Biomedical subjects

Yuan Wei

Publications and source records attributed to Yuan Wei.

30 records · Page 2Linked to original sources

PubMed2004

Arachidonic acid inhibits epithelial Na channel via cytochrome P450 (CYP) epoxygenase-dependent metabolic pathways.

We used the patch-clamp technique to study the effect of arachidonic acid (AA) on epithelial Na channels (ENaC) in the rat cortical collecting duct (CCD). Application of 10 microM AA decreased the ENaC activity defined by NPo from 1.0 to 0.1. The dose-response curve of the AA effect on ENaC shows that 2 microM AA inhibited the ENaC activity by 50%. The effect of AA on ENaC is specific because neither 5,8,11,14-eicosatetraynoic acid (ETYA), a nonmetabolized analogue of AA, nor 11,14,17-eicosatrienoic acid mimicked the inhibitory effect of AA on ENaC. Moreover, inhibition of either cyclooxygenase (COX) with indomethacin or cytochrome P450 (CYP) omega-hydroxylation with N-methylsulfonyl-12,12-dibromododec-11-enamide (DDMS) failed to abolish the effect of AA on ENaC. In contrast, the inhibitory effect of AA on ENaC was absent in the presence of N-methylsulfonyl-6-(propargyloxyphenyl)hexanamide (MS-PPOH), an agent that inhibits CYP-epoxygenase activity. The notion that the inhibitory effect of AA is mediated by CYP-epoxygenase-dependent metabolites is also supported by the observation that application of 200 nM 11,12-epoxyeicosatrienoic acid (EET) inhibited ENaC in the CCD. In contrast, addition of 5,6-, 8,9-, or 14,15-EET failed to decrease ENaC activity. Also, application of 11,12-EET can still reduce ENaC activity in the presence of MS-PPOH, suggesting that 11,12-EET is a mediator for the AA-induced inhibition of ENaC. Furthermore, gas chromatography mass spectrometry analysis detected the presence of 11,12-EET in the CCD and CYP2C23 is expressed in the principal cells of the CCD. We conclude that AA inhibits ENaC activity in the CCD and that the effect of AA is mediated by a CYP-epoxygenase-dependent metabolite, 11,12-EET.

Animals↗

PubMed2004

Dietary K intake regulates the response of apical K channels to adenosine in the thick ascending limb.

We used the patch-clamp technique to study the effect of adenosine on the apical 70-pS K channel in the thick ascending limb (TAL) of the rat kidney. Application of 1 microM cyclohexyladenosine (CHA), an adenosine analog, stimulated apical 70-pS K channel activity and increased the product of channel open probability and channel number (NP(o)) from 0.34 to 0.7. Also, addition of CGS-21680, a specific A(2a) adenosine receptor agonist, mimicked the effect of CHA and increased NP(o) from 0.33 to 0.77. The stimulatory effect of CHA and CGS-21680 was completely blocked by H89, an inhibitor of protein kinase A (PKA), or by inhibition of adenylate cyclase with SQ-22536. This suggests that the stimulatory effect of adenosine analogs is mediated by a PKA-dependent pathway. The effect of adenosine analog was almost absent in the TAL from rats on a K-deficient (KD) diet for 7 days. Application of DDMS, an agent that inhibits cytochrome P-450 hydrolase, not only significantly increased the activity of the 70-pS K channel but also restored the stimulatory effect of CHA on the 70-pS K channel in the TAL from rats on a KD diet. Also, the effect of CHA was absent in the presence of 20-HETE. Inhibition of PKA blocked the stimulatory effect of CHA on the apical 70-pS K channel in the presence of DDMS in the TAL from rats on a KD diet. We conclude that stimulation of adenosine receptor increases the apical 70-pS K channel activity via a PKA-dependent pathway and that the effect of adenosine on the apical 70-pS K channel is suppressed by low-K intake. Moreover, the diminished response to adenosine is the result of increase in 20-HETE formation, which inhibits the cAMP-dependent pathway in the TAL from rats on a KD diet.

Adenosine↗

PubMed2004

Vascular CO counterbalances the sensitizing influence of 20-HETE on agonist-induced vasoconstriction.

