IL-β의 인슐린 분비 자극효과와 그 과정에 관여하는 인자들

정인경,오승훈,강동묵,정재훈,민용기,이명식,이문규,김광원 대한당뇨병학회 2002 Diabetes and Metabolism Journal Vol.24 No.4

연구배경: IL­1β는 용량과 노출시간에 따라 인슐린분비에 대해 상반되는 효과를 가지고 있다. IL­1β의 인슐린 분비 억제효과는 제1혁명 당뇨병의 자가면역 기전과 관련되어 잘 알려져 있으나 인슐린 자극효과에 대해서는 아직 명백히 밝혀지지 않았다. 이에 저자 등은 IL­1β의 다양한 농도에 따라 백서의 췌도세포에서 인슐린 분비에 미치는 영향을 살펴보고, 그 기전으로 인슐린 생합성, iNOS의 발현, 칼슘통로의 활성도 변화여부를 알아보고자 하였다. 방법:200∼300g인 수컷 Sprague­Dawley 백서의 췌도를 변형된 Lacy&Kostianovsky's 방법으로 분리한 후 IL­1β의 다양한 농도(0, 0.5, 5, 50, 500pmol/L)에 2, 6, 24시간 노출시켜 췌도세포의 형태, 생존능을 관찰하고 인슐린 분비능 및 췌도세포내 인슐린 함량을 측정하였으며, 전전구 인슐린 mRNA발현, iNOS mRNA발현을 RT­PCR을 통해 확인하였고, 세포의 칼슘 통로 활성도 변화 여부를 측정하였다. 결과:1) IL­1β에 노출되 췌도의 생존능:2시간 노출시는 대조군과 차이가 없었으나, 6시간 고농도군과 24시간 모든 군에서 생존능이 감소되었다. 2)인슐린 분비능은 IL­1β를 2시간 5poml/L이상의 고농도와 6시간 0.5pmol/L 저농도 처리시 대조군에 비해 의미있게 증가하였으나, 6시간과 24시간 5poml/L이상의 고농도에서는 의미있게 인슐린의 분비가 억제 되었다. 3)췌도내 인슐린양의 변화는 IL­1β의 시간과 농도에 따라 배지내의 인슐린 변화와 비슷한 경향을 보였으나 통계학적으로 의미있는 차이는 없었다. 4)전전구인슐린의 mRNA발현은 2시간 50pmol/L이상 고농도의 IL­1β에서 의미있게 증가하였고, 6시간 처리군에서는 의미있는 차이는 없었으나, 24시간에서 IL­1β의 용량의존적으로 억제되었다. 5)iNOS mRNA는 IL­1β 처리 후 2시간부터 발현되기 시작하여, 6시간에 최고에 달한 후 24시간에는 점차 감소하였다. IL­1β의 처리시간과 무관하게 5poml/L이상의 고농도에서 용량에 따라 발현이 증가하였다. 6)칼슘통로 활성도는 IL­1β의 농도나 시간에 따라 유의한 차이가 없었다. 결론: IL­1β는 단시간 고용량이나 장기간 저용량에서 인슐린 분비와 생합성을 증가시키며, 이런 효과는 iNOS나 칼슘 통로 활성도 변화와는 무관한 것으로 생각된다. Background : The inhibitory effort of IL-1β on the insulin secretion has been validated in pathogenesis of type 1 diabetes, but complex results about the stimulatory effect of IL-1β have been reported. The aims of this study are to clarify the effects of IL-1βon insulin secretion of pancreatic islets and to investigate the mechanisms in terms of preproinsulin synthesis, inducible NOS expression, and calcium channel activity. Method : Islets were isolated from male Sprague-Dawley (SD) rat by modified Lacy-Kostianovsky's method. After islets were treated with different concentrations (0, 0.5, 5, 50, 500 pmol/L) and exposure time (2, 6, 24 hours) of IL-1β, morphology, viability, static stimulation of insulin to glucose, insulin content, preproinsulin mRNA expression, iNOS mRNA expression and calcium channel activity were measured. Results : 1) Viability o islets was reduced in high concentrations of long term exposure of IL-1β. 2) Insulin secretion was stimulated in islets treated with 5, 50, and 500 pmol/L of IL-1β for 2 hours. 3) Insulin content was not significantly different regardless of concentration and exposure time of IL-1β. 4) Preproinsulin mRNA expression increased in islets treated with 50, 500 pmol/L of IL-1β for 2 hours. After 24 hours, it decreased in dose dependent manner. 5) iNOS mRNA expression was detectable after 2 hours in the presence of IL-1β, peaks at 6 hour and decreased after 24 hours. It was increased above 5 pmol/L of IL-1β in dose dependent manner. 6) Activities of the voltage-dependent Ca^2+ channels were not different among groups. Conclusion : IL-1β plays a positive role in terms of insulin secretion and insulin synthesis in high concentration of short term or low concentration of long term. These effects of IL-1β might be neither dependent of iNOS pathway nor Ca^2+ channel activity (J Kor Diabetes Asso 431~443, 2000).

Evidence for Adenosine Triphosphate (ATP) as an Excitatory Neurotransmitter in Guinea-Pig Gastric Antrum

Tong Mook Kang,Wenxie Xu,Sung Joon Kim,Seung Cheol Ahn,Young Chul Kim,Insuk So,Myoung Kyu Park,Dae-Yong Uhm,Ki Whan Kim 대한생리학회-대한약리학회 1999 The Korean Journal of Physiology & Pharmacology Vol.3 No.2

<P> We explore the question of whether adenosine 5 -triphosphate (ATP) acts as an excitatory neurotransmitter in guinea-pig gastric smooth muscle. In an organ bath system, isometric force of the circular smooth muscle of guinea-pig gastric antrum was measured in the presence of atropine and guanethidine. Under electrical field stimulation (EFS) at high frequencies (>20 Hz), NO-mediated relaxation during EFS was followed by a strong contraction after the cessation of EFS (a "rebound-contraction"). Exogenous ATP mimicked the rebound-contraction. A known P<SUB>2Y</SUB>-purinoceptor antagonist, reactive blue 2 (RB-2), blocked the rebound-contraction while selective desensitization of P<SUB>2X</SUB>-purinoceptor with α,β-MeATP did not affect it. ATP and 2-MeSATP induced smooth muscle contraction, which was effectively blocked by RB-2 and suramin, a nonselective P<SUB>2</SUB>-purinoceptor antagonist. Particularly, in the presence of RB-2, exogenous ATP and 2-MeSATP inhibited spontaneous phasic contractions, suggesting the existence of different populations of purinoceptors. Both the rebound-contraction and the agonist-induced contraction were not inhibited by indomethacin. The rank orders of agonists potency were 2-MeSATP > ATP ≥ UTP for contraction and α,β-MeATP ≥ β,γ-MeATP for inhibition of the phasic contraction, that accord with the commonly accepted rank order of the classical P<SUB>2Y</SUB>-purinoceptor subtypes. Electrical activities of smooth muscles were only slightly influenced by ATP and 2-MeSATP, whereas α,β-MeATP attenuated slow waves with membrane hyperpolarization. From the above results, it is suggested that ATP acts as an excitatory neurotransmitter, which mediates the rebound-contraction via P<SUB>2Y</SUB>-purinoceptor in guinea-pig gastric antrum.

Two Types of Voltage-dependent Outward Potassium Currents in Smooth Muscle Cells of Rabbit Basilar Atery

Tong Mook Kang,Insuk So,Dae Yong Uhm,Ki Whan Kim 대한생리학회-대한약리학회 1997 The Korean Journal of Physiology & Pharmacology Vol.1 No.2

<P> We have investigated the two types of voltage-dependent outward potassium (K) currents, i.e. delayed rectifier K current (I<SUB>K(V)</SUB>) and A-like transient outward K current (I<SUB>to</SUB>) with patch-clamp technique in single smooth muscle cells (SMCs) isolated from rabbit basilar artery, and investigated the characteristics of them. The time-courses of activation were well fitted by exponential function raised to second power (n<SUP>2</SUP>) in I<SUB>K(v)</SUB> and fourth power (n<SUP>4</SUP>) in I<SUB>to</SUB>. The activation, inactivation and recovery time courses of I<SUB>to</SUB> were much faster than that of I<SUB>K(V)</SUB>. The steady-state activation and inactivation of I<SUB>K(V)</SUB> was at the more hyperpolarized range than that of I<SUB>to</SUB> contrary to the reports in other vascular SMCs. Tetraethylammonium chloride (TEA; 10 mM) markedly inhibited I<SUB>K(V)</SUB> but little affected I<SUB>to</SUB>. 4-Aminopyridine (4-AP) had similar inhibitory potency on both currents. While a low concentration of Cd<SUP>2+</SUP> (0.5 mM) shifted the current- voltage relationship of I<SUB>to</SUB> to the positive direction without change of maximum conductance, Cd<SUP>2+</SUP> did not cause any appreciable change for I<SUB>K(V)</SUB>.

PKC-Independent Stimulation of Cardiac Na<SUP>+</SUP>/Ca<SUP>2+</SUP> Exchanger by Staurosporine

Tong Mook Kang 대한생리학회-대한약리학회 2008 The Korean Journal of Physiology & Pharmacology Vol.12 No.5

[Ca²⁺]_i transients by reverse mode of cardiac Na⁺/Ca²⁺exchanger (NCX1) were recorded in fura-2 loaded BHK cells with stable expression of NCX1. Repeated stimulation of reverse NCX1 produced a long-lasting decrease of Ca²⁺transients ( rundown ). Rundown of NCX1 was independent of membrane PIP₂ depletion. Although the activation of protein kinase C (PKC) was observed during the Ca²⁺ transients, neither a selective PKC inhibitor (calphostin C) nor a PKC activator (PMA) changed the degrees of rundown. By comparison, a non-specific PKC inhibitor, staurosporine (STS), reversed rundown in a dose-dependent and reversible manner. The action of STS was unaffected by pretreatment of the cells with calphostin C, PMA, or forskolin. Taken together, the results suggest that the stimulation of reverse NCX1 by STS is independent of PKC and/or PKA inhibition.

Two Types of Voltage-dependent Outward Potassium Currents in Smooth Muscle Cells of Rabbit Basilar Atery

Kang, Tong-Mook,So, In-Suk,Uhm, Dae-Yong,Kim, Ki-Whan The Korean Society of Pharmacology 1997 The Korean Journal of Physiology & Pharmacology Vol.1 No.2

We have investigated the two types of voltage-dependent outward potassium (K) currents, i.e. delayed rectifier K current ($I_{K(V)}$) and 'A-like' transient outward K current ($I_{to}$) with patch-clamp technique in single smooth muscle cells (SMCs) isolated from rabbit basilar artery, and investigated the characteristics of them. The time-courses of activation were well fitted by exponential function raised to second power ($n^2$) in $I_{K(V)}$ and fourth power ($n^4$) in $I_{to}$. The activation, inactivation and recovery time courses of $I_{to}$ were much faster than that of $I_{K(V)}$. The steady-state activation and inactivation of $I_{K(V)}$ was at the more hyperpolarized range than that of $I_{to}$ contrary to the reports in other vascular SMCs. Tetraethylammonium chloride (TEA; 10 mM) markedly inhibited $I_{K(V)}$ but little affected $I_{to}$. 4-Aminopyridine (4-AP) had similar inhibitory potency on both currents. While a low concentration of $Cd^{2+}$ (0.5 mM) shifted the current- voltage relationship of $I_{to}$ to the positive direction without change of maximum conductance, $Cd^{2+}$ did not cause any appreciable change for $I_{K(V)}$.

PKC-Independent Stimulation of Cardiac $Na^+/Ca^{2+}$ Exchanger by Staurosporine

Kang, Tong-Mook The Korean Society of Pharmacology 2008 The Korean Journal of Physiology & Pharmacology Vol.12 No.5

$[Ca^{2+}]_i$ transients by reverse mode of cardiac $Na^+/Ca^{2+}$ exchanger (NCX1) were recorded in fura-2 loaded BHK cells with stable expression of NCX1. Repeated stimulation of reverse NCX1 produced a long-lasting decrease of $Ca^{2+}$ transients ('rundown'). Rundown of NCX1 was independent of membrane $PIP_2$ depletion. Although the activation of protein kinase C (PKC) was observed during the $Ca^{2+}$ transients, neither a selective PKC inhibitor (calphostin C) nor a PKC activator (PMA) changed the degrees of rundown. By comparison, a non-specific PKC inhibitor, staurosporine (STS), reversed rundown in a dose-dependent and reversible manner. The action of STS was unaffected by pretreatment of the cells with calphostin C, PMA, or forskolin. Taken together, the results suggest that the stimulation of reverse NCX1 by STS is independent of PKC and/or PKA inhibition.

