveolar Lavage Fluid↗

PubMed1999

Primary oral squamous cell carcinoma: an analysis of 703 cases in southern Taiwan.

We retrospectively analyzed the records of 703 cases of oral squamous cell carcinoma (SCC) collected from 1 January 1985 to 31 December 1996 at a teaching hospital in southern Taiwan, to identify the characteristics of patients and factors associated with survival. There was an overwhelming male predominance (male:female = 15:1). The mean age of the patients was 52. The peak age of oral SCC patients declined from 50 to 59 years in the first six years (1985-1990) and 40-49 years in the last six years (1991-1996). The most common site of oral SCC was the buccal mucosa with 263 patients (37.4%). Most patients (346/703 patients; 49.2%) had stage III cancer. The most common site of occurrence of SCC was the buccal mucosa (263/703 patients; 37.4%), both overall and in patients who chewed betel quid alone or in combination with cigarette smoking and/or alcohol consumption; the tongue was the most common site among patients without any oral habits (18/48 patients; 37.5%). Furthermore, the age of occurrence was on average 6-12 years younger among patients who chewed betel quid than in those who did not. Of the 703 patients, 496 received treatment with surgery, chemotherapy, and/or radiation therapy. Of these, 209 (42.1%) died. The cancer stage significantly influenced mortality: the 5-year survival rate in patients treated from 1985 to 1991 was 72% in those with stage I, 38.9% in those with stage II, 26.7% in those with stage III, and 11.8% in those with stage IV cancer. Six variables were found to significantly affect survival: tumor size, lymph node involvement, surgery, betel quid chewing, staging, and histological differentiation (all p < 0.05, Kaplan-Meier analysis with log rank test). Of these, surgery and cancer stage independently affected survival in a proportional hazards model (both p < 0.0001). Therefore, the early surgical intervention, and the withdrawal from oral habits, especially betel quid chewing, will be advantageous to patients' survival.

Adolescent↗

PubMed1999

Cellular locations of Pseudomonas syringae pv. syringae HrcC and HrcJ proteins, required for harpin secretion via the type III pathway.

The complete hrp-hrc-hrmA cluster of Pseudomonas syringae pv. syringae 61 encodes 28 polypeptides. A saprophytic bacterium carrying this cluster is capable of secreting HrpZ-a harpin encoded by hrpZ-in an hrp-dependent manner, which suggests that this cluster contains sufficient components to assemble functional type III secretion machinery. Sequence data show that HrcJ and HrcC are putative outer membrane proteins, and nonpolar mutagenesis demonstrates they are all required for HrpZ secretion. In this study, we investigated the cellular localization of the HrcC and HrcJ proteins by Triton solubilization, sucrose-gradient isopycnic centrifugation, and immunogold labeling of the bacterial cell surface. Our results indicate that HrcC is indeed an outer membrane protein and that HrcJ is located between both membranes. Their membrane localization suggests that they might be involved in the formation of a supramolecular structure for protein secretion.

Bacterial Outer Membrane Proteins↗

PubMed1999

Predicting factors for outcome of tube thoracostomy in complicated parapneumonic effusion for empyema.

STUDY OBJECTIVES: To determine the predicting factors for outcome of tube thoracostomy in patients with complicated parapneumonic effusion (CPE) or empyema. DESIGN AND SETTINGS: Retrospective chart review over a 55-month period at a tertiary referred medical center. PATIENTS AND MEASUREMENTS: The medical charts of patients with empyema or CPE were reviewed. Data including age, gender, clinical symptoms, important underlying diseases, leukocyte count, duration of preadmission symptoms, interval from first procedure to second procedure, the time from first procedure to discharge (recovery time), the amount of effusion drained, administration of intrapleural streptokinase, chest tube size and position, loculation of pleural effusion, and characteristics and culture results of pleural effusion were recorded and compared between groups of patients with successful and failed outcome of tube thoracostomy drainage. RESULTS: One hundred twenty-one patients were selected for study. One hundred of these patients had received tube thoracostomy drainage with 53 successful outcomes and 47 failed outcomes of chest tube drainage. Nineteen patients received decortication directly, and the other two received antibiotics alone. Univariate analysis showed that pleural effusion leukocyte count, effusion amount, and loculation of pleural effusion were significantly related to the outcome of chest tube drainage. Multiple logistic regression analysis demonstrated that loculation and pleural effusion leukocyte count < or = 6,400/uL were the only independent predicting factors related to failure of tube thoracostomy drainage. CONCLUSIONS: Loculation and pleural effusion leukocyte count < or = 6,400/microL were independent predicting factors of poor outcome of tube thoracostomy drainage. These results suggest that if the initial attempt at chest tube drainage fails, early surgical intervention should be considered in good surgical candidates with loculated empyema or pleural effusion with leukocyte count < or = 6,400/microL.

