장내분비 K-세포에서 인슐린-발현세포로의 분화

이에스더 ( Esder Lee ),유준모 ( Jun Mo Yu ),이민경 ( Min Kyung Lee ),류경렬 ( Gyeong Ryul Ryu ),고승현 ( Seung Hyun Ko ),안유배 ( Yu Bae Ahn ),문성대 ( Sung Dae Moon ),송기호 ( Ki Ho Song ) 대한당뇨병학회 2009 Diabetes and Metabolism Journal Vol.33 No.6

연구배경: 최근 제1형 당뇨병 치료법으로 췌도 이식의 새로운 발견에도 불구하고 췌장 공여자가 부족한 것이 여전히 중요한 장애물이다. 장상피에 존재하는 내분비세포(장내분비 세포)와 췌장 베타세포는 발생과정에서 유사한 분화경로를 공유한다. 특히, GIP를 분비하는 K-세포는 베타세포에서 관찰되는 중요한 단백질들이 많이 발현됨이 알려져있다. 따라서 저자들은 K-세포의 생물학적인 특성이 베타세포와 매우 유사하다는 점에 착안하여 이를 베타세포로 분화시키고자 본 연구를 시행하였다. 방법: 생쥐 췌장의 내분비 종양으로부터 기원한 복합 STC-1세포에서 K-세포를 순수 분리하였다. 또한, 안정적으로 Nkx6.1을 발현하는 K-세포 클론("Kn4-세포"로 명명)을 성공적으로 확보하였다. 혈청이 없는 조건에서 exendin-4를 처리함으로써 K-세포 또는 Kn4-세포가 시험관내에서 베타세포로 분화하도록 유도하였다. 인슐린 mRNA와 인슐린 단백질의 발현은 RT-PCR과 면역세포화학염색으로 조사하였다. 세포내 인슐린양도 측정하였다. 결과: K-세포에서 RT-PCR과 western blot 분석을 통하여 glucokinase와 GIP가 발현함을 확인하였다. RT-PCR의 결과에서 K-세포에서는 Pdx-1, NeuroD1/Beta2, MafA mRNA가 발현됨을 확인하였으나 Nkx6.1은 발현하지 않았다. Kn4-세포를 혈청이 없는조건에서 exendin-4을 처리한 결과, 인슐린 mRNA와 인슐린 또는 C-펩타이드 단백질이 뚜렷이 발현됨을 관찰하였다. 또한 세포내 인슐린양도 유의하게 증가하였다. 결론: K-세포는 특정한 조건에서 인슐린을 발현하는 세포로 분화가 되므로 장차 제1형 당뇨병의 세포치료법으로써 활용될 가능성이 있다. Background: Despite a recent breakthough in human islet transplantation for treating type 1 diabetes mellitus, the limited availability of donor pancreases remains a major obstacle. Endocrine cells within the gut epithelium (enteroendocrine cells) and pancreatic β cells share similar pathways of differentiation during embryonic development. In particular, K-cells that secrete glucose-dependent insulinotropic polypeptide (GIP) have been shown to express many of the key proteins found in β cells. Therefore, we hypothesize that K-cells can be transdifferentiated into β cells because both cells have remarkable similarities in their embryonic development and cellular phenotypes. Methods: K-cells were purified from heterogeneous STC-1 cells originating from an endocrine tumor of a mouse intestine. In addition, a K-cell subclone expressing stable Nkx6.1, called "Kn4-cells," was successfully obtained. In vitro differentiation of K-cells or Kn4-cells into β cells was completed after exendin-4 treatment and serum deprivation. The expressions of insulin mRNA and protein were examined by RT-PCR and immunocytochemistry. The interacellular insulin content was also measured. Results: K-cells were found to express glucokinase and GIP as assessed by RT-PCR and Western blot analysis. RT-PCR showed that K-cells also expressed Pdx-1, NeuroD1/Beta2, and MafA, but not Nkx6.1. After exendin-4 treatment and serum deprivation, insulin mRNA and insulin or C-peptide were clearly detected in Kn4-cells. The intracellular insulin content was also increased significantly in these cells. Conclusion: K-cells are an attractive potential source of insulin-producing cells for treatment of type 1 diabetes mellitus. However, more experiments are necessary to optimize a strategy for converting K-cells into β cells. (Korean Diabetes J 33:475-484, 2009)

제1형 당뇨병 쥐 모델에서 유전공학적 제조 K-세포 이식을 통한 당뇨병의 치료

심주연 ( Ju Yeon Sim ),김주희 ( Ju Hee Kim ),안유배 ( Yu Bae Ahn ),송기호 ( Ki Ho Song ),한제호 ( Je Ho Han ),차봉연 ( Bong Yun Cha ),이숙경 ( Sook Kyung Lee ),문성대 ( Sung Dae Moon ) 대한당뇨병학회 2009 Diabetes and Metabolism Journal Vol.33 No.6

