2.Alterations in Calcium-Activated Potassium Channel Expressions in Human Prostate Cancer.
Duk Yoon KIM ; Eun Kyoung YANG ; Jung Wook KIM
Korean Journal of Urology 2006;47(5):548-552
PURPOSE: Recent studies have shown that potassium (K+) and sodium channels are involved in prostate cell growth. However, a great many of the studies have been done in prostate cancer cell lines and there are only scant studies on prostate cancer and benign prostatic hypertrophy (BPH) tissue. The present study was aimed to evaluate the alterations of the calcium-activated K+ channel (KCa) expression in prostate cancer, and to compare them with the expression profiles in human BPH tissue to understand their potential role in the progression of prostate cancer. MATERIALS AND METHODS: The prostate tissues obtained from radical prostatectomy (n=10) and transurethral resection of the prostate (n=18) were quickly frozen in liquid nitrogen for the RNA measurements. The protein and mRNA levels of the KCa subtypes and connexins were measured by performing immunoblot analysis and reverse-transcription polymerase chain reaction, respectively. RESULTS: The mRNA levels of type 2 (SK2) and type 3 (SK3) small-conductance and large-conductance (BK) KCas in the prostate cancer tissues were decreased more than 50% compared with those in the BPH samples. In addition, the BK and SK2 protein levels in prostate cancer were also significantly lower than those in the BPH. As reported previously, the connexin 26 and 43 transcript signals in the prostate cancer were significantly reduced compared with those in the BPH samples. CONCLUSIONS: These results suggest that the impaired expression of KCas may have a role in tumor progression via aberrant and uncontrolled prostate cell growth.
Cell Line
;
Connexins
;
Humans*
;
Large-Conductance Calcium-Activated Potassium Channels
;
Nitrogen
;
Polymerase Chain Reaction
;
Potassium
;
Potassium Channels, Calcium-Activated*
;
Prostate*
;
Prostatectomy
;
Prostatic Hyperplasia
;
Prostatic Neoplasms*
;
RNA
;
RNA, Messenger
;
Small-Conductance Calcium-Activated Potassium Channels
;
Sodium Channels
3.Unique interactions between scorpion toxins and small conductance Ca(2+)-activated potassium channels.
Fan YANG ; Zong-Yun CHEN ; Ying-Liang WU
Acta Physiologica Sinica 2015;67(3):255-260
Small conductance Ca(2+)-activated potassium channels (SK channels) distributing in the nervous system play an important role in learning, memory and synaptic plasticity. Most pharmacological properties of them are determined by short-chain scorpion toxins. Different from most voltage-gated potassium channels and large-conductance Ca(2+)-activated potassium channels, SK channels are only inhibited by a small quantity of scorpion toxins. Recently, a novel peptide screener in the extracellular pore entryway of SK channels was considered as the structural basis of toxin selective recognition. In this review, we summarized the unique interactions between scorpion toxins and SK channels, which is crucial not only in deep-researching for physiological function of SK channels, but also in developing drugs for SK channel-related diseases.
Animals
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Memory
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Neuronal Plasticity
;
Scorpion Venoms
;
chemistry
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Scorpions
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Small-Conductance Calcium-Activated Potassium Channels
;
antagonists & inhibitors
4.Construction and identification of the expression plasmid of SK2 (KCNN2) gene from human atrial myocytes with overlapping PCR.
Xiao-Qiu TAN ; Gui-Lan CHEN ; Tao LI ; Liang MAO ; Isao INOUE ; Yan YANG ; Xiao-Rong ZENG
Chinese Journal of Applied Physiology 2012;28(4):381-384
OBJECTIVESmall conductance calcium activated potassium channels type 2 (SK2) play a crucial role in atrial repolarization. It is difficult to acquire the full-length of its coded gene KCNN2 by RT-PCR with one step. We aim to get the full-length of KCNN2 gene and construct the plasmid by Overlapping PCR, and further more discuss the application of Overlapping PCR.
METHODSTotal RNA was extracted from human right atrial tissue and cDNA was acquired with reverse transcription. Overlapping PCR was conducted with three pairs of primers which were designed according to the sequence of KCNN2 (AY258141) gene. The expression plasmid of pIRES-hrGFP-SK2 was constructed by directed cloning with restriction enzyme site and identified by enzyme cutting and sequencing.
RESULTSThree parts of PCR amplification were consistent with predicted size. The sequence of the plasmid was consistent with the gene-bank data except two sites, however, which were the same as gene in different tissues.