Aged↗

PubMed1999

Characterization of a putative Pseudomonas UDPglucose pyrophosphorylase.

A UDP-glucose pyrophosphorylase encoding gene was identified through functional complementation screening by using an Escherichia coli galU mutant. Sequence analysis of the gene indicated that it is most likely derived from a Pseud monas sp. The gene is located immediately upstream and transcribed in the same direction of the gor (glutathione reductase) gene and is capable of encoding a protein 30,943 daltons in size. The gene product synthesized in Escherichia coli was purified and its biochemical properties characterized. The recombinant UDP-glucose pyrophosphorylase exhibited a molecular weight of 130 kDa, suggesting a tetrameric organization of the gene product. Two mutant forms of the enzyme were identified. The activity of the mutant enzyme with a tyrosine to histidine (Y26 1H) substitution was found to be greatly reduced. On the other hand, the tyrosine to cysteine (Y84C) substitution resulted in an enzyme that functions normally at 37 degrees C but rather poorly at temperatures lower than 30 degrees C.

Amino Acid Sequence↗

PubMed1999

Noonan syndrome presented with cystic hygroma and chylothorax: case report.

A cystic hygroma was found in a full-term (39 week-old) male newborn delivered by cesarian section. Polyhydramnios was diagnosed by sonography at 21 weeks of gestation. Clinical manifestations at birth, such as respiratory distress, tachypnea and chylothorax were successfully relieved by the insertion of chest tube and medical therapy. Other conditions, which included mild hydronephrosis of left kidney, thickening of aortic and pulmonary valves with mild mitral and tricuspid valves regurgitation, subsided within six months. The cystic hygroma regressed to a webbed neck. In general, the features and clinical manifestations of the present case resembled the criteria of Noonan syndrome including various congenital heart defects; webbed neck; chest deformity; a characteristic facial appearance comprising of a broad forehead, ocular hypertelorism, antimongoloid slant of palpebral fissures, low set ears; and bilateral undescended testis etc., suggested by Noonan and other subsequent authors. The outcome of this infant was satisfactory following medical and surgical therapy (chest tube, orchiopexy) and a special dietary regimen.

Chylothorax↗

PubMed1999

Aortic regurgitation detected with Doppler echocardiography in apparently healthy adolescents.

In this study, we sought to determine the prevalence and characteristics of aortic regurgitation (AR) in apparently healthy adolescent students. A total of 315 healthy junior high school students underwent echocardiographic examination. There were 158 boys and 157 girls, with a mean +/- standard deviation age of 13.3 +/- 0.9 years (range, 13-15 yr). AR was found in six (2%) students. Five of the six (83%) with AR had minimally thickened aortic valves: three (50%) had a thickened right coronary cusp and four (67%) had a thickened noncoronary cusp, but none had a thickened left coronary cusp. The AR was mild in five (83%) students and moderate in one (17%). Two of the six students had aortic valve prolapse, both of whom had minimal thickening. One student with AR did not have any abnormal structural changes, and one had aortic root dilatation. No cardiac chamber dilatation was noted in students with AR. These findings indicate a relatively high prevalence of AR in apparently healthy adolescents.