We examined the influence of interactions between CO and 20-hydroxyeicosatetraenoic acid (20-HETE) on vascular reactivity to phenylephrine and vasopressin. Renal interlobar arteries incubated in Krebs buffer released CO at a rate that is decreased (from 125.0+/-15.2 to 46.3+/-8.8 pmol/mg protein per hour, P<0.05) by the heme oxygenase inhibitor chromium mesoporphyrin (CrMP; 30 micromol/L). The level of 20-HETE in vessels was not affected by CrMP (74.3+/-6.1 versus 72.5+/-16.2 pmol/mg protein), but was decreased (P<0.05) by CO (1 micromol/L; 33.2+/-7.9 pmol/mg protein) or the cytochrome P450-4A inhibitor N-methylsulfonyl-12,12-dibromododec-11-enamide (DDMS; 30 micromol/L; 11.4+/-3.3 pmol/mg protein). Phenylephrine elicited development of isometric tension in vascular rings mounted on a wire-myograph (EC(50), 0.29+/-0.02 micromol/L; R(max), 3.78+/-0.19 mN/mm). The sensitivity to phenylephrine was decreased (P<0.05) by CO (1 micromol/L; EC(50), 0.60+/-0.04 micromol/L) or DDMS (EC(50), 0.71+/-0.12 micromol/L) and increased (P<0.05) by 20-HETE (10 micromol/L; EC(50), 0.08+/-0.02 micromol/L) or CrMP (EC(50), 0.11+/-0.02 micromol/L). Notably, neither CO nor CrMP changed the sensitivity to phenylephrine in vessels treated with DDMS. Refractoriness to CO and CrMP in such a setting was eliminated by inclusion of 20-HETE (1 micromol/L) in the bathing buffer. The aforementioned interventions affected the vascular reactivity to vasopressin in a similar manner. These data indicate that the reactivity of renal arteries to phenylephrine and vasopressin is reciprocally influenced by CO and 20-HETE of vascular origin and that CO desensitizes the vascular smooth muscle to constrictor agonists by interfering with the sensitizing influence of 20-HETE.

Adrenergic Agonists↗

PubMed2004

Dual effect of insulin-like growth factor on the apical 70-pS K channel in the thick ascending limb of rat kidney.

We used the patch-clamp technique to study the effect of insulin-like growth factor I (IGF-I) on the apical 70-pS K channel in the isolated thick ascending limb (TAL) of the rat kidney. The isolated TAL was cut open to gain access to the apical membrane. Addition of 25 nM IGF-I stimulates the apical 70-pS K channel and increases channel activity, defined by the product of channel open probability and channel number, from 0.31 to 1.21. The stimulatory effect of IGF-I is not mediated by nitric oxide- or protein tyrosine phosphatase-dependent mechanisms, because inhibition of nitric oxide synthase or blocking protein tyrosine phosphatase did not abolish the stimulatory effect of IGF-I on the 70-pS K channel. In contrast, inhibition of mitogen-activated protein (MAP) kinase with PD-98059 or U0126 abolished the stimulatory effect of IGF-I. This suggests that MAP kinase is responsible for mediating the effect of IGF-I on the apical K channels. Moreover, the effect of IGF-I on the apical 70-pS K channel is biphasic because high concentrations (>200 nM) inhibit apical 70-pS K channels. Application of 400 nM IGF-I decreased channel activity from 1.45 to 0.2. The inhibitory effect of IGF-I is not blocked by calphostin C (an inhibitor of PKC), but inhibition of protein tyrosine kinase with herbimycin A abolished the IGF-induced inhibition. We conclude that IGF-I has a dual effect on the apical 70-pS K channel in the TAL: low concentrations of IGF-I stimulate, whereas high concentrations inhibit the channel activity. The stimulatory effect of IGF-I is mediated by a MAP kinase-dependent pathway, whereas the inhibitory effect is the result of stimulation of protein tyrosine kinase.

Animals↗

PubMed2004

PKC expression is regulated by dietary K intake and mediates internalization of SK channels in the CCD.

We have used Western blot analysis and immunocytochemistry to determine the effect of dietary K intake on the expression of protein kinase C (PKC) isoforms in the kidney. Western blot has demonstrated that conventional PKC isoforms (alpha and beta), novel PKC isoforms (delta, epsilon, and eta), and atypical PKC isoforms (zeta) are expressed in the renal cortex and outer medulla. Moreover, a low K intake significantly increases the expression of PKC-epsilon in the renal cortex and outer medulla but does not change the expression of PKC-alpha, PKC-beta, PKC-delta, PKC-eta, and PKC-zeta. Also, immunocytochemistry shows that PKC-epsilon isoform is expressed in the cortical collecting duct (CCD) and outer medullary collecting duct (OMCD) and that the intensity of PKC-epsilon staining is higher in the kidney from rats on a K-deficient diet than those on a control diet. Also, we used the patch-clamp technique to study the role of PKC in mediating internalization of ROMK (Kir 1.1)-like small-conductance K (SK) channels induced by phenylarsine oxide (PAO), an agent that inhibits protein tyrosine phosphatase and has been shown to stimulate the internalization of the SK channel in the CCD (Sterling H, Lin DH, Qu RM, Dong K, Herbert SC, and Wang WH. J Biol Chem 277: 4317-4323, 2002). Inhibition of PKC with calphostin C and GF-109203x had no significant effect on channel activity but abolished the inhibitory effect of PAO on SK channels. In conclusion, a low K intake increases the expression of PKC-epsilon isoform in the renal cortex and outer medulla, and PKC is involved in mediating the internalization of SK channels in the CCD induced by stimulation of protein tyrosine kinase activity.