아세틸콜린과 콜레시스토키닌에 의한 사람 담낭 수축시 이용되는 칼슘의 근원

강동묵,이종균,이풍렬,이종철,윤용범,엄대용 대한소화기학회 1998 대한소화기학회지 Vol.31 No.6

Background/Aims: Gallbladder contraction regulates the bile flow during digestive period and pevents stasis of the lithogenic bile. In addition, abnormal gallbladder motility may cause recurtent biliary pain or biliary dyskinesia. Acetylcholine (ACh) and cholecystokinin (CCK) are major neurohormonal mediators in gallbladder contraction. In addition, an increase in cytosolic calcium concentration is a common event resulting in smooth muscle contraction. This study was designed to identify the sources of calcium utilized in gallbladder smooth muscle contraction mediated by ACh and CCK in human. Methods: By measuring the contractile forces of the muscle strips and the intracellular free Ca^(2+) conentration, Ca^(2+) sources utilized in muscle contraction were estimated. Results: The ACh-induced contractile response was accentuated after a twofold increment of the extracellular calcium concentration. The contractile response to ACh was markedly blocked by verapamil (10 μM) and was significantly potentiated by Bay-K 8644 (1 μM). Preventing Ca^(2+)release from the internal stores by strontium (4 mM) reduced the contractile response to ACh and CCK. The inhibitory effect was greater in the response to CCK than in the response to ACh Depletion of IP3-sensitive calcium stores by a high concentration of hista mine (50 μM) significantly inhibited the contractile response to CCK. The intfacellular caleium concentration was increased slowly on ACh-induced contraction but rapidly on CCK-induced contraction which was similar to the behavior of caffeine. Conclusions: These results suggest that ACh utilizes both intracellular and extracellular calcium, mainly extracellular calcium, whereas CCK utilizes calcium from intracellular calcium stores.

탄소고정 건설재료 및 구조시스템 연구실 (연세대-UNIST)

한동석,박경수,임윤묵,김건,Tong-Seok Han,Kyoungsoo Park,Yun Mook Lim,Gun Kim 한국건설순환자원학회 2023 한국건설순환자원학회지 Vol.18 No.1

Ca<SUP>2+</SUP>-dependent Long-term Inactivation of Cardiac Na<SUP>+</SUP>/Ca<SUP>2+</SUP> Exchanger

Jee-Eun Lee,Tong Mook Kang 대한생리학회-대한약리학회 2007 The Korean Journal of Physiology & Pharmacology Vol.21 No.1

Using BHK cells with stable expression of cardiac Na<SUP>+</SUP>/Ca<SUP>2+</SUP> exchanger (BHK-NCX1), reverse mode (i.e. Ca<SUP>2+</SUP> influx mode) of NCX1 current was recorded by whole-cell patch clamp. Repeated stimulation of reverse NCX1 produced a cytosolic Ca<SUP>2+</SUP>-dependent long-term inactivation of the exchanger activity. The degrees of inactivation correlated with NCX1 densities of the cells and were attenuated by reduced Ca<SUP>2+</SUP> influx via the reverse exchanger. The inactivation of NCX1 was attenuated by (i) inhibition of Ca<SUP>2+</SUP> influx with reduced extracellular Ca<SUP>2+</SUP>, (ii) treatment with NCX1 blocker (Ni<SUP>2+</SUP>), and (iii) increase of cytoplasmic Ca<SUP>2+</SUP> buffer (EGTA). In BHK-NCX1 cells transiently expressing TRPV1 channels, Ca<SUP>2+</SUP> influx elicited by capsaicin produced a marked inactivation of NCX1. We suggest that cytoplasmic Ca<SUP>2+</SUP> has a dual effect on NCX1 activities, and that allosteric Ca<SUP>2+</SUP> activation of NCX1 can be opposed by the Ca<SUP>2+</SUP>-dependent long-term inactivation in intact cells.

소(우(牛)) 식도구 윤상근의 비아드레날린 비콜린성 이완 및 수축

강동묵,한호재,양일석,Kang, Tong-mook,Han, Ho-jae,Yang, Il-suk 대한수의학회 1995 大韓獸醫學會誌 Vol.35 No.2

To characterize non-adrenergic non-cholinergic(NANC) nerve mediated contractile responses in circular smooth muscle of bovine reticular groove, we investigated NANC relaxation and contraction induced by electric field stimulation to enteric nerves. In the presence of atropine($1{\mu}M$) and guanethidine($50{\mu}M$), electric field stimulation at frequency of 1 to 16Hz(square pulses, 0.5ms duration, 70V) evoked clear-cut relaxations through stimulations. Transient 'rebound contraction' was occured when the stimulus was switched off. All of the responses (relaxation and rebound contraction) were dose-dependently blocked by Nw-nitro-$_{\small{L}}$-arginine methyl ester(L-NAME), an inhibitor of nitric oxide synthesis, and methylene blue, and inhibitor of soluble guanylate cyclase. Tetraethyl ammonium(TEA), a potassium channel blocker, did not block the NANC relaxations.