Dietary oleate has beneficial effects on every step of non-alcoholic Fatty liver disease progression in a methionine- and choline-deficient diet-fed animal model.

BACKGROUND: Non-alcoholic fatty liver disease (NAFLD) is increasingly recognized as a major cause of liver-related morbidity and mortality. The underlying mechanisms of disease progression remain poorly understood, and primary therapy of NAFLD is not yet established. We investigated the effects of dietary oleate on the development and progression of NAFLD in a methionine- and choline-deficient (MCD) diet-fed animal model. METHODS: A total of 30 C57BL/6J mice were randomly divided into three groups (n=10 in each group) and fed various experimental diets for four weeks: chow, MCD diet, or OMCD (MCD diet with oleate, 0.5 mg/g/day). Liver samples were examined for steatohepatitis and fibrosis parameters and associated genes. RESULTS: Additional dietary oleate dramatically reduced MCD diet-induced hepatic steatosis. Hepatic carbohydrate responsive element-binding protein was overexpressed in MCD diet-fed mice, and dietary oleate prevented this overexpression (P<0.001). Dietary oleate partially prevented MCD diet-induced serum level increases in aspartate aminotransferase and alanine aminotransferase (P<0.001, respectively). The mRNA expressions of hepatic monocyte chemoattractant protein 1, tumor necrosis factor-α and matrix metalloproteinase-9 were increased in MCD diet-fed mice, and this overexpression of inflammatory molecules was prevented by dietary oleate (P<0.001). Hepatic pericellular fibrosis was observed in MCD diet-fed mice, and dietary oleate prevented this fibrosis. Altogether, dietary oleate prevented MCD diet-induced hepatic steatosis, inflammation and fibrosis. CONCLUSION: Dietary oleate has beneficial effects in every step of NAFLD development and progression and could be a nutritional option for NAFLD prevention and treatment.ope

A Case of Cephalosporin-induced DRESS (Drug Rash with Eosinophilia and Systemic Symptoms) Syndrome with Acute Renal Failure

Drug rash with eosinophilia and systemic symptoms (DRESS) syndrome, a kind of hypersensitivity syndrome, is a life-threatening condition with various clinical manifestations such as fever, skin rash, lymph node enlargement and internal organ involvement. We report a case of skin rash and eosinophilia followed by acute renal failure and mild hepatitis after cephalosporin administration.ope

로지글리타존이 간의 LRP1 발현에 미치는 영향: 티아졸리딘디온 제제가 당뇨병에서 발생하는 죽상경화성 지질대사이상을 호전시키는 새로운 기전

Dept. of Medicine/박사Hepatic low-density lipoprotein receptor-related protein 1 (LRP1) plays an important role in the clearance of circulating remnant lipoproteins. In this study, the effect of thiazolidinedione, an insulin sensitizing drug on the expression and function of hepatic LRP1 was investigated. This study may present a novel mechanism of the improvement of atherogenic dyslipidemia by a thiazolidinedione drug in diabetic patients. For in vitro evaluation, HepG2 cells were treated with various concentrations of rosiglitazone. For in vivo study, male Otsuka Long-Evans Tokushima Fatty (OLETF) rats and Long-Evans Tokushima Otsuka (LETO) rats were used as a diabetic animal model and its normal counterpart respectively. Rats were treated with rosiglitazone for 5 weeks. The expression and function of LRP1 in HepG2 cells and liver samples of rats were analyzed.LRP1 mRNA and protein expressions were increased by 0.5 and 5 μM of rosiglitazone in HepG2 cells. However, the expression of LRP1 did not change compared to that in non-treated cells at concentrations above 50 μM of rosiglitazone. The electrophoretic mobility shift assay and the reporter assay showed that rosiglitazone increased the transcriptional activity of the LRP1 promoter by binding peroxisome proliferator-activated receptor-γ to a perxisome proliferator response element of the LRP1 promoter in HepG2 cells. The rosiglitazone-induced up-regulation of the LRP1 promoter activity showed the same tendency with the LRP1 expression. The uptake of ApoE through LRP1 in HepG2 cells was also increased by rosiglitazone. The serum triglyceride level was increased in OLETF rats compared to that in LETO rats and partially recovered with rosiglitazone treatment. Hepatic LRP1 was reduced in OLETF rats compared to that in LETO rats and rosiglitazone treatment increased the hepatic LRP1 in OLETF rats. A high-glucose condition (25 mM of glucose in culture media) reduced the expression of LRP1 in HepG2 cells, and this reduced LRP1 expression was recovered with rosiglitazone.In conclusion, our data suggest that decreased hepatic LRP1 in a diabetic condition is associated with the development of atherogenic dyslipidemia and that increased hepatic LRP1 by thiazolidinediones contributes to an improvement in atherogenic lipid profiles in diabetic patients.ope