연구배경: 인슐린 유전자 치료법의 이상적인 표적세포로 K-세포가 알려져 있다. 이전 연구에서 본 연구자들은 EBV-유래 에피솜 벡터를 이용하여 K-세포에서 포도당농도 의존적인 인슐린 분비가 이루어지는 것을 실험실 환경에서 확인한 바 있다. 본 연구에서는 스트렙토조토신(STZ)으로 유발된 제1형 당뇨병 쥐의 신장 피막에 유전공학적으로 제조된 K-세포를 이식하여 당뇨병이 치료되는지를 관찰해 보았다. 방법: 인슐린이 분비되도록 제조된 K-세포를 STZ으로 유도된 BALB/c Nude 마우스의 신장 피막에 이식한 후 혈당과 몸무게를 측정하였다. 마우스는 정상군과 STZ으로 유도된 당뇨병군 그리고 당뇨병군의 일부는 K-세포를 이식한 치료군 등으로 분류하여 실험을 하였다. 이식 4주 후 모든 마우스에서 6시간 동안 금식한 후 2 g/kg의 포도당을 복강내로 주사하여 당내성검사를 하였다. 당내성검사가 끝나자마자 모든 마우스의 신장과 췌장을 적출하여 인슐린, 글루카곤, C-peptide 등으로 면역조직화학염색 및 면역형광염색을 한 후 그 결과를 분석하였다. 결과: STZ을 복강에 주사한 마우스는 주사 3일 후 혈당이 300±50 mg/dL로 상승하였다. 정상군과 K-세포를 이식한 당뇨병군에서는 몸무게가 점차 증가하였으나, STZ만을 주사한 당뇨병군에서는 실험기간 동안 낮게 유지되었다. 그리고 정상군에서는 정상범위의 혈당분포를 유지하였으나, STZ만을 주사한 당뇨병군에서는 지속적으로 혈당이 높게 유지되었다. 그리고 K-세포를 이식한 당뇨병군에서는 STZ만을 주사한 군에서보다 혈당이 서서히 떨어졌으며 이식 2주 후부터는 급격히 감소하였다. 당부하검사 결과 정상군과 K-세포를 이식한 당뇨병군에서는 서로 유사한 내당력을 보였으나 STZ만을 주사한 군에서는 지속적으로 포도당 불내성을 보였다. 조직면역검사 결과 정상군의 췌장에서는 인슐린과 글루카곤, C-peptide 모두에 염색이 되었지만 STZ만을 주사한 마우스의 췌장에서는 췌도파괴로 인슐린은 물론 글루카곤, C-peptide 모두에 염색이 되지 않았다. 그러나, K-세포를 이식한 당뇨병군의 신장에서는 면역조직화학염색에서 인슐린과 C-peptide 모두에서 염색이 되었으며 면역형 광염색에서도 인슐린이 발현하는 것을 관찰할 수 있었다. 결론: 유전공학적으로 제조된 K-세포가 마우스 신장 피막에 착상되어 혈당농도 의존적으로 인슐린을 분비하여 당뇨병이 치료되는 것을 생체 내에서도 확인할 수 있었으며, EBV-유래 에피솜벡터를 이용한 유전공학적 조제 K-세포는 제1형 당뇨병의 치료를 위한 대체베타세포로 사용될 수 있을 것으로 생각된다. Background: K-cells function as targets for insulin gene therapy. In a previous study, we constructed EBV-based plasmids expressing rat preproinsulin controlled by glucose-dependent insulinotropic polypeptide promoters. In the present study, we attempted to correct hyperglycemia in vivo using genetically engineered K-cells in a mouse model of type 1 diabetes. Methods: K-cells expressing insulin were transplanted under the kidney capsules of STZ-induced diabetic mice. The blood glucose levels and body weights of the experimental animals were measured daily. After four weeks, the mice were injected intra-peritoneally with 2 g/kg glucose following a 6 hr fast. Blood glucose levels were measured immediately following glucose injections. All animals were sacrificed at the end of the glucose tolerance study, and pancreas and graft-bearing kidney tissue samples were stained with antibodies against insulin, glucagon, and C-peptide. Results: The body weights of K-cell-transplanted diabetic mice increased after transplantation, whereas those of untreated diabetic control mice continued to decline. The blood glucose levels of K-cell-transplanted diabetic mice decreased gradually during the two weeks following transplantation. After intra-peritoneal injection of glucose into K-cell-transplanted diabetic mice, blood glucose levels increased at 30 minutes, and were restored to the normal range between 60 and 90 minutes, while untreated control diabetic mice continued to experience hyperglycemia. Kidney capsules containing transplanted K-cells were removed, and sections were stained with anti-insulin antibodies. We detected insulin-positive cells in the kidney capsules of K-cell-transplanted diabetic mice, but not in untreated control mice. Conclusion: We detected glucose-dependent insulin secretion in genetically engineered K-cells in a mouse model of type 1 diabetes. Our results suggest that genetically modified insulin producing K-cells may act as surrogate β-cells to effectively treat type 1 diabetes. (Korean Diabetes J 33:466-474, 2009)

