Diabetes & Metabolism Journal

Original Article

The Association of Serum Cystatin C with Glycosylated Hemoglobin in Korean Adults: Eun Hee Sim, Hye Won Lee, Hyun Ju Choi, Dong Wook Jeong, Seok Man Son, Yang Ho Kang; Diabetes Metab J. 2016;40(1):62-69. Published online November 27, 2015; DOI: https://doi.org/10.4093/dmj.2016.40.1.62

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Background

Cystatin C has been known to be associated not only with early renal impairment but also with the incidence of diabetic conditions (prediabetes plus diabetes). However, it is not clear whether cystatin C levels are associated with the prevalence of diabetic conditions in Asian populations. We evaluated this association using glycosylated hemoglobin (HbA1c) levels as the definition of diabetes in Korean adults.

Methods

We analyzed data from 1,559 Korean adults (937 men and 622 women) with available serum cystatin C and HbA1c values.

Results

The serum cystatin C levels in subjects with prediabetes and diabetes were significantly increased (0.91±0.14 mg/L in prediabetes and 0.91±0.17 mg/L in diabetes vs. 0.88±0.13 mg/L in patients with normal glucose levels, P=0.001). At increasing cystatin C levels, the prevalence of subjects with prediabetes (30.2% vs. 14.6%, P<0.001) and those with diabetes (10.6% vs. 8.0%, P<0.001) significantly increased in the group with the highest cystatin C levels. The group with the highest cystatin C levels had a significantly increased odds ratio (OR) for the presence of diabetic conditions compared to the group with the lowest values in total subjects (OR, 2.35; 95% confidence interval [CI], 1.54 to 3.58; P<0.001) and in women (OR, 4.13; 95% CI, 1.97 to 8.65; P<0.001), though there was no significant increase after adjusting for multiple variables.

Conclusions

Higher levels of serum cystatin C are associated with an increased prevalence of diabetic conditions in Korean adults. Our findings may extend the positive association of cystatin C with diabetes incidence to an Asian population.

Citations

Citations to this article as recorded by

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Detection of decline in estimated glomerular filtration rate in patients with type 2 diabetes by cystatin C-based equations
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Cystatin C as a Predictor for Diabetes according to Glycosylated Hemoglobin Levels in Korean Patients
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Diabetes & Metabolism Journal.2016; 40(1): 32. CrossRef
Prevalence of Reduced Kidney Function by Estimated Glomerular Filtration Rate Using an Equation Based on Creatinine and Cystatin C in Metabolic Syndrome and Its Components in Korean Adults
Yang Ho Kang, Dong Wook Jeong, Seok Man Son
Endocrinology and Metabolism.2016; 31(3): 446. CrossRef
Letter: The Association of Serum Cystatin C with Glycosylated Hemoglobin in Korean Adults (Diabetes Metab J 2016;40:62-9)
Kyung-Soo Kim
Diabetes & Metabolism Journal.2016; 40(2): 171. CrossRef
Response: The Association of Serum Cystatin C with Glycosylated Hemoglobin in Korean Adults (Diabetes Metab J 2016;40:62-9)
Yang Ho Kang
Diabetes & Metabolism Journal.2016; 40(2): 173. CrossRef

Letter

Letter: Diabetic Retinopathy and Endothelial Dysfunction in Patients with Type 2 Diabetes Mellitus (Diabetes Metab J 2013;37:262-9): Seok Man Son; Diabetes Metab J. 2013;37(5):391-392. Published online October 17, 2013; DOI: https://doi.org/10.4093/dmj.2013.37.5.391

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Editorial

C-Peptide and Vascular Complications in Type 2 Diabetic Subjects: Seok Man Son; Diabetes Metab J. 2012;36(5):345-349. Published online October 18, 2012; DOI: https://doi.org/10.4093/dmj.2012.36.5.345

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Citations

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The Correlation Between C-Peptide and Severity of Peripheral Atherosclerosis in Type 2 Diabetes Mellitus
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Review

Reactive Oxygen and Nitrogen Species in Pathogenesis of Vascular Complications of Diabetes: Seok Man Son; Diabetes Metab J. 2012;36(3):190-198. Published online June 14, 2012; DOI: https://doi.org/10.4093/dmj.2012.36.3.190