CONCLUSIONThe expression plasmid pIRES-hrGFP-SK2 was constructed successfully. Overlapping PCR is a good choice for amplifying these genes with long size or low expression.
Base Sequence ; Gene Expression ; Humans ; Myocytes, Cardiac ; Plasmids ; genetics ; Polymerase Chain Reaction ; methods ; Small-Conductance Calcium-Activated Potassium Channels ; genetics
5.Diabetes mellitus reduces the expression of SK3 in rat cavernous tissues.
Jin-hai ZHU ; Rui-peng JIA ; Lu-wei XU ; Jian-ping WU ; Kai LIANG ; Qi SU
National Journal of Andrology 2010;16(3):236-239
OBJECTIVESK3, one of the small conductance calcium-activated potassium channels, is the key substance of the endothelium-derived hyperpolarizing factor (EDHF) passway. This study aimed to investigate the expression of SK3 in the cavernous tissue of rats with diabetes mellitus (DM).
METHODSTwenty-six DM models were made by injection of streptozocin (STZ) out of 50 male Sprague-Dawley rats, and another 15 that failed to be modeled were included in an STZ group. Ten healthy male rats were taken as blank controls. Eight weeks later, the penile erectile function of the rats was detected by injection of apomorphine (APO) at 80 microg/kg, and the expression of SK3 in the cavernous tissue was determined by RT-PCR and Western blot.
RESULTSPenile erection was observed in 14 (54%) of the 26 DM rat models and in all the rats of the STZ and blank control groups. Both the mRNA and protein expressions of SK3 were significantly lower in the DM (0.50 +/- 0.09 and 0.65 +/- 0.06) than in the STZ (1.15 +/- 0.03 and 1.28 +/- 0.04) and blank control groups (1.21 +/- 0.04 and 1.34 +/- 0.05) (P < 0.05). There were no significant differences between the STZ and blank control groups in either penile erection or mRNA and protein expressions of SK3 (P > 0.05).
CONCLUSIONDiabetes mellitus can significantly reduce erectile function in rats, which may be related to the decreased expression of SK3 in the corpus cavernosum.
Animals ; Diabetes Mellitus, Experimental ; metabolism ; Erectile Dysfunction ; metabolism ; Male ; Penis ; metabolism ; Potassium Channels, Calcium-Activated ; metabolism ; RNA, Messenger ; genetics ; Rats ; Rats, Sprague-Dawley ; Small-Conductance Calcium-Activated Potassium Channels
6.Increased small conductance calcium-activated potassium channel (SK2 channel) current in atrial myocytes of patients with persistent atrial fibrillation.
Miao-ling LI ; Tao LI ; Ming LEI ; Xiao-qiu TAN ; Yan YANG ; Tai-peng LIU ; Jie PEI ; Xiao-rong ZENG
Chinese Journal of Cardiology 2011;39(2):147-151
OBJECTIVETo compare the amplitude of the SK2 current (small conductance calcium-activated potassium channel) in human atrial myocytes with or without persistent atrial fibrillation (AF).
METHODSRight atrial appendage was obtained from 15 patients with sinus rate (SR) and 7 patients with AF underwent surgical valve replacement. Single myocyte was isolated by enzymatic dissociation method and the SK2 channel current density was recorded using whole-cell patch clamp techniques to detect the changes. Immunofluorescence was used to observe SK2 channel protein distribution on right atrial appendage.
RESULTSUsing the whole cell patch-clamp recording techniques, an inward rectifier K(+) mix currents could be obtained from both SR (n = 15) and AF (n = 7) samples, I(K1) mix currents density in single myocyte of AF group was significantly increased than in SR group [(-16.42 ± 5.32) pA/pF vs (-6.59 ± 2.24) pA/pF, P < 0.01], which could be partially inhibited by apamin (100 nmol/L). The apamin-sensitive current was obtained by subtraction of the currents before and after treatment with apamin. SK2 current density was significantly increased in AF group than that of SR group [(-9.81 ± 2.54) pA/pF vs (-3.67 ± 0.37) pA/pF, P < 0.01]. SK2 channel protein was evidenced with immunofluorescence method in right atrial appendage from AF group and SR group.
CONCLUSIONSK2 channel protein and current were present in atrial myocytes. The SK2 current density was significantly increased in AF group than in SR group suggesting that the increase of SK2 current might contribute to the electrical remodeling in AF patients.
Apamin ; pharmacology ; Atrial Fibrillation ; metabolism ; Cells, Cultured ; Female ; Humans ; Male ; Myocytes, Cardiac ; metabolism ; Patch-Clamp Techniques ; Small-Conductance Calcium-Activated Potassium Channels ; drug effects ; metabolism
7.Role of endothelium-derived hyperpolarizing factor in shear stress-induced endothelium-dependent relaxations of rats.
Hui-Ying ZHAO ; Quan LIU ; Bao-Rong CHI
Acta Pharmaceutica Sinica 2005;40(6):491-495
AIMTo investigate the role and mechanism of endothelium-derived hyperpolarizing factor (EDHF) in shear stress induced vasorelaxation of rat mesenteric artery.
METHODSThe changes in vessel diameter in response to variable flow (0-300 microL.min(-1)) were continuously examined. The contribution of prostacyclin (PGI2), NO and EDHF to shear stress induced relaxation were analyzed by inhibitory effects of indomethacin, N(G)-nitro-L-arginine (L-NA) and KCl. The nature and hyperpolarizing mechanism of EDHF were examined by the inhibitory effects of inhibitors of cytochrome P450 pathway and of various K+ channels.
RESULTSThe shear stress-induced relaxation were endothelium dependent and the contribution of NO was more prominent in large mesenteric arteries (400-500 microm) than that in resistance arteries (150-250 microm), whereas that of EDHF was noted in both-sized blood vessels. Tetrabutylammonium (a nonselective inhibitor of K channels) almost abolished, whereas the combination of charybdotoxin (an inhibitor of both large and intermediate-conductance Ca2+-activated K channels) and apamin (an inhibitor of small-conductance Ca2+-activated K channels) significantly inhibited the EDHF-mediated component of the shear stress-induced relaxations.
CONCLUSIONEDHF plays an important role in shear stress-induced endothelium-dependent relaxations, and K channels especially calcium-activated K channels appear to be involved.
Animals ; Apamin ; pharmacology ; Biological Factors ; physiology ; Charybdotoxin ; pharmacology ; Cytochrome P-450 Enzyme Inhibitors ; Endothelium, Vascular ; drug effects ; physiology ; In Vitro Techniques ; Large-Conductance Calcium-Activated Potassium Channels ; antagonists & inhibitors ; Male ; Mesenteric Arteries ; drug effects ; physiology ; Nitric Oxide ; physiology ; Potassium Channel Blockers ; pharmacology ; Proadifen ; pharmacology ; Quaternary Ammonium Compounds ; pharmacology ; Rats ; Rats, Wistar ; Small-Conductance Calcium-Activated Potassium Channels ; antagonists & inhibitors ; Vasodilation ; drug effects
8.Oxidative stress impairs IKCa- and SKCa-mediated vasodilatation in mesenteric arteries from diabetic rats.
Limei ZHAO ; Yan WANG ; Xiaozhen MA ; Yawen WANG ; Xiuling DENG
Journal of Southern Medical University 2013;33(7):939-944
OBJECTIVETo investigate the role of oxidative stress in impaired intermediate-conductance Ca(2+)-activated potassium channels (IKCa)- and small-conductance Ca(2+)-activated potassium channels (SKCa)-mediated relaxation in diabetic resistance arteries.
METHODSRat diabetic model was induced by a high fat and high glucose diet and streptozotocin (STZ) injection. Endothelial function of mesenteric arteries was assessed with the use of wire myography. The expression levels of IKCa and SKCa in cultured human umbilical vein endothelial cells (HUVECs) treated with H2O2 and/or antioxidant alpha lipoic acid (ALA) were measured using Western blotting.
RESULTSIKCa- and SKCa-mediated vasodilatation in response to acetylcholine was impaired in the third-order mesenteric arterioles of diabetic rats. In cultured HUVECs, H2O2 significantly decreased the protein expression of IKCa and SKCa. ALA alleviated the impairment of both vasodilatation function of the mesenteric arterioles ex vivo and enhanced the expression of IKCa and SKCa challenged with H2O2 in cultured HUVECs.
CONCLUSIONOur data demonstrated for the first time that impaired IKCa- and SKCa-mediated vasodilatation in diabetes was induced, at least in part, by oxidative stress via down-regulation of IKCa and SKCa channels.
