Xylazine-ketamine 병용마취가 토끼의 혈중 ACTH, corticosterone 및 glucose 농도에 미치는 영향

박권무,이용화,한성규,류판동,Park, Kwon-moo,Li, Long-hua,Han, Seong-kyu,Ryu, Pan-dong 대한수의학회 1999 大韓獸醫學會誌 Vol.39 No.1

Anesthetic agents are useful in inducing the anesthesia for surgical operations and various biological experiments, but they can disturb the body homeostasis and cause the stress in animals. Much efforts have been directed on reducing such side effects of anesthesia. In this work, we measured the serum ACTH, corticosterone and glucose concentration in rabbits to compare the degree of stress induced by two commonly-used anesthetics, ketamine, xylazine, and the combination of xylazine and ketamine. 1. The anesthesia was induced in about 10 min in the rabbits treated with xyalzine, ketamine and xylazine-ketamine. The duration of complete loss of righting reflex were 12, 13 and 115 min in the groups treated with xylazine, ketamine and xylazine-ketamine, respectively. 2. Serum ACTH concentrations in all treatment groups were higher than those in control group. At 30 min after the administration of the drugs, serum ACTH levels in ketamine-treated group were significantly higher than those in control, xylazine- and xylazine-ketamine-treated groups. However, at 1, 2, 5 and 9 hours after the drug administration, serum ACTH levels in xylazine-treated-group were higher than those in control. 3. Serum corticosterone levels in xylazine- and xylazine-ketamine-treated groups were lower than those in control or ketamine-treated groups at 0.5 and 1 hour after the administration. However, at 5 and 9 hours after the administration, serum corticosterone levels in xylazine- and xylazine-ketamine-treated groups were significantly higher than those in ketamine-treated group or control. 4. Serum glucose levels transiently increased to 3 times of the pre-injection levels at 0.5 and 1 hours after the administration in xylazine or xylazine-ketamine-treated groin, but were not changed in control and ketamine-treated group. These results indicate that xylazine-induced stress lasts longer than ketamine-induced, suggesting that the difference in stress-related hormone levels during anesthesia could be due to the differences in modes of actions of individual drugs used and the depth of anesthesia.

신생리 : 허혈/재관류에 의한 급성신부전에 있어 isocitrate dehydrogenase의 역할

박권무 ( Kwon Moo Park ),김진우 ( Jin U Kim ),김기영 ( Ki Young Kim ) 대한신장학회 2008 춘계학술대회 초록집 Vol.28 No.1

신장 근위세뇨관세포에서 고포도당이 IGF-I 결합과 포도당운반계에 미치는 영향

한호재,박권무,손창호,윤용달,Han, Ho-jae,Park, Kwon-moo,Son, Chang-ho,Yoon, Yong-dal 대한수의학회 1997 大韓獸醫學會誌 Vol.37 No.2

Diabetes mellitus is associated with a wide range of pathophysiological in the kidney. This study was designed to examine the effects of high glucose concentration on IGF-I binding and glucose transporters in renal proximal tubule cells. The results were as follows : The binding of $^{125}I-IGF-I$ reached the peak at the 30 minutes and gradually decreased by the time dependent manner. The binding of $^{125}I-IGF-I$ was inhibited by the unlabelled IGF-I($10^{-14}{\sim}10^{-8}M$) in a concentration dependent manner. The relative affinity of IGF-I receptor for IGF-I, IGF-II and insulin exhibited typical type 1 binding(IGF-I > insulin > IGF-II). However IGF-II did not compete for the cultured cell membrane $^{125}I-IGF-I$ binding site at $10^{-14}{\sim}10^{-8}M$. Under optimal conditions, IGF-I binding to the membranes from 5mM and 20mM glucose treated cells was analyzed. It was found that 20mM glucose treated cells exhibited higher binding activity for IGF-I. In order to further substantiate this increase in IGF-I binding sites, we performed affinity-labelling studies. The cross-linked cell membrane subjected to SDS-PAGE; labelled material was detected by autoradiography. 20mM glucose treated cells exhibited higher levels. The initial rate of $methyl-{\alpha}-D-glucopyranoside({\alpha}-MG)$ uptake was significantly lower($74.41{\pm}6.71%$) in monolayers treated with 20mM glucose than those of 5mM glucose. However, 3-O-methyl-D-glucose(3-O-MG) uptake was not affected by glucose concentration in culture media. IGF-I significantly increased ${\alpha}-MG$ uptake in both 5mM and 20mM glucose treated cells. However, 3-O-MG uptake was not affected by IGF-I in both conditions. In conclusion, 20mM glucose increased binding sites of $^{125}I-IGF-I$, inhibited Na/glucose cotransporter activity. But 20mM glucose did not change facilitated glucose transporter.