Algorithms↗

PubMed2004

Serial monitoring of BCR-ABL transcripts in chronic myelogenous leukemia (CML) treated with imatinib mesylate.

Survival among chronic myelogenous leukemia (CML) patients can be linked to the reduction in leukemic cell burden. Treatment with imatinib mesylate results in a high frequency of complete cytogenetic response, which can be further stratified using quantitative reverse-transcriptase polymerase chain reaction (qRT-PCR). We have serially monitored peripheral blood and bone marrow BCR-ABL transcripts using qRT-PCR in CML patients commencing imatinib therapy, and compared the results with bone marrow cytogenetics. Seventeen patients (aged 25-74 yr) with Philadelphia chromosome positive CML in first chronic phase were treated with imatinib targeting a dose of 400 mg/d. The median follow up is 30 mo (range 9-33 mo). Every third month the product of the BCR-ABL fusion gene was evaluated in both blood and bone marrow specimens by real-time RT-PCR using the TaqMan probe system. In 113 simultaneously obtained blood and bone marrow samples, the BCR-ABL transcript values agreed well with cytogenetic data. Blood and bone marrow specimens gave comparable values for BCR-ABL transcripts. Before start of imatinib therapy there was a considerable variation in BCR-ABL transcripts among the patients, ranging approximately one log (base 10). Similarly, patients with a complete cytogenetic response following imatinib therapy had variable BCR-ABL transcript levels, ranging at least three logs (base 10). The major decline in BCR-ABL transcripts occurred within 6 mo after start of imatinib therapy. The decline in BCR-ABL transcripts, following imatinib therapy, appears to level off at 12-15 mo. Two late responders were identified with a still decreasing level in BCR-ABL transcripts after 24 mo of treatment. It is concluded that BCR-ABL mRNA quantification in peripheral blood is suitable for routine monitoring of the response to treatment and long-term disease status in CML, especially in patients who have achieved a complete cytogenetic response. A plateau in BCR-ABL transcripts seems to have been reached after 12-15 mo of imatinib treatment; however, some "late responders" are seen.

Adult↗

PubMed2003

Acute application of TNF stimulates apical 70-pS K+ channels in the thick ascending limb of rat kidney.

TNF has been shown to be synthesized by the medullary thick ascending limb (mTAL) (21). In the present study, we used the patch-clamp technique to study the acute effect of TNF on the apical 70-pS K+ channel in the mTAL. Addition of TNF (10 nM) significantly stimulated activity of the 70-pS K+ channel and increased NPo [a product of channel open probability (Po) and channel number (N)] from 0.20 to 0.97. The stimulatory effect of TNF was observed only in cell-attached patches but not in excised patches. Moreover, addition of TNF had no effect on the ROMK-like small-conductance K+ channels in the TAL. The dose-response curve of the TNF effect yielded a Km value of 1 nM, a concentration that increased channel activity to 50% maximal stimulatory effect of TNF. The concentrations required for reaching the plateau of the TNF effect were between 5 and 10 nM. The stimulatory effect of TNF on the 70-pS K+ channel was observed in the presence of N(omega)-nitro-L-arginine methyl ester. This indicated that the effect of TNF was not mediated by a nitric oxide-dependent pathway. Also, inhibition of PKA did not affect the stimulatory effect of TNF. In contrast, inhibition of protein tyrosine kinase not only increased activity of the 70-pS K+ channel but also abolished the effect of TNF. Moreover, inhibition of protein tyrosine phosphatase (PTP) blocked the stimulatory effect of TNF on the 70-pS K+ channel. The notion that the TNF effect results from stimulation of PTP activity is supported by PTP activity assay in which treatment of mTAL cells with TNF significantly increased the activity of PTP. We conclude that TNF stimulates the 70-pS K+ channel via stimulation of PTP in the mTAL.