Adolescent↗

PubMed1998

The Rana catesbeiana rcr gene encoding a cytotoxic ribonuclease. Tissue distribution, cloning, purification, cytotoxicity, and active residues for RNase activity.

Rana catesbeiana ribonuclease (RC-RNase) is a pyrimidine-guanine sequence-specific ribonuclease found in R. catesbeiana (bullfrog) oocytes. It possesses both ribonuclease activity and cytotoxicity against tumor cells. We report here for the first time the cloning of RC-RNase cDNA from liver rather than from oocytes where RC-RNase is stored. An internal fragment of cDNA was obtained by reverse transcription-PCR using deduced oligonucleotides as primers. Full-length cDNA was obtained by 5'- and 3'-RACE technique. The cDNA clone, named rcr gene, contained a 5'-untranslated region, a putative signal peptide (22 amino acids), a mature protein (111 amino acids), a 3'-untranslated region, and a polyadenylation site. The cDNA which encoded the mature protein was fused upstream with a modified pelB signal peptide DNA and inserted into pET11d for expression in Escherichia coli strain BL21(DE3). The secretory RC-RNase in the culture medium was enzymatically active and was purified to homogeneity. The recombinant RC-RNase had the same amino acid sequence, specific activity, substrate specificity, antigenicity, and cytotoxicity as that of native RC-RNase from frog oocytes. Amino acid residues His-10, Lys-35, and His-103 are involved in RC-RNase catalytic activity. Ribonucleolytic activity was involved in and may be essential for RC-RNase cytotoxicity. DNA sequence analysis showed that RC-RNase had approximately 45% identity to that of RNase superfamily genes. This indicates that RC-RNase is a distinct ribonuclease gene in the RNase superfamily.

Amino Acid Sequence↗

PubMed1998

The additive effect of sclerotherapy to patients receiving repeated endoscopic variceal ligation: a prospective, randomized trial.

Endoscopic variceal ligation (EVL) is a new technique designed to manage esophageal varices. The effect of sclerotherapy following repeated banding ligation remains unknown. Seventy-two patients with a history of esophageal variceal bleeding received regular EVL until variceal disappearance or until left with residual small varices. Subsequently, patients were randomized to receive sclerotherapy (Group 1, 37 patients) or serve as a control (Group 2, 35 patients). Group 1 received one to two sessions of low-dose sclerotherapy to achieve complete variceal disappearance. After a mean follow-up of 2 years, 4 months, recurrent esophageal varices developed in 14% of Group 1 and 43% of Group 2 (P < .02). Rebleeding was encountered in 8% of Group 1 versus 31% of Group 2 (P = .01). One case of esophageal stricture (2.7%) was encountered in Group 1. One patient in Group 1, compared with 3 patients in Group 2, died of massive variceal bleeding (P > .05). The multivariate Cox model indicated that treatment was the only factor predictive of variceal recurrence, and both Child-Pugh class and treatment were factors predictive of variceal rebleeding. The addition of low-dose sclerotherapy following repeated banding ligations proved safe and effective in the prevention of recurrence of esophageal varices and rebleeding.

Aged↗

PubMed1998

The possible mechanisms of the antiproliferative effect of fullerenol, polyhydroxylated C60, on vascular smooth muscle cells.

