K-685, a TRPV1 antagonist, blocks PKC-sensitized TRPV1 activation and improves the inflammatory pain in a rat complete Freund's adjuvant model.

Transient receptor potential vanilloid 1 (TRPV1) is a Ca(2+)-permeable non-selective cation channel that transmits pain signals. TRPV1 is activated by multiple stimuli such as capsaicin, acid, and heat. During inflammation, TRPV1 is reported to be sensitized by protein kinase C (PKC) in dorsal root ganglia (DRG) neurons, which leads to reduction in the threshold of the temperature for TRPV1 activation to body temperature. This sensitization is considered to contribute to chronic inflammatory pain. In a previous study, we discovered orally active 5,5-diarylpentadienamide TRPV1 antagonists. To examine the effects of our TRPV1 antagonists on PKC-sensitized TRPV1, we developed an in vitro assay system to monitor the TRPV1 sensitization by PKC. In this assay system, our TRPV1 antagonists, such as (2E,4Z)-N-[(3R)-3-hydroxy-2-oxo-1,2,3,4-tetrahydro-5-quinolyl]-5-(4-isopropoxyphenyl)-5-(4-trifluoromethylphenyl)-2,4-pentadienamide (K-685), inhibited the activation of TRPV1 sensitized by PKC. The potentiation of heat-induced inward currents by PKC was seen in rat DRG neurons, and K-685 attenuated these currents. Furthermore, K-685 reversed the thermal hyperalgesia and mechanical allodynia in a rat complete Freund's adjuvant-induced inflammatory pain model. These results therefore suggest that K-685 has a strong potential as a new analgesic drug for the treatment of inflammatory pain.

The generation of rat dorsal root ganglion cell lines to identify the target of KW-7158, a novel treatment for overactive bladder.

KW-7158 is a drug candidate for the treatment of overactive bladder. Although pharmacological studies have suggested that it suppresses afferent nerve conduction, its molecular target is unknown. We herein report the establishment of dorsal root ganglion (DRG) cell lines useful for identification of the target of this compound. First, we confirmed that the target exists in rat primary DRG by [(3)H]KW-7158 binding. To establish DRG cell lines, we used DRG from transgenic rats harboring the temperature-sensitive large T-antigen. The immortalized cells were initially screened for their expression of neuronal markers, and 72 positive clones were obtained (designated as TRD cells). Next, in order to select TRD cells expressing the target of KW-7158, we measured the binding affinity and amount of the binding sites present in each clone. Most clones expressed two binding sites, one with low affinity and one with high affinity. Differential binding of KW-7158 derivatives to each site revealed that the high affinity site is pharmacologically relevant. Therefore, we successfully identified "TRD-10" which express the largest amount of the high affinity site. These cell lines will therefore be useful tools to identify the target of KW-7158.

Trichoblastic carcinoma arising from colostomy.

Trichoblastic carcinoma is a rare skin cancer originating from hair germ cells. We report a case of an 84-year-old man who presented with a tumor on the stoma of the descending colon, which was preoperatively diagnosed as colon cancer. He underwent colectomy with adjacent skin, and the tumor was diagnosed as trichoblastic carcinoma by postoperative pathological examination. We are not aware of any similar cases published in the English literature. Therefore, we report this case because it is quite a rare condition.

Mechanism underlying the block of human Cav3.2 T-type Ca2+ channels by benidipine, a dihydropyridine Ca2+ channel blocker.