Mutation in Ret Proto - Oncogene in Medullary Thyroid Cancer and Multiple Endocrine Neoplasia Type 2A ( Men 2A ) and Men 2B with Hirschcprung`s Disease

유순집(Soon Jib Yoo),이희진(Hee Jin Lee),안유배(Yu Bae Ahn),한제호(Je Ho Han),윤건호(Kun Ho Yoon),강무일(Moo Il Kang),차봉연(Bong Yun Cha),이광우(Kwang Woo Lee),손호영(Ho Young Son),강성구(Sung Koo Kang),김영실(Young Sill Kim),김병기(By 대한내과학회 1998 대한내과학회 추계학술대회 Vol.55 No.-

그레이브스 병에서 항갑상성제 투여중 발생한 갑상성기능저하증의 치료

송기호(Ki Ho Song),김병수(Byung Su Kim),장상아(Sang A Jang),안유배(Yu Bae Ahn),한제호(Je Ho Han),유순집(Soon Jip Yoo),이종민(Jong Min Lee),손현식(Hyun Sik Son),윤건호(Kun Ho Yoon),감무일(Moo Il Kang),차봉연(Bong Yun Cha),이광우(Kwang Wo 대한내과학회 1996 대한내과학회지 Vol.51 No.3

N/A Objectives : Antithyroid drug is highly effective and safe in the treatment of Graves` disease. However, some patients may be rendered hypothyroid by large dosage of andithyroid drug : at this point reduction of dosage is common but often difficult to quantitate. Alternatively, combination therapy of antithyroid drug with L-thyroxine is often used, but there are few reports on its long-term effectiveness. Methods : 43 patients with Graves` disease who developed hypothyroidism during methimazole treatment were studied. They received combination therapy of 100mg methimazole with 50 ㎍(group 1, n=16) or 100 ㎍(group 2, n=27) L-thyroxine per day. Hypothyroidism was defined as elevation of seruum thyroid-stimulating hormone (TSH) concentrations above normal ranges. Results : 1) Hypothyroidism occurred within 5.8±3.7(mean ± SD) months after methimazole treatment and mean doses of methimazole were 22,1±8.4mg per day. 2) Serum total thyroxine concentrations increased significantly from 3.7±2,4㎍/dl in the hypothyroid stage to 9.4+3.0 ㎍/dl after 2 months of combination therapy(p<0.0001). 3) Serum TSH concentrations decreased significantly from 34.1+30.6μU/ml in the hypothyroid stage to 3.4+5.2 μU/ml after 2 months of combination therapy(p<0.0001). 4) After 6 months of combination therapy, 74.2Yo of patients became euthyroid. 5) There was no significant difference in the frequency of euthyroid patients after combination therapy betwen group 1 and group 2. Conclusion: These results suggest that long- term combination therapy of antithyroid drug with L-thyroxine in small fixed doses could be effective and convenient to maintain euthyroidism in patients with Graves' disease who developed hypothyroidism during antithyroid drug treatment.

사람의 부분 췌장으로부터 분리한 췌관세포를 이용한 베타세포의 분화에 관한 연구

송기호 ( Ki Ho Song ),김명미 ( Myung Mee Kim ),이민경 ( Min Kyung Lee ),류경렬 ( Gyeong Ryul Ryu ),고승현 ( Seung Hyun Ko ),문성대 ( Sung Dae Moon ),안유배 ( Yu Bae Ahn ),윤건호 ( Kun Ho Yoon ),차봉연 ( Bong Yun Cha ),이광우 ( Kwa 대한당뇨병학회 2007 Diabetes and Metabolism Journal Vol.31 No.3

Pyloric Septum

박두호(Doo Ho Park),최상욱(Sang wook Choi),양진모(Jin Mo Yang),서은주(Eun Joo Seo),조현미(Hyun Mee Cho),김희제(Hee Je Kim),안유배(Yu Bae Ahn),이숙영(Suk Young Lee),변재영(Jae Young Byun) 대한소화기학회 1991 대한소화기학회지 Vol.23 No.3

Double pylorus is one of the rare anomalies of the intestinal tract. The medical literature has had descriptions of double pylorus, pyloric septum, pyloric band, and pyloric duplication as synonyms by a band of varying width. True double pylorus has the second channel been complete, resulting in a double pyloric canal, with no other malformation. It is widely recognized that true double pylorus is usually an acquired lesion as the end results of duodenal or gastric ulcers which upon undermining normal tissue, recommunicate with either the lumen of the duodenum or the stomach itself. The length of the undermined area 5etermines of a septum or a double pylorus is formed. We wish to report here a case of pyloric septum diagnosed endoscopically in a patient with shallow gastric ulcer. A 53-year-old man was found to have pyloric septum during examinations for long lasting recurrent epigastric pain. We think that the formation of pyloric septum in this case resulted from recurrent peptic ulcer.