1. The possible mechanisms of the antiproliferative effect of polyhydroxylated fullerene (fullerenol), a novel free radical trapper, were studied in rat vascular smooth muscle cells (A7r5 cells) and compared with the effect of ascorbic acid. 2. Fullerenol-1 and ascorbic acid inhibited the proliferative responses in a number of cells, including rat aortic smooth muscle cells (A7r5 cells), human coronary artery smooth muscle cells, and human CEM lymphocytes (CEM cells) in a concentration dependent manner. 3. At the concentration range of 10(-6) to 10(-2) M, fullerenol-1 and ascorbic acid concentration-dependently inhibited the proliferative responses stimulated by serum in A7r5 cells. Fullerenol-1 was more potent than ascorbic acid. 4. The production of O2- induced by alloxan, a diabetogenic compound, was reduced by fullerenol-1 (10(-4) M) in the presence of A7r5 cells. 5. The cytosolic protein kinase C activity of A7r5 cells stimulated by phorbol ester was reduced by 10(-3) M fullerenol-1, but not ascorbic acid (10(-4)-10(-2) M) and fullerenol-1 at lower concentrations (10(-6)-10(-4) M). 6. In contrast, the membraneous protein tyrosine kinase activity of A7r5 cells stimulated by foetal calf serum was significantly reduced by fullerenol-1 (10(-6)-10(-3) M) and ascorbic acid (10(-4)-10(-2) M). Again, the inhibitory activity of fullerenol-1 was greater than that of ascorbic acid. 7. Our results demonstrate that fullerenol-1 and ascorbic acid exhibit inhibitory effects on transduction signals in addition to their antioxidative property. It is suggested that the antiproliferative effect of fullerenol-1 on vascular smooth muscle cells may partly be mediated through the inhibition of protein tyrosine kinase.

Animals↗

PubMed1998

Epigallocatechin suppression of proliferation of vascular smooth muscle cells: correlation with c-jun and JNK.

1. The mechanisms of the antiproliferative effect of epigallocatechin, one of the catechin derivatives found in green tea, in vascular smooth muscle cells were studied. The proliferative response was determined from the uptake of tritiated thymidine. 2. In the concentration range of 10(-6) to 10(-4) M, catechin, epicatechin, epigallocatechin, epicatechin gallate and epigallocatechin, epigallocatechin gallate, concentration-dependently inhibited the proliferative response stimulated by serum in rabbit cultured vascular smooth muscle cells. Catechin and epicatechin were less effective in inhibiting the serum-stimulated smooth muscle cell proliferation, indicating that the galloyl group may be important for full inhibitory activity. 3. Epigallocatechin (EGC) inhibited the proliferative responses in different cells including rat aortic smooth muscle cells (A7r5 cells), rabbit cultured aortic smooth muscle cells, human coronary artery smooth muscle cells, and human CEM lymphocytes in a concentration-dependent manner. The possible mechanisms of the antiproliferative effect of EGC were further studied in A7r5 cells. 4. The membranous protein tyrosine kinase activity stimulated by serum in A7r5 cells was significantly reduced by 10(-5) M EGC. In contrast, the cytosolic protein kinase C activity stimulated by phorbol ester was unaffected by directly incubating with EGC (10(-6)-10(-4) M). 5. We also performed Western blot analysis using the anti-phosphotyrosine monoclonal antibody PY20. EGC (10(-5) M) reduced the levels of tyrosine phosphorylated proteins with different molecular weights, indicating that EGC may inhibit the protein tyrosine kinase activity or stimulate the protein phosphatase activity. 6. Reverse transcription-polymerase chain reaction analysis of c-fos, c-jun and c-myc mRNA levels demonstrated that c-jun mRNA level after serum-stimulation was significantly reduced by 10(-5) M EGC. However, the reduction of c-fos and c-myc mRNA levels by 10(-5) M EGC did not achieve significance. 7. Western blot analysis using the antibody against JNK (c-jun N-terminal kinase) and ERK (extracellular signal-regulated kinase) demonstrated that the level of phosphorylated JNK1, but not phosphorylated ERK1 and ERK2, was reduced by 10(-5) M EGC. Direct measurement of kinase activity by immune complex kinase assay confirmed that JNK1 activity was inhibited by EGC treatment. These results demonstrate that EGC preferentially reduced the activation of JNK/SAPK (stress-activated protein kinase) signal transduction pathway. 8. It is suggested that the antiproliferative effect of epigallocatechin on vascular smooth muscle cells may partly be mediated through inhibition of protein tyrosine kinase activity, reducing c-jun mRNA expression and inhibiting JNK1 activation. Tea catechins may be useful as a template for the development of drugs to prevent the pathological changes of atherosclerosis and post-angioplasty restenosis.

Animals↗