Long term, serial effect of pioglitazone on anthropometrics and metabolic parameters in patients with type 2 di

의학과/석사[한글] 제 2형 당뇨병의 병인기전은 인슐린 저항성 및 인슐린 분비능의 저하로 알려져 있다. 본 연구는 인슐린 저항성을 호전시키는 것으로 알려진 티아졸리딘디온 제제인 피오글리타존 제제를 1년 이상 투여한 한국인 제 2형 당뇨병 환자군을 대상으로 한 코호트연구를 통해, 투여기간에 따른 신체계측 및 대사지표의 변화를 관찰하고 피오글리타존 제제 투여에 높은 효과를 보이는 환자군의 특성을 알아보고자 하였다.2003년 3월부터 연세대학교 세브란스병원에 내원하는 113명의 제 2형 당뇨병 환자에게 피오글리타존 제제를 투여하면서 전향적 코호트 연구를 수행하였다. 대상 환자군을 투여 전, 투여 후 3개월, 6개월, 12개월에 추적 관찰하여 신체계측 및 대사지표를 측정하였다.대상 환자군에서 피오글리타존 제제 투여 3개월 후 체중 및 제질량지수의 증가가 나타났고(68.8±12.2 vs 69.8±11.9kg, p<0.01) 이후 지속적인 체중 및 체질량지수의 증가는 보이지 않았으며 이러한 체중 및 체질량지수의 증가는 여자에서 더 크게(3개월 체중변화, 0.6±1.7kg vs 1.6±1.7kg, p<0.01), 더 오랜 기간(3개월 vs 6개월) 나타났다. 피오글리타존 제제의 혈당 강하 효과 역시 투여 후 3개월에 나타났고(공복혈당, 143.4±41.6 vs 125.3±36.3 mg/dL, p<0.01; 당화혈색소, 7.7±1.5 vs 7.0±1.1%, p<0.01), 남자보다 여자에서 그 효과가 더 크게 나타났다(공복혈당, -1.6±31.0 vs -32.4±45.8 mg/dL, p<0.001; 당화혈색소, -0.4±1.1 vs -0.9±1.3%, p<0.05). 고밀도지단백-콜레스테롤 수치가 피오글리타존 제제 투여 후 3개월에 증가하여(45.0±9.2 vs 50.6±10.8 mg/dL, p<0.01) 지속적으로 유지되었고 중성지방은 투여 후 3개월에 감소하였다가(205.4±166.7 vs 166.3±107.3 mg/dL, p<0.01) 다시 증가하였다. 피오글리타존 제제 투여 후 간 효소 수치의 감소도 관찰되었으며, 특히 alanine transaminase 수치는 투여 후 3개월에 감소하여(30.8±23.7 vs 24.5±18.5 IU/L, p<0.01) 이후 유지되었다. 피오글리타존 투여 후 혈당 강하 효과는 투여 전 체질량지수 및 체지방률이 높을수록 크게 나타났다.결론적으로 한국인 제 2형 당뇨병 환자에서 장기간 피오글리타존 제제의 투여시 혈당 강하, 지질 대사 개선 및 간 기능 개선에 있어 우수한 임상 경과를 보였고 체중 증가는 제한적으로 나타났다. [영문]Type 2 diabetes is characterized by impaired insulin secretion and/or insulin resistance. Thiazolidindiones have been shown to ameliorate insulin resistance. The purpose of the present study was to evaluate the long term serial effect of pioglitazone on anthropometrics and metabolic parameters in Korean type 2 diabetes patients.One hundred thirteen type 2 diabetes patients (male, 67; female, 46; mean age, 49.1 years) were studied before and after 3 months, 6 months and 12 months of treatment with pioglitazone. anthropometric parameters and metabolic parameters were measured.Body weight and body mass index (BMI) were increased in 3 months after pioglitazone treatment (68.8 ± 12.2 vs 69.8 ± 11.9 kg, p < 0.01) without continuouos increase. In female patients, body weight and BMI tend to increase more (0.6 kg vs 1.6 kg, p < 0.01) and longer (3 months vs 6 months) than male patients. Fasting plasma glucose (FPG) and glycosylated hemoglobin (HbA1c) were decreased in 3 months after pioglitazone treatment (FPG, 143.4 ± 41.6 vs 125.3 ± 36.3 mg/dL, p < 0.01; HbA1c, 7.7 ± 1.5 vs 7.0 ± 1.1 %, p < 0.01). Hypoglycemic effect of pioglytazone was prominent in female patients. High density lipoprotein cholesterol was increased after pioglitazone treatment (45.0 ± 9.2 vs 50.6 ± 10.8 mg/dL, p < 0.01). Triglyceride was decreased until 6 months after pioglitazone treatment (205.4 ± 166.7 vs 166.3 ± 107.3 mg/dL, p < 0.01), and recovered at 12 months after. Liver enzyme, especially alanine transaminase decreased after pioglitazone treatments (30.8 ± 23.7 vs 24.5 ±18.5 IU/L, p < 0.01). Hypoglycemic effect of pioglitazone was associated with basal BMI, fat contents and serum leptin level.Korean type 2 diabetic patients with pioglitazone use show good clinical course for glycemic control, lipid metabolism and liver function, but body weight increase by pioglitazone is limited.ope

제조공정 최적화를 통한 무세포 백일해 백신 정제공정 개발

학위논문(박사) - 한국과학기술원 : 생명과학과, 2024.2,[iv, 82 p. :]Pertussis, also known as whooping cough, is a highly contagious respiratory disease transmitted directly between humans. Pertussis is caused by the bacterium Bordetella pertussis and can affect people of all ages, with serious or even deadly outcomes for babies under one year of age. The whole-cell pertussis (wP) vaccine developed in the late 1940s protected against pertussis and rapidly reduced incidence. However, adverse reactions such as febrile convulsions and , very rarely, hypotonic-hyporesponsive episodes were observed at a high frequency after administration of the wP vaccine, underlining the need for a safer pertussis vaccine. Acellular pertussis (aP) vaccine composing thress components including such as pertussis toxin (PT), filamentous hemagglutinin (FHA), and pertactin (PRN) from B. pertussis, is safer and less reactogenic than the wP vaccine. However, due to difficulties in the manufacturing process of acellular pertussis vaccines, only two acellular pertussis vaccines are approved and sold. The purpose of this study is to develop a manufacturing process that overcomes the major technical limitations of the PT, FHA, and PRN antigen manufacturing process. Three types of antigens can be obtained by Bodetella pertussis cell culture, PRN can be obtained from cell pellets and PT and FHA can be obtained from culture supernatant. PRN, a non-fibrial outer membrane protein of Bordetella pertussis, is a limiting component of cell-free pertussis vaccine due to its low expression, so the study of PRN manufacturing process was conducted with the main goal of improving productivity. We compared the extraction of PRN from the cell membrane using urea with the commonly used heating method and found that more membrane proteins were extracted using the urea method. Through the study of various conditions, it was found that the cell pellet processing conditions, including freezing and thawing, substantially affect the PRN yield. Finally, a single cycle of rapid freezing of the cell pellet with slow thawing resulted in up to 5-fold higher PRN yields than conditions without freezing and thawing. We also explored urea treatment conditions and found the optimal conditions. After extraction, residual impurities were removed by sequential anion exchange, hydrophobic interaction, and gel filtration chromatography to obtain highly purified PRN antigen with a purity of more than 95%, thus establishing a PRN manufacturing process. PT and FHA are expressed in relatively large amounts and are obtained from the culture supernatant. Due to its strong toxicity, PT must undergo a detoxification process after the purification process. Because only PT undergoes detoxification process, cross-contamination of PT and FHA may cause safety concerns during vaccine manufacturing. So it is necessary to completely separate PT and FHA, and conditions capable of completely separating the two antigens were established through optimization of affinity column chromatography conditions. The design of experiment(DoE) method was used to optimize the conditions, and a design space capable of stably isolating antigens was selected. Through sequential hydroxyapatite, hydrophobic interaction, and affinity chromatography processes, PT and FHA antigens with a purity of more than 95% were obtained. Scale-up study was conducted up to 200 L scale and reproducible results are confirmed. Acellular pertussis vaccine for adults was prepared by mixing three types of PT, FHA, and PRN antigens produced from a 200 L scale with diphtheria toxoid and tetanus toxoid, and its safety and efficacy were confirmed through non-clinical and clinical studies. In conclusion, an integrated acelluar pertussis vaccine purification process for three types of antigens from Bodetella pertussis culture was developed. The membrane protein extraction process using urea developed in this study may also serve as a reference for manufacture of other membrane proteins and the methodology for finding antigen separation using affinity chromatography can be applied to the production of a variety of proteins. In addition, this pertussis antigen manufacturing method will be applicable to the development of various combination vaccines based on acellular pertussis.한국과학기술원 :생명과학과