Animals↗

PubMed2002

Tetanus toxin abolishes exocytosis of ROMK1 induced by inhibition of protein tyrosine kinase.

We used confocal microscopy, patch-clamp, and biotin-labeling techniques to examine the role of soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins in mediating the effect of inhibition of PTK on ROMK1 trafficking in HEK-293 cells transfected with c-Src and green fluorescent protein (GFP)-ROMK1. Inhibition of c-Src with herbimycin A significantly decreased the tyrosine phosphorylation level of ROMK1. Patch-clamp studies demonstrated that addition of herbimycin A increased the activity of ROMK1 in cell-attached patches. Confocal microscopic imaging showed that herbimycin A decreased the intracellular intensity of GFP-ROMK1. The biotin-labeling technique demonstrated that the inhibition of c-Src increased surface ROMK1 by 110%. In contrast, inhibition of c-Src did not increase the K channel number in HEK cells transfected with R1Y337A, a ROMK1 mutant in which tyrosine residue 337 was mutated to alanine. This suggests that tyrosine residue 337 is essential for the herbimycin A-induced increase in surface ROMK1 channels. To determine whether SNARE proteins are involved in mediating exocytosis of ROMK1 induced by the inhibition of c-Src, we examined the effect of herbimycin A on ROMK1 trafficking in cells treated with tetanus toxin. The incubation of cells in a medium containing tetanus toxin abolished the herbimycin A-induced increase in the number of surface ROMK1. In contrast, inhibition of c-Src still increased the numbers of surface ROMK1 in cells treated with boiled tetanus toxin. We conclude that tyrosine dephosphorylation enhances the exocytosis of ROMK1 and that SNARE proteins are required for exocytosis induced by inhibition of PTK.

Amino Acid Substitution↗

PubMed2002

Angiotensin II stimulates basolateral K channels in rat cortical collecting ducts.

We used the patch-clamp technique to study the effects of angiotensin II (ANG II) on basolateral K channels in cortical collecting ducts (CCDs). Application of ANG II (100 pM-100 nM) increased the activity of basolateral 18-pS K channels. This effect of ANG II was completely abolished by losartan, which is an antagonist of type 1 angiotensin (AT(1)) receptors. In contrast, inhibition of type 2 angiotensin (AT(2)) receptors did not block the stimulatory effect of ANG II. Also, application of ANG II significantly increased intracellular Ca(2+) concentrations, which were measured with fura 2 dye. To explore the role of Ca(2+)-dependent pathways in the regulation of basolateral K channels, the effects of ANG II on channel activity were examined in the presence of arachidonyltrifluoromethyl ketone to inhibit phospholipase A(2) (PLA(2)), GF-109203X [a protein kinase C (PKC) inhibitor], and N(G)-nitro-l-arginine methyl ester (l-NAME) to inhibit nitric oxide synthase. Inhibition of either PLA(2) or PKC did not block the effect of ANG II on basolateral K-channel activity. However, the stimulatory effect of ANG II was absent in the CCDs treated with l-NAME. Moreover, addition of the membrane-permeant 8-bromo-guanosine 3',5'-cyclic monophosphate (8-bromo-cGMP) not only increased channel activity but also abolished the stimulatory effect of ANG II on channel activity. We conclude that ANG II increases basolateral K-channel activity via the stimulation of AT(1) receptors, and the stimulatory effect of ANG II is mediated by a nitric oxide-dependent cGMP pathway.

Angiotensin II↗

PubMed2002

Reference value of semen quality in Chinese young men.

The purpose of the study was to evaluate semen quality in young Chinese men and to establish reference values. Normal healthy young men from seven geographical areas were enrolled. The study showed that the mean sperm volume was 2.61 mL, and mean percent of sperm with forward progression was 59.89, while median of semen viability was 79.0%, and geometric mean of semen density was 55.45 x 10(6)/mL. Proportion of routine semen indexes that met World Health Organization (WHO) criteria were as follows: 81.9% for semen volume, 91.1% for liquefaction time, 93.4% for viscosity, 90.8% for pH, 81.3% for sperm with forward progression, 65.3% for sperm viability, 93.8% for semen density, 98.8% for normal sperm morphology, and 89.1% for total sperm count. Participants whose sperm met all WHO standard parameters accounted for 42.3%. Because the infertility rate in China is about 10-15%, the fifteenth percentile of semen parameters might be used as the lower limit of reference values, which may be more appropriate for young Chinese men. The fifteenth percentiles of parameters in this study were as follows: 1.5 mL for semen volume, 7.2 for pH value, 45% for proportion of sperm with forward progression, 68% for sperm viability, 30 x 10(6)/mL for semen density, 68% for proportion of sperm with normal morphology, and 50 x 10(6) for total sperm count.