Animals ; Cells, Cultured ; Diabetes Mellitus, Experimental ; metabolism ; physiopathology ; Human Umbilical Vein Endothelial Cells ; drug effects ; pathology ; Humans ; Hydrogen Peroxide ; pharmacology ; Intermediate-Conductance Calcium-Activated Potassium Channels ; metabolism ; Male ; Mesenteric Arteries ; physiopathology ; Oxidative Stress ; Rats ; Rats, Sprague-Dawley ; Small-Conductance Calcium-Activated Potassium Channels ; metabolism ; Thioctic Acid ; pharmacology ; Vasodilation
9.Colonic Dysmotility in Murine Partial Colonic Obstruction Due to Functional Changes in Interstitial Cells
Qianqian WANG ; Jingyu ZANG ; Xu HUANG ; Hongli LU ; Wenxie XU ; Jie CHEN
Journal of Neurogastroenterology and Motility 2019;25(4):589-601
BACKGROUND/AIMS: Interstitial cells play important roles in gastrointestinal (GI) neuro-smooth muscle transmission. The underlying mechanisms of colonic dysmotility have not been well illustrated. We established a partial colon obstruction (PCO) mouse model to investigate the changes of interstitial cells and the correlation with colonic motility. METHODS: Western blot technique was employed to observe the protein expressions of Kit, platelet-derived growth factor receptor-α (Pdgfra), Ca²⁺-activated Cl⁻ (Ano1) channels, and small conductance Ca²⁺- activated K⁺ (SK) channels. Colonic migrating motor complexes (CMMCs) and isometric force measurements were employed in control mice and PCO mice. RESULTS: PCO mice showed distended abdomen and feces excretion was significantly reduced. Anatomically, the colon above the obstructive silicone ring was obviously dilated. Kit and Ano1 proteins in the colonic smooth muscle layer of the PCO colons were significantly decreased, while the expression of Pdgfra and SK3 proteins were significantly increased. The effects of a nitric oxide synthase inhibitor (L-NAME) and an Ano1 channel inhibitor (NPPB) on CMMC and colonic spontaneous contractions were decreased in the proximal and distal colons of PCO mice. The SK agonist, CyPPA and antagonist, apamin in PCO mice showed more effect to the CMMCs and colonic smooth muscle contractions. CONCLUSIONS: Colonic transit disorder may be due to the downregulation of the Kit and Ano1 channels and the upregulation of SK3 channels in platelet-derived growth factor receptor-α positive (PDGFRα⁺) cells. The imbalance between interstitial cells of Cajal-Ano1 and PDGFRα-SK3 distribution might be a potential reason for the colonic dysmotility.
Abdomen
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Animals
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Apamin
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Blotting, Western
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Chloride Channels
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Colon
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Down-Regulation
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Feces
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Interstitial Cells of Cajal
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Mice
;
Muscle, Smooth
;
Myoelectric Complex, Migrating
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Nitric Oxide Synthase
;
Platelet-Derived Growth Factor
;
Silicon
;
Silicones
;
Small-Conductance Calcium-Activated Potassium Channels
;
Up-Regulation
10.Effect of Kv1.3 and KCa3.1 potassium ion channels on the proliferation and migration of monocytes/macrophages.
Shuang-Xia ZHANG ; Xian-Pei WANG ; Chuan-Yu GAO ; Chen-Hui JU ; Li-Jie ZHU ; Yi-Mei DU
Acta Physiologica Sinica 2015;67(5):505-512
This study was aimed to investigate the effects of blockade of Ca(2+) activated channel KCa3.1 and voltage-gated potassium channel Kv1.3 of the monocytes/macrophages on inflammatory monocyte chemotaxis. Chemotaxis assay was used to test the inflammatory Ly-6C(hi) monocyte chemotaxis caused by the monocytes/macrophages. The proliferation of monocytes/macrophages was detected by cell counting kit-8 (CCK8). Enzyme-linked immunosorbent assay (ELISA) was applied to detect the C-C motif ligand 7 (CCL7) in cultured media. The results showed that the recruitment of Ly-6C(hi) monocyte induced by monocytes/macrophages was suppressed by the potent Kv1.3 blocker Stichodactyla helianthus neurotoxin (ShK) or the specific KCa3.1 inhibitor TRAM-34. Meanwhile, the proliferation of monocytes/macrophages was significantly inhibited by ShK. The response of Ly-6C(hi) monocyte pretreated with ShK or TRAM-34 to CCL2 was declined. These results suggest that KCa3.1 and Kv1.3 may play an important role in monocytes/macrophages' proliferation and migration.
Cell Movement
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Cell Proliferation
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Cnidarian Venoms
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pharmacology
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Enzyme-Linked Immunosorbent Assay
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Humans
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Kv1.3 Potassium Channel
;
antagonists & inhibitors
;
physiology
;
Macrophages
;
cytology
;
Monocytes
;
cytology
;
Protein Structure, Tertiary
;
Pyrazoles
;
pharmacology
;
Small-Conductance Calcium-Activated Potassium Channels
;
antagonists & inhibitors
;
physiology