초대배양된 토끼 신장 근위세뇨관세포의 성장과 기능분화에 대한 insulin과 IGF의 효과 - Na⁺ uptake에 대한 IGF-I의 효과 -

한호재,박권무,이장헌,양일석,Han, Ho-jae,Park, Kwon-moo,Lee, Jang-hern,Yang, IL-suk The Korean Society of Veterinary Science 1996 大韓獸醫學會誌 Vol.36 No.4

이온운반계는 생체의 각기 다른 세포의 성장을 조절하는 성장조절인자들의 효과를 매개하는데 깊은 관련이 있는 것으로 보고되고 있다. 신장 근위세뇨관에서 솔변 연 $Na^+/H^+$ 상호운반계는 사구체에서 여과된 나트륨의 재흡수와 수소이온의 분비를 조절하는 중요한 기능을 수행한다. 이 연구는 초대배양된 신장 근위세뇨관세포의 나트륨 운반을 Insulin-like Growth Factor-I(IGF-I)이 어떤 경로를 통하여 조절하는지를 알아보고자 실시하였다. 결과는 아래와 같다. 1. 초대배양된 신장 근위세뇨관세포에서 $Na^+$ uptake는 시간의존적으로 증가되었으며, 30분동안 $Na^+$ uptake를 실시한 결과 세포외 NaCl 농도의존적으로 $Na^+$ uptake를 유의성있게 감소시켰다(대조군; $40.11{\pm}1.76$, 140mM군; $17.82{\pm}0.94pmole\;Na^+/mg\;protein/min$). 2. $Na^+$ uptake는 iodoacetic acid(IAA, $1{\times}10^{-4}M$) 또는 valinomycin($5{\times}10^{-6}M$)처리시 대조군에 비해 각각 $50.51{\pm}4.4%$와 $57.65{\pm}2.27%$ 억제되었으며, ouabain($5{\times}10^{-5}M$)을 처리한 경우는 $140.23{\pm}3.37%$ 증가되었다. IGF-I($1{\times}10^{-5}M$)으로 배양한 세포를 actinomycin D($1{\times}10^{-7}M$)와 cycloheximide($4{\times}10^{-5}M$)로 처리시 $Na^+$ uptake는 대조군에 비해 각각 $90.21{\pm}2.39%$와 $89.64{\pm}3.69%$로 감소되었다. 3. IGF-I으로 배양한 세포에서 세포외 cAMP는 농도의존적($10^{-8}-10^{-4}M$)으로 $Na^+$ uptake를 유의성있게 감소시켰고, 3-isobutyl-1-methyl-xanthine(IBMX, $5{\times}10^{-5}M$)도 억제시켰다. Pertussis toxin(PTX, 50pg/ml)이나 cholera toxin(CTX, $1{\mu}g/ml$)의 처리시에도 $Na^+$ uptake는 억제되었다. 세포외 phorbol 12-myristate 13 acetate(PMA) 또한 농도의존적(1-100ng/ml)으로 $Na^+$ uptake를 감소시켰다. 그러나 staurosporine($1{\times}10^{-7}M$)은 $Na^+$ uptake에 영향을 미치지 않았으며 PMA와 stauiosporine을 동시에 처리했을 때도 $Na^+$ uptake는 억제되지 않았다. 결론적으로 초대배양된 토끼 신장 근위세뇨관세포에서 $Na^+$ uptake는 막전위와 세포내 에너지 의존적이며 IGF-I은 부분적으로 단백질 및 RNA 합성을 통해서 그리고 세포내 cAMP나 PKC 경로를 통해서 $Na^+$ uptake를 조절하는 것으로 생각된다. It has been suggested that ion transport systems are intimately involved in mediating the effects of growth regulatory factors on the growth of a number of different types of animal cells in vivo. The functional importance of the apical membrane $Na^+/H^+$ antiporter in the renal proximal tubule is evidenced by estimates that this transporter mediates the reabsorption of approximately one third of the filtered load of sodium and the bulk of the secretion of hydrogen