심근염과 완전 방실차단 및 췌장염이 합병된 장티푸스 1예

안유배,김양리,유진홍,신완식,강문원 대한감염학회 1993 감염 Vol.25 No.3

The report of completer atrioventricular (AV) block wit myocarditis complicating typhoid fever is a rare occurrence. The patient was a 30-year-old woman and experienced fever, chill and abdominal pain two weeks earlier followed by mental confusion. Typhoid fever was confirmed by islolation of S. typhi in blood and stool. Laboratory examination showed the elevation of cardiac enzymes and serum amylase level. On the fifth hospital day, complete AV block developed transiently and disappeared without any intervention. The patient was treated with dexamethasone and a two-week course of chloramphenicol. We report a case of typhoid fever complicated by complete AV block, myocarditis and pancreatitis.

갈색세포종 제거후 성공적으로 당뇨병이 완치된 1예

홍창균,안유배,김설혜,우영식,이성구,고승현,송기호,윤건호,강무일,차봉연,이광우,손호영,강성구 대한내분비학회 2001 Endocrinology and metabolism Vol.16 No.4-5

갈색세포종 제거후 성공적으로 당뇨병이 완치된 1예

홍창균,안유배,김설혜,우영식,이성구,고승현,송기호,윤건호,강무일,차봉연,이광우,손호영,강성구 대한내분비학회 2001 Endocrinology and metabolism Vol.16 No.4

Pheochromocytoma is usually associated with a combination of various clinical manifestations caused by the overproduction of catecholamines. It is frequently accompanied by impaired glucose tolerance operating through 2 - and - adrenergic mechanisms. A 41-year-old-woman was admitted to the hospital because of poorly-controlled diabetes mellitus and hypertension. She had suffered intermittent paroxysmal attacks of headache and chest discomfort and had been treated intermittently over a 2 year period for diabetes mellitus and hypertension. At admission, the levels of serum epinephrine, norepinephrine urinary excretion of total metanephrine, and VMA were all abnormally elevated. Adrenal CT showed a well-defined, homogenous mass in the right adrenal region and the tumor was diagnosed as pheochromocytoma. After tumor resection, the increased blood level of catecholamines, the urinary excretion of total metanephrine, and VMA were normalized, as was the hyperglycemia state. Diabetes mellitus of the patient was considered permanently resolved after tumor removal by the result of glucose tolerance in 75 g oral glucose tolerance test

분만 후 심한 저혈당이 동반된 림프구성 뇌하수체염 1예의 경과 관찰

손장명,고승현,안유배,주강,이정록,양성은,송기호,손호영,전정수 대한내분비학회 2003 Endocrinology and metabolism Vol.18 No.3

저자들은 출산 후 발생한 심한 저혈당을 주소로 내원한 31세 여자 환자에서 뇌 자기공명영상검사에서 뇌하수체종괴가 관찰되고 복합 뇌하수체 기능검사 결과전반적인 뇌하수체 기능저하증의 소견을 보여 림프구성 뇌하수체염을 진단하고, 호르몬 보충요법만으로 경과를 관찰하여 호전된 예를 경험하였기에 문헌 고찰과 함께 보고하는 바이다. Lymphocytic hypophysitis, an inflammatory disease of the pituitary gland that is often associated with pregnancy, is caused by autoimmune destruction of the pituitary gland evidenced by diffuse inflammatory cell infiltration of the pituitary gland and some kinds of detectable autoantibodies. We report a case of lymphocytic hypophysitis in a 31-year-old woman presenting with severe hypoglycemia and hyponatremia after delivery. Hormonal study revealed panhypopituitarism and magnetic resonance imaging with enhancement showed the bulging contour of the right side pituitary gland with an ill-defined mass-like lesion and nodular thickening of the stalk The patient's symptoms and biochemical data improved greatly with replacement of L-thyroxine and glucocorticoid. Partial recovery of panhypopituitarism was also seen. The follow-up tests revealed dramatic resolution of the pituitary lesion (J Kor SOC Endocrinol 1 8 : 3 25 -33 1, 2003).