On-line measurement and control of methanol concentration using alcohol sensor in fed-batch fermentation

학위논문(석사) - 한국과학기술원 : 생물과학과, 2003.2, [ [v], 50 p. ]한국과학기술원 : 생물과학과

The effect of rosiglitazone on LRP1 expression and amyloid β uptake in human brain microvascular endothelial cells: a possible role of a low-dose thiazolidinedione for dementia treatment.

Thiazolidinediones, such as rosiglitazone or pioglitazone, are anti-diabetic agents that have been expected to show a beneficial effect in Alzheimer's disease (AD) because of their anti-inflammatory effect. However, these agents have failed to show a significant beneficial effect on AD in recent clinical trials. Here, we suggest that low-dose rosiglitazone treatment, and not the conventional doses, has an amyloid β (Aβ)-clearing effect by increasing LRP1, an Aβ outward transporter in the blood-brain barrier. Rosiglitazone up-regulated LRP1 mRNA and protein expression and LRP1 promoter activity in human brain microvascular endothelial cells (HBMECs). Aβ uptake through LRP1 in HBMECs was also increased by rosiglitazone. This increase in LRP1 and Aβ uptake was observed in up to 10 nm rosiglitazone concentration. At concentrations above 20 nm rosiglitazone, the LRP1 expression and Aβ uptake in HBMECs were not altered. The possible mechanism of this unusual dose response is discussed. This study suggests a new therapeutic application of thiazolidinediones for AD at a much lower dose than the doses used for diabetes treatment.ope

Decreased endothelial progenitor cells and increased serum glycated albumin are independently correlated with plaque-forming carotid artery atherosclerosis in type 2 diabetes patients without documented ischemic disease.

BACKGROUND: The aim of the present study was to investigate the serum levels of endothelial progenitor cells (EPCs) in type 2 diabetic patients without documented ischemic disease and the association between EPCs and atherosclerotic plaque formation in the carotid artery. METHODS AND RESULTS: A clinic-based, prospective study of type 2 diabetic patients was conducted. A total of 73 subjects were enrolled in this study after cardiac magnetic resonance imaging and ankle-brachial index measurements to exclude patients with ischemic disease. Plaque formation in the carotid artery was measured on ultrasonography. Circulating EPCs (CD34(+)/CD133(+)/CD309(+) cells) were counted on flow cytometry. Compared to subjects without carotid artery plaques, patients with plaques were significantly older (P=0.006) and had decreased EPC count (P=0.027). Serum glycated albumin (GA) level and the GA/glycated hemoglobin ratio tended to decrease in patients with plaques (P=0.091 and 0.067, respectively). Other cardiovascular disease risk factors were not significantly different between the 2 groups. On binary logistic regression analysis old age, low EPC count, and high serum GA level were independently correlated with carotid artery plaque formation. CONCLUSIONS: EPC count and serum GA level appear to be a protective and an aggravating factor for endothelial damage, respectively, and therefore, a reduced EPC count or an increased GA level results in atherosclerotic plaque formation in type 2 diabetic patients.ope