ions. This study was designed to investigate the pathway utilized by IGF-I in regulating sodium transport in primary cultured renal proximal tubule cells. Results were as follows : 1. $Na^+$ was observed to accumulate in the primary cells as a function of time. Raising the concentration of extracellular NaCl induced an decrease in $Na^+$ uptake compared with control cells in a dose dependent manner. The rate of $Na^+$ uptake into the primary cells was about two times higher in the absence of NaCl($40.11{\pm}1.76pmole\;Na^+/mg\;protein/min$) than in the presence of 140mM NaCl($17.82{\pm}0.94pmole\;Na^+/mg\;protein/min$) at the 30 minute uptake. 2. $Na^+$ uptake was inhibited by IAA($1{\times}10^{-4}M$) or valinomycin($5{\times}10^{-6}M$) treatment($50.51{\pm}4.04$ and $57.65{\pm}2.27$ of that of control, respectively). $Na^+$ uptake by the primary proximal tubule cells was significantly increased by ouabain($5{\times}10^{-5}M$) treatment($140.23{\pm}3.37%$ of that of control). When actinomycin D($1{\times}10^{-7}M$) or cycloheximide($4{\times}10^{-5}M$) was applied, $Na^+$ uptake was decreased to $90.21{\pm}2.39%$ or $89.64{\pm}3.69%$ of control in IGF-I($1{\times}10^{-5}M$) treated cells, respectively. 3. Extracellular cAMP decreased $Na^+$ uptake in a dose-dependent manner($10^{-8}-10^{-4}M$). IBMX($5{\times}10^{-5}M$) also inhibited $Na^+$ uptake. Treatment of cells with pertussis toxin(50pg/ml) or cholera toxin($1{\mu}g/ml$) inhibited $Na^+$ uptake. Extracellular PMA decreased $Na^+$ uptake in a dose-dependent manner(1-100ng/ml). 100 ng/ml PMA concentration significantly inhibited $Na^+$ uptake in IGF-I treated cells. However, staurosporine($1{\times}10^{-7}M$) had no effect on $Na^+$ uptake. When PMA and staurosporine were added together, the inhibition of $Na^+$ uptake was not observed. In conclusion, sodium uptake in primary cultured rabbit renal proximal tubule cells was dependent on membrane potentials and intracellular energy levels. IGF-I stimulates sodium uptake through mechanisms that involve some degree of de novo protein and/or RNA synthesis, and cAMP and/or PKC pathway mediating the action mechanisms of IGF-I.

초대배양한 신장 근위세뇨관세포에서 estradiol-17β와 IGF-I 수용체 발현과의 상관관계

한호재,남성안,박권무,Han, Ho-jae,Nam, Seong-ahn,Park, Kwon-moo 대한수의학회 1997 大韓獸醫學會誌 Vol.37 No.2

The mechanisms of $estradiol-17{\beta}$ regulating growth of both normal and neoplastic cells are not clear until now. In studies using various estrogen-dependent breast cell lines, it is recently known that estrogen controls the cell growth by regulating the expression of growth factors and/or their receptors. In the present study, we investigated the effects of $estradiol-17{\beta}$on cell growth and IGF-I binding sites using primary cultured renal proximal tubule cells. We have obtained results as follows : $Estradiol-17{\beta}(10^{-9})$ has stimulatory effects in cell growth. Cotreatment of $estradiol-17{\beta}(10^{-9}M)$ and $IGF-I(5{\times}10^{-8}M)$ significantly increased the growth of primary rabbit renal proximal tubule cells compared to that of $estradiol-17{\beta}$ or IGF-I alone treated cells. In binding studies, we found that the binding of $^{125}IGF-I$ on cell membranes was incubation time- and temperature-dependent. Incubation at $37^{\circ}C$ results in higher binding of $^{125}IGF-I$ than that of $23^{\circ}C$ or $4^{\circ}C$. Maximum binding was observed at $37^{\circ}C$ between 30 and 60 minutes. The binding of $^{125}IGF-I$ to both control and $estradiol-17{\beta}-treated$ cells was inhibited by unlabelled $IGF-I(10^{-8}{\sim}10^{-12}M)$ in a concentration-dependent manner. However, EGF did not compete for $^{125}IGF-I$ binding at $10^{-8}{\sim}10^{-12}M$. IGF-I binding to the membranes from both control and $estradiol-17{\beta}-treated$ cells was also analyzed. We found that $estradiol-17{\beta}-treated$ cells exhibited higher binding activity for IGF-I. When $estradiol-17{\beta}$ or tamoxifen alone, or $estradiol-17{\beta}$ and tamoxifen cotreated cells were compared, the binding ratio of $^{125}I-IGF-I$ of $estradiol-17{\beta}-treated$ cell was significantly increased but was similar to control in both $estradiol-17{\beta}$ and tamoxifen cotreated cell. These results suggest that $estradiol-17{\beta}$ in part controls cell proliferation by regulating the expression of IGF-I receptors in primary rabbit renal proximal tubule cells.

호르몬 한정배지를 이용한 세포 초대배양계의 확립

한호재,강주원,박권무,이장헌,양일석,Han, Ho-jae,Kang, Ju-won,Park, Kwon-moo,Lee, Jang-hern,Yang, Il-suk 대한수의학회 1996 大韓獸醫學會誌 Vol.36 No.3

This study investigated the properties of primary cultured proximal tubule cells in hormonally defined(insulin, transferrin, and hydrocortisone), serum-free medium or 10% serum-supplemented medium. The growth rate of the primary cultured proximal tubule cells was lower in the hormonally defined, serum-free medium than in the 10% serum- supplemented medium(p < 0.05), while the activities of brush border marker enzymes, alkaline phosphatase(AP), leucine aminopeptidase(LAP), and y-glutamyl transpeptidase(${\gamma}$-GTP) were increased(p < 0.05). The activities of these enzymes, however, decreased with the lapse of incubation time to 50-70% after 6 days culture compared to those of the freshly-prepared proximal tubules. The enzymatic activities of the primary cultured proximal tubul cells on 6, 9, 12, and 15 days of culture were significantly increased in the hormonally defined, serum-free medium compared to the 10% serum-supplemented medium(p < 0.05). The functional differentiation of the primary culture was examined by observing multicellular domes of the confluent monolayer, which is indicative of transepithelial solute transport. The dome formation by the proximal tubule cultures occurred at a higher frequency in the hormonally defined, serum-free medium than in the 10% serum-supplemented medium(p < 0.05). Upon electron microscopic examination, an increased density of the brush border was observed in the hormonally defined, serum-free medium compared to the cells grown in 10% serum-supplemented medium. The activities of $Na^+$glucose cotransporter($^{14}C$-a-MG uptake), $Na^+$phosphate cotransportere($^{32}P$ uptake) and $Na^+$ transporter($^{22}Na^+$ uptake) in the brush border membrane, and of $Na^+/K^+$-ATPase($^{86}Rb$ uptake) in the basolateral membrane were significantly stimulated in the hormonally defined, serum-free medium than in 10% serum-supplemented medium(p < 0.05). In conclusion, the primary cultured proximal tubule cells grown in the hormonally defined, serum-free medium demonstrated a slower growth rate, but the functions of cell were enhanced.

느린 세포주기를 가진 성체줄기세포의 생쥐 장기 별 분포

김진우(Jinu Kim),박권무(Kwon Moo Park) 대한해부학회 2007 Anatomy & Cell Biology Vol.40 No.4

성체줄기세포는 자가재생, 분화능력 그리고 느린 세포주기(slow cell cycle)를 가지고 있다. 본 연구에서는 성체의 어떤 장기에 그리고 장기의 어느 부위에 느린 세포주기 세포(Slow-cycling cell)가 있는가를 조사하기 위해 갓 태어난 생쥐를 3일간 bromodeoxyuridine (BrdU)를 주입하여 8주 후에 성체에서 BrdU를 가지고 있는 느린 세포주기 세포들의 위치를 장기 별로 조사하였다. 형광 또는 광학현미경을 통한 조직학적 관찰을 통해 고환에서는 정세관의 가장자리에 위치한 정조세포, 대뇌에서는 뇌실 아래층의 내벽에 존재하는 세포와 대뇌겉질 세포, 피부의 털주머니에서는 털망울을 둘러싼 주변세포, 눈의 망막에서는 속핵층에 위치한 세포, 허파에서는 세기관지의 상피세포층에 존재하는 세포, 콩팥에서는 겉질과 속질의 경계부위와 속속질부의 세관에 존재하는 상피세포, 그리고 심장에서는 심장근육세포 등에 세포주기가 느린 BrdU 양성 세포가 존재함을 확인하였다. 결론적으로 성체의 다양한 장기들은 세포주기가 느린 세포들을 가지고 있었고 이들은 성체장기의 정상적인 재생(turnover)과 손상 후 복원에 관여할 것으로 여겨진다. Adult stem cells possess the characteristics of self-renewal, multipotent, plasticity as well as slow cycling rate. We investigated a location of slow-cycling cells, that is, adult stem-like cells, in various organs in the 8 week-old mice which administered bromodeoxyuridine (BrdU) at neonatal phase. BrdU-retaining cells (slow-cycling cells) were observed in speramtogonia at the edge of seminiferous tubules in testes, hair root cells surrounding hair follicles, the cells in the inner nuclear layer of the retina, the myocytes in the hearts, the cells in the parenchyma and the Glisson’s capsule of liver, the cells in the epithelial layer of bronchioles, and the tubular epithelial cells in the kidneys. In conclusion, various organs of adult mouse expressed slow-cycling cells, indicating that the cells may associate with normal cell turnover and repair after damages.

$17{\beta}$-estradiol Attenuates Renal Fibrosis in Mice with Obstructive Uropathy

조민현,장희성,정경진,박권무,Cho, Min-Hyun,Jang, Hee-Seong,Jung, Kyung-Jin,Park, Kwon-Moo Korean Society of Pediatric Nephrology 2011 Childhood kidney diseases Vol.15 No.2

목적: 일반적으로 남자는 여자에 비해 만성 신장병의 발병이 많고 말기 신부전으로의 진행이 더 흔한 것으로 알려져 있다. 본 연구는 일측성 요관 폐쇄를 가진 생쥐에서 신섬유화에 대한 성별과 성호르몬의 효과를 규명하기 위해 시행되었다. 방법: 일측성 완전 요관 폐쇄 7일째 암컷과 수컷 생쥐의 신장에서 ${\alpha}$-smooth muscle actin (${\alpha}$-SMA)의 발현을 측정한 후, 암컷 생쥐에서 난소를 제거하거나 제거 후 다시 $17{\beta}$-estradiol을 보충하여 나타나는 신섬유화 정도를 비교 분석하였다. 결과:일측성 요관 폐쇄를 가진 암컷 신장의 ${\alpha}$-SMA의 발현이 수컷 신장에 비해 현저히 낮았다. 난소 제거와 $17{\beta}$-estradiol의 보충은 일측성 요관 폐쇄를 가진 암컷 신장의 안지오텐신 II 1형 수용체의 발현에는 의미 있는 영향을 주지 않았지만, 안지오텐신 II 2형 수용체의 발현은 정상 암컷과 난소 제거 후 $17{\beta}$-estradiol를 보충한 암컷에서 현저히 증가되었다. 또한, inducible nitric oxide synthase (iNOS) 역시 유사한 변화를 보였다. 결론 : 여성은 폐쇄성 요로병증에서 신섬유화에 대한 저항성과 연관이 있으며 이러한 성별의 차이는 $17{\beta}$-estradiol에 의한 안지오텐신 II 2형 수용체와 iNOS의 발현 증가와 연관이 있을 것으로 사료된다. Purpose : Men are generally more prone to chronic renal disease and progression to end stage renal disease than women. The purpose of this study is to prove the effect of gender and sex hormone on renal fibrosis in mice with unilateral ureteral obstruction (UUO) and to elucidate the specific underlying mechanisms. Methods :We compared the expression of ${\alpha}$-smooth muscle actin (${\alpha}$-SMA) in female and male mice with complete UUO (day 7). After this, we estimated the changes of renal fibrosis in the female mice with oophorectomy and in the female mice with oophorectomy and replacement of $17{\beta}$-estradiol, respectively. Results : The level of ${\alpha}$-SMA in the female kidney with UUO was significantly lower than that in the male kidney with UUO. oophorectomy and replacement of $17{\beta}$-estradiol did not change the expression of angiotensin II type 1 (AT1) receptor in the female kidney with UUO, whereas the expression of angiotensin II type 2 (AT2) receptor was significantly more elevated in the intact female (IF) and the oophorectomized female with estrogen (OF+E) than that in the oophorectomized female (OF). The expressions of inducible nitric oxide synthase (iNOS) in the IF and OF+E mice were significantly more elevated than that in the OF mice, which was similar to the expression of AT2 receptor. Conclusion : The female gender is associated with resistance to renal fibrosis in obstructive uropathy and this gender difference may originate from the existence of $17{\beta}$-estradiol, which has an anti-fibrotic effect via upregulation of the AT2 receptor and iNOS.

BMP 신호 하부 인자인 PV.1 단백질의 결합 단백질 동정

황유석(Yoo-Seok Hwang),채정필(Jeong-Pil Chae),김동선(Dong-Sun Kim),박권무(Kwon Moo Park),배용철(Yong Chul Bae),박매자(Mae Ja Park) 대한해부학회 2007 Anatomy & Cell Biology Vol.40 No.3

척추동물에서 BMP의 배(ventral)쪽 신호 전달 경로의 하위에 있는 전사조절인자들은 등배(dorsalventral)쪽 축을 따라 형성되는 중배엽에 있어서도 그 기능을 공유 한다는 것이 보고 되었다. 본 연구에서는 BMP 신호 물질의 하위 유전자인 PV.1 단백질이 배쪽과 등쪽에서의 신호전달에 있어서 차별적 기능을 알기 위하여 PV.1과 상호 결합하는 단백질을 검색하였다. 효모 이중 접합법을 이용하여 24개의 PV.1 결합 단백질을 검색하였고, 이중 결합하는 Xvent-2와 Xclaudin-6에 대한 PV.1 단백질의 도메인(domain) 연구를 하였다. PV.1 단백질의 C-terminus인 197-241 지역이 Xclaudin-6와 결합하는 반면에 Xvent-2는 PV.1 단백질의 C-terminus 전반에서 결합한다는 것을 확인하였다. 또한 PV.1과 결합하는 Xvent-2는 동형 이중 결합(Homodimerization)뿐만 아니라 PV.1과 결합하는 Xclaudin-6과도 이형 이중 결합(Heteodimerization)을 한다는 것을 확인하였다. Homeodomain transcription factors functioning downstream of BMP ventral pathway have been reported to share similar domain of roles in mesoderm patterning along the dorsal-ventral axis. To elucidate the differential role of PV.1 in the aspect of relationship between dorsal and ventral region, we tried to screen PV.1- interacting proteins. Twenty-four PV.1-interacting proteins were identified by yeast two-hybrid screening. Xvent-2 and Xclaudin-6 among these, went under domain study. The C-terminus of PV.1, more specifically 197-241 region was found to interact with Xclaudin-6. Meanwhile Xvent-2 has mild affinity to overall C-terminal region of PV.1. At the same time it was found that Xvent-2 homodimerizes and also binds to Xclaudin-6.

Transient Unilateral Renal Ischemia/reperfusion Induces Alteration in the Contralateral Kidney

장희성 ( Hee Seong Jang ),정경진 ( Kyoung Jin Jung ),박권무 ( Kwon Moo Park ) 대한신장학회 2008 춘계학술대회 초록집 Vol.28 No.1