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Abstract PDF PubReader ePub

Macrovascular and microvascular diseases are currently the principal causes of morbidity and mortality in subjects with diabetes. Disorders of the physiological signaling functions of reactive oxygen species (superoxide and hydrogen peroxide) and reactive nitrogen species (nitric oxide and peroxynitrite) are important features of diabetes. In the absence of an appropriate compensation by the endogenous antioxidant defense network, increased oxidative stress leads to the activation of stress-sensitive intracellular signaling pathways and the formation of gene products that cause cellular damage and contribute to the vascular complications of diabetes. It has recently been suggested that diabetic subjects with vascular complications may have a defective cellular antioxidant response against the oxidative stress generated by hyperglycemia. This raises the concept that antioxidant therapy may be of great benefit to these subjects. Although our understanding of how hyperglycemia-induced oxidative stress ultimately leads to tissue damage has advanced considerably in recent years, effective therapeutic strategies to prevent or delay the development of this damage remain limited. Thus, further investigation of therapeutic interventions to prevent or delay the progression of diabetic vascular complications is needed.

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Original Articles

The Association between Apolipoprotein A-II and Metabolic Syndrome in Korean Adults: A Comparison Study of Apolipoprotein A-I and Apolipoprotein B: Dong Won Yi, Dong Wook Jeong, Sang Yeoup Lee, Seok Man Son, Yang Ho Kang; Diabetes Metab J. 2012;36(1):56-63. Published online February 17, 2012; DOI: https://doi.org/10.4093/dmj.2012.36.1.56

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Abstract PDF PubReader ePub

Background

Apolipoprotein A-II (apoA-II) is the second-most abundant apolipoprotein in human high-density lipoprotein and its role in cardio metabolic risk is not entirely clear. It has been suggested to have poor anti-atherogenic or even pro-atherogenic properties, but there are few studies on the possible role of apoA-II in Asian populations. The aim of this study is to evaluate the role of apoA-II in metabolic syndrome (MetS) compared with apolipoprotein A-I (apoA-I) and apolipoprotein B (apoB) in Korean adults.

Methods

We analyzed data from 244 adults who visited the Center for Health Promotion in Pusan National University Yangsan Hospital for routine health examinations.

Results

The mean apoB level was significantly higher, and the mean apoA-I level was significantly lower, in MetS; however, there was no significant difference in apoA-II levels (30.5±4.6 mg/dL vs. 31.2±4.6 mg/dL, P=0.261). ApoA-II levels were more positively correlated with apoA-I levels than apoB levels. ApoA-II levels were less negatively correlated with homocysteine and high sensitivity C-reactive protein levels than apoA-I levels. The differences in MetS prevalence from the lowest to highest quartile of apoA-II were not significant (9.0%, 5.7%, 4.9%, and 6.6%, P=0.279). The relative risk of the highest quartile of apoA-II compared with the lowest quartile also was not significantly different (odds ratio, 0.96; 95% confidence interval, 0.95 to 1.04; P=0.956).

Conclusion

Compared with apoA-I (negative association with MetS) and apoB (positive association with MetS) levels, apoA-II levels did not show any association with MetS in this study involving Korean adults. However, apoA-II may have both anti-atherogenic and pro-atherogenic properties.

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Interaction between Apo A-II –265T > C polymorphism and dietary total antioxidant capacity on some oxidative stress and inflammatory markers in patients with type 2 diabetes mellitus
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British Journal of Nutrition.2022; 128(1): 13. CrossRef
Dietary acid load modifies the effects of ApoA2–265 T > C polymorphism on lipid profile and serum leptin and ghrelin levels among type 2 diabetic patients
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High Glucose and/or Free Fatty Acid Damage Vascular Endothelial Cells via Stimulating of NAD(P)H Oxidase-induced Superoxide Production from Neutrophils.: Sang Soo Kim, Sun Young Kim, Soo Hyung Lee, Yang Ho Kang, In Ju Kim, Yong Ki Kim, Seok Man Son; Korean Diabetes J. 2009;33(2):94-104. Published online April 1, 2009; DOI: https://doi.org/10.4093/kdj.2009.33.2.94