AIMS: Benidipine, a dihydropyridine Ca(2+) channel blocker, has been reported to block T-type Ca(2+) channels; however, the mechanism underlying this effect was unclear. In this study, we characterized the mechanism responsible for this blocking activity. Furthermore, the blocking activity was compared between two enantiomers of benidipine, (S, S)- and (R, R)-benidipine. MAIN METHODS: Human Ca(v)3.2 (hCa(v)3.2) T-type Ca(2+) channels stably expressed in the human embryonic kidney cell line, HEK-293, were studied in whole-cell patch-clamp recordings and Ca(2+) mobilization assay. KEY FINDINGS: In whole-cell patch-clamp recordings, benidipine blocked hCa(v)3.2 T-type Ca(2+) currents elicited by depolarization to a comparable extent as efonidipine. The block was dependent on stimulation frequency and holding potential, but not test potential. Benidipine significantly shifted the steady-state inactivation curve to the hyperpolarizing direction, but had no effect on the activation curve. Benidipine prolonged the recovery from inactivation of hCa(v)3.2 T-type Ca(2+) channels without any effect on the kinetics of activation, inactivation, or deactivation. In the Ca(2+) mobilization assay, benidipine was more potent than efonidipine in blocking Ca(2+) influx through hCa(v)3.2 T-type Ca(2+) channels. (S, S)-Benidipine was more potent than (R, R)-benidipine in blocking hCa(v)3.2 T-type Ca(2+) currents, but there was no difference in blocking the Ca(2+) influx. SIGNIFICANCE: We have characterized the blocking activity of benidipine against hCa(v)3.2 Ca(2+) channels and revealed the difference between the two enantiomers of benidipine. The blocking action of benidipine could be mediated by stabilizing hCa(v)3.2 Ca(2+) channels in an inactivated state.

Acute afferent loop necrosis after Roux-en-Y cholangiojejunostomy.

Afferent loop necrosis after Roux-en-Y cholangiojejunostomy biliary reconstruction is rare. We present the case of a 36-year-old woman with acute necrotic afferent loop obstruction. The peripheral area of the Roux-en-Y limb, including the cholangiojejunostomy portion, was twisted just proximal to the cholangiojejunostomy. Cholangiojejunostomy was completely separated due to necrosis of the Roux-en-Y jejunum. In addition to the case report, we discuss features of cholangiojejunostomy that require special attention.

Blockade of T-type voltage-dependent Ca2+ channels by benidipine, a dihydropyridine calcium channel blocker, inhibits aldosterone production in human adrenocortical cell line NCI-H295R.

Benidipine, a long-lasting dihydropyridine calcium channel blocker, is used for treatment of hypertension and angina. Benidipine exerts pleiotropic pharmacological features, such as renoprotective and cardioprotective effects. In pathophysiological conditions, the antidiuretic hormone aldosterone causes development of renal and cardiovascular diseases. In adrenal glomerulosa cells, aldosterone is produced in response to extracellular potassium, which is mainly mediated by T-type voltage-dependent Ca2+ channels. More recently, it has been demonstrated that benidipine inhibits T-type Ca2+ channels in addition to L-type Ca2+ channels. Therefore, effect of calcium channel blockers, including benidipine, on aldosterone production and T-type Ca2+ channels using human adrenocortical cell line NCI-H295R was investigated. Benidipine efficiently inhibited KCl-induced aldosterone production at low concentration (3 and 10 nM), with inhibitory activity more potent than other calcium channel blockers. Patch clamp analysis indicated that benidipine concentration-dependently inhibited T-type Ca2+ currents at 10, 100 and 1000 nM. As for examined calcium channel blockers, inhibitory activity for T-type Ca2+ currents was well correlated with aldosterone production. L-type specific calcium channel blockers calciseptine and nifedipine showed no effect in both assays. These results indicate that inhibition of T-type Ca2+ channels is responsible for inhibition of aldosterone production in NCI-H295R cells. Benidipine efficiently inhibited KCl-induced upregulation of 11-beta-hydroxylase mRNA and aldosterone synthase mRNA as well as KCl-induced Ca2+ influx, indicating it as the most likely inhibition mechanism. Benidipine partially inhibited angiotensin II-induced aldosterone production, plus showed additive effects when used in combination with the angiotensin II type I receptor blocker valsartan. Benidipine also partially inhibited angiotensin II-induced upregulation of the above mRNAs and Ca2+ influx inhibitory activities of benidipine for aldosterone production. T-type Ca2+ channels may contribute to additional benefits of this drug for treating renal and cardiovascular diseases, beyond its primary anti-hypertensive effects from blocking L-type Ca2+ channels.