Up-regulation of hepatic low-density lipoprotein receptor-related protein 1: a possible novel mechanism of antiatherogenic activity of hydroxymethylglutaryl-coenzyme A reductase inhibitor Atorvastatin and hepatic LRP1 expression

Low-density lipoprotein receptor-related protein 1 (LRP1) binds to apolipoprotein E and serves as a receptor for remnant lipoproteins in the liver, thus playing an important role in clearing these atherogenic particles. In this study, we investigated the effect of atorvastatin, a hydroxymethylglutaryl-coenzyme A reductase inhibitor, on hepatic LRP1 expression. We used HepG2 and Hep3B cells for in vitro study, and Otsuka Long-Evans Tokushima fatty and Sprague-Dawley rats for in vivo study. We used relatively high pharmacologic dose of atorvastatin in this study (in vitro, 0.5 μmol/L in culture media, for 48 hours; in vivo, 20 mg/[kg d], for 6 weeks). Atorvastatin increased LRP1 and low-density lipoprotein (LDL) receptor expression in HepG2 and Hep3B cells and induced hepatic LRP1 and LDL receptor expression in chow diet-fed Sprague-Dawley rats and high-fat diet-fed Otsuka Long-Evans Tokushima fatty rats. Atorvastatin decreased intracellular sterol level and increased the amount of the nuclear form of sterol response element-binding protein-2 (SREBP-2) in both HepG2 and Hep3B cells as well as in two animal models. Treatment of HepG2 cells with LDL increased intracellular sterol level and reduced LRP1, LDL receptor, and SREBP-2. When SREBP-2 in HepG2 cells was knocked down by small interfering RNA, the induction of LRP1 expression by atorvastatin did not take place. In conclusion, up-regulation of hepatic LRP1 might be a novel mechanism by which statin treatment decreases remnant lipoproteins. In addition, SREBP-2 acts as a mediator of atorvastatin-induced up-regulation of hepatic LRP1. Future studies using standard doses of atorvastatin in humans are needed to elucidate clinical relevance of these findings.ope

Fat redistribution preferentially reflects the anti-inflammatory benefits of pioglitazone treatment.

Thiazoledinedione is known to have an anti-inflammatory effect besides a hypoglycemic effect. We investigated changes in high-sensitivity C-reactive protein (hsCRP), a proinflammatory marker, after pioglitazone treatment in association with the resulting changes in various metabolic and anthropometric parameters. A total of 93 type 2 diabetes mellitus patients (47 men and 46 women; mean age, 50.0 ± 10.8 years) who were being treated with a stable dose of sulfonylurea or metformin were enrolled in the study. Pioglitazone (15 mg/d) was added to their treatment regimen for 12 weeks, and metabolic and anthropometric measurements were taken before and after pioglitazone treatment. Pioglitazone treatment for 12 weeks decreased serum hsCRP levels (0.83 [1.14] to 0.52 [0.82] mg/L, P < .001) and improved glycemic control (fasting glucose, P < .001; glycosylated hemoglobin, P < .001) and lipid profiles (triglyceride, P = .016; high-density lipoprotein cholesterol, P < .001). Between responders and nonresponders to the hsCRP-lowering effect of pioglitazone, there were significant differences in baseline hsCRP levels and changes in the postprandial glucose and the ratio of visceral fat thickness (VFT) to subcutaneous fat thickness (SFT) (P = .004, .011, and .001, respectively). The percentage change in hsCRP levels after treatment was inversely correlated with baseline hsCRP levels (r = -0.497, P < .001) and directly correlated with the change in postprandial glucose (r = 0.251, P = .021), VFT (r = 0.246, P = .030), and VFT/SFT ratio (r = 0.276, P = .015). Logistic regression analysis revealed that the hsCRP-lowering effect of pioglitazone was affected by baseline hsCRP levels (odds ratio [OR] = 7.929, P = .007) as well as changes in postprandial 2-hour glucose (OR = 0.716, P = .025) and VFT/SFT ratio (OR = 0.055, P = .009). In conclusion, treatment with pioglitazone produced an anti-inflammatory effect, decreasing serum hsCRP levels; and a decrease in the VFT/SFT ratio was independently and most strongly associated with the hsCRP-decreasing effect. These results suggest that abdominal fat redistribution preferentially reflects the anti-inflammatory benefits of pioglitazone treatment.ope