Adult↗

PubMed2002

K depletion enhances the extracellular Ca2+-induced inhibition of the apical K channels in the mTAL of rat kidney.

We have shown previously that raising extracellular Ca(2)+ inhibited the apical 70-pS K channel in the thick ascending limb (TAL; Wang, W.H., M. Lu, and S.C. Hebert. 1996. Am. J. Physiol. 270:C103-C111). We now used the patch-clamp technique to study the effect of increasing the extracellular Ca(2)+ on the 70-pS K channel in the mTAL from rats on a different K diet. Increasing the extracellular Ca(2)+ from 10 microM to 0.5, 1, and to 1.5 mM in the mTAL from rats on a K-deficient (KD) diet inhibited the channel activity by 30, 65, and 90%, respectively. In contrast, raising the extracellular Ca(2)+ to 1.5 mM had no significant effect on channel activity in the mTAL from animals on a high K (HK) diet and further increasing the extracellular Ca(2)+ to 2.5, 3.5, and 5.5 mM decreased the channel activity by 29, 55, and 90%, respectively. Inhibition of the cytochrome P450 monooxygenase completely abolished the effect of the extracellular Ca(2)+ on channel activity in the mTAL from rats on a different K diet. In contrast, blocking cyclooxygenase did not significantly alter the responsiveness of the 70-pS K channel to the extracellular Ca(2)+. Moreover, addition of sodium nitropruside, a nitric oxide (NO) donor, not only increased the channel activity, but also blunted the inhibitory effect of the extracellular Ca(2)+ on the 70-pS K channel and decreased 20-hydroxyeicosatetraenoic acid (20-HETE) concentration in the mTAL from rats on a KD diet. In contrast, inhibiting NOS with L-NAME enhanced the inhibitory effect of the extracellular Ca(2)+ on the channel activity and increased 20-HETE concentration in the mTAL from rats on a high K diet. Western blot has further shown that the expression of inducible NO synthase (iNOS) is significantly higher in the renal medulla from rats on an HK diet than that on a KD diet. Also, addition of S-nitroso-N-acetylpenicillamine abolished the inhibitory effect of arachidonic acid on channel activity in the mTAL, whereas it did not block the inhibitory effect of 20-HETE. We conclude that a low dietary K intake increases the sensitivity of the 70-pS K channel to the extracellular Ca(2)+, and that a decrease in NOS activity is involved in enhancing the inhibitory effect of the extracellular Ca(2)+ on channel activity in the mTAL during K depletion.

Animals↗

PubMed2002

Role of the cytoskeleton in mediating effect of vasopressin and herbimycin A on secretory K channels in CCD.

We have previously demonstrated that inhibiting protein tyrosine kinase (PTK) and stimulating protein kinase A (PKA) increase the activity of the small-conductance K (SK) channel in the cortical collecting duct (CCD) of rat kidneys (Cassola AC, Giebisch G, and Wang WH. Am J Physiol Renal Fluid Electrolyte Physiol 264: F502-F509, 1993; Wang WH, Lerea KM, Chan M, and Giebisch G. Am J Physiol Renal Physiol 278: F165-F171, 2000). In the present study, we used the patch-clamp technique to study the role of the cytoskeleton in mediating the effect of herbimycin A, an inhibitor of PTK, and vasopressin on the SK channels in the CCD. The addition of colchicine, an inhibitor of microtubule assembly, or taxol, an agent that blocks microtubule reconstruction, had no significant effect on channel activity. However, colchicine and taxol treatment completely abolished the stimulatory effect of herbimycin A on the SK channels in the CCD. Removal of the microtubule inhibitors restored the stimulatory effect of herbimycin A. In contrast, treatment of the tubules with either taxol or colchicine did not block the stimulatory effect of vasopressin on the SK channels. Moreover, the effect of herbimycin A on the SK channels was also absent in the CCDs treated with either cytochalasin D or phalloidin. In contrast, the stimulatory effect of vasopressin was still observed in the tubules treated with phalloidin. However, cytochalasin D treatment abolished the effect of vasopressin on the SK channels. Finally, the effects of vasopressin and herbimycin A are additive because inhibiting PTK can still increase the channel activity in CCD that has been challenged by vasopressin. We conclude that an intact cytoskeleton is required for the effect on the SK channels of inhibiting PTK and that the SK channels that are activated by inhibiting PTK were differently regulated from those stimulated by vasopressin.

Actins↗