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J Yun,
H Park,
J-H Ko,
W Lee,
K Kim,
T Kim,
J Shin,
J-H Song,
Y-H Noh, H Bang,
I Lim
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ABSTRACT: Cell proliferation and apoptosis are responsible for maintaining normal tissue homeostasis, and K(+) currents play important roles in regulating the physiological balance between them. This function of Ca(2+)-activated K(+) (K(Ca)) channels has been demonstrated in many types of tissues, but not in dermal fibroblasts. We investigated the expression of K(Ca) channels and their effects on proliferation and apoptosis in human dermal fibroblasts.
We used discoidin domain receptor 2 immunostaining to identify human dermal fibroblasts, and reverse transcription polymerase chain reaction, Western blot analysis and electrophysiological patch clamp recordings to evaluate the expression and characteristics of 3 members of the K(Ca) channel family, large-conductance K(Ca) (BK), intermediate-conductance K(Ca) (IK) and small-conductance K(Ca) channels. We also used the 3-(4,5-dimethyl-2-yl)-2,5-diphenyltetrazolium bromide assay, flow cytometry, Hoechst 33258 staining and Depsipher staining to investigate the effects of K(Ca) channels on cell proliferation and the mechanisms involved.
All 3 members of the K(Ca) channel family were found in fibroblasts. 1,3-Dihydro-1-[2-hydroxy-5-(trifluoromethyl)phenyl]-5-(trifluoromethyl)-2H-benzimidazol-2-one (NS1619, a BK channel activator) or 1-ethyl-2-benzimidazolinone (EBIO, an IK channel activator) decreased the proliferation of fibroblasts and induced apoptotic changes by mitochondrial membrane potential disruption. However, a pan-caspase inhibitor (Z-VAD-fmk) failed to prevent the apoptotic changes. Our findings indicate that 3 types of functional K(Ca) channels are expressed in human dermal fibroblasts and are involved in apoptosis of the cells through the mitochondrial apoptotic pathway, but seemingly in a caspase-independent manner.
Skin pharmacology and physiology 12/2009; 23(2):91-104. · 2.92 Impact Factor
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ABSTRACT: Differential effects of ATP on tetrodotoxin-sensitive (TTX-S) and tetrodotoxin-resistant (TTX-R) sodium currents in rat dorsal root ganglion neurons were studied using the whole-cell variation of path-clamp technique. Currents were evoked by step depolarizations to 0 mV from a holding potential of -80 mV. ATP suppressed TTX-S sodium currents while it increased TTX-R sodium currents. The effects were concentration-dependent and were reversible upon washout with ATP-free external solution. ATP-gamma-S, a hydrolysis-resistant ATP analog, also affected two types of sodium currents similarly to ATP, excluding the possibility that the effects were caused by the products of ATP hydrolysis, namely adenosine. ATP by modulating sodium currents may exert profound effects on the transmission of sensory information such as nociception.
Neuroreport 01/2002; 12(17):3659-62. · 1.66 Impact Factor
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ABSTRACT: In order to investigate the large-conductance Ca(2+)-activated K(+) (BK(Ca)) channel and determine the effects of nitric oxide (NO) on the channel in human skin fibroblasts, we performed electrophysiological patch clamp recordings on 5th-passage cells of human genital skin cultures. The whole-cell outward K(+) current was increased with depolarization, and proved to be sensitive to NS1619 (a selective BK(Ca) channel activator) and iberiotoxin (a specific BK(Ca )channel inhibitor). The single-channel currents showed 226 pS of mean conductance in symmetrical K(+). Sodium nitroprusside (SNP; an NO donor) significantly increased the K(+) current amplitude in the whole-cell mode, and open probability of the channel (NPo) in the cell-attached mode, but not in the inside-out mode. S-nitroso-N-acetylpenicillamine (an NO donor) and 8-Br-cGMP (a membrane-permeant cGMP analogue) also increased the BK(Ca )channel activity. The stimulatory effect of SNP on BK(Ca) channels was inhibited by pretreatment with 1H-[1,2,4]-oxadiazolo[4,3-a]quinoxalin-1-one (a soluble guanylyl cyclase inhibitor), or KT5823 [a specific protein kinase G (PKG) inhibitor]. Cytoplasmic PKG also increased the channel activity in inside-out patches. In conclusion, the present data indicate that BK(Ca) channels constitute a significant fraction of K(+) current in human skin fibroblasts, and that NO increases NPo of BK(Ca) channels, which are mediated via the cGMP/PKG pathway, without direct effects on the channel.
Skin pharmacology and physiology 18(6):279-87. · 2.92 Impact Factor