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Abstract PDF: BACKGROUND
Oxidative stress and inflammation are important factors in the pathogenesis of diabetes and contribute to the development of diabetic complications. To understand the mechanisms that cause vascular complications in diabetes, we examined the effects of high glucose and/or free fatty acids on the production of superoxide from neutrophils and their role in endothelial cell damage. METHODS: Human neutrophils were incubated in the media containing 5.5 mM D-glucose, 30 mM D-glucose, 3 nM oleic acid, or 30 microM oleic acid for 1 hour to evaluate superoxide production through NAD(P)H oxidase activation. Human aortic endothelial cells were co-cultured with neutrophils exposed to high glucose and oleic acid. We then measured neutrophil adhesion to endothelial cells, neutrophil activation and superoxide production, neutrophil-mediated endothelial cell cytotoxicity and subunits of neutrophil NAD(P)H oxidase. RESULTS: After 1 hour of incubation with various concentrations of glucose and oleic acid, neutrophil adherence to high glucose and oleic acid-treated endothelial cells was significantly increased compared with adhesion to low glucose and oleic acid-treated endothelial cells. Incubation of neutrophils with glucose and free fatty acids increased superoxide production in a dose-dependent manner. High glucose and oleic acid treatment significantly increased expression of the membrane components of NAD(P)H oxidase of neutrophil (gp91(phox)). Endothelial cells co-cultured with neutrophils exposed to high glucose and oleic acid showed increased cytolysis, which could be prevented by an antioxidant, N-acetylcysteine. CONCLUSION: These results suggest that high glucose and/orfree fatty acidsincrease injury of endothelial cells via stimulating NAD(P)H oxidase-induced superoxide production from neutrophils.

Association of Serum Cystatin C with Metabolic Syndrome and Its Related Components in Korean Adults.: Sun Young Kim, Sang Heon Song, Yun Kyung Jeon, Ji Ryang Kim, Jung Ho Bae, Sang Soo Kim, Soo Hyung Lee, Seok Man Son, In Ju Kim, Yong Ki Kim, Yang Ho Kang; Korean Diabetes J. 2008;32(5):409-417. Published online October 1, 2008; DOI: https://doi.org/10.4093/kdj.2008.32.5.409

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Abstract PDF

BACKGROUND
Serum cystatin C has been reported as a better marker than serum creatinine for estimation of kidney function and may be associated with cardiovascular disease. The aim of this study was to elucidate the association of serum cystatin C with metabolic syndrome (MS), a constellation of cardiovascular risk factors, and its related components and the usefulness of serum cystatin C for the cardiovascular risk assessment. METHODS: 1,468 healthy subjects (814 men and 655 women), who visited health promotion center of Pusan National University Hospital for routine medical checkup were included. MS was defined by modified, revised National Cholesterol Education Program (NCEP) Adult Treatment Panel (ATP) III criteria. RESULTS: Mean serum cystatin C value was 0.87 +/- 0.17 mg/L. In partial correlation analysis adjusted by age, sex and Glomerular Filtration Rate (GFR), cystatin C was associated with most of metabolic parameters and especially had significant positive correlation with waist circumference (r = 0.215), triglyceride (TG) (r = 0.141), diastolic blood pressure (BP) (r = 0.116), and correlated negatively with high density lipoprotein (HDL) cholesterol (r = -0.152) (all P < 0.001). There were increasing trends of prevalence of MS with the increase of quartiles of cystatin C and as the number of MS components increased, cystatin C values significantly increased. Serum cystatin C was also significantly increased in MS (0.90 +/- 0.19 mg/L vs. 0.86 +/- 0.16 mg/L). In stepwise multiple regression analysis including the components of MS, Waist circumference, diastolic BP, triglyceride, and HDL cholesterol were independent determinants of serum cystatin C, but with creatinine, only waist circumference was independent determinant. CONCLUSIONS: Serum cystatin C was closely associated with MS and its related cardiovascular risk factors and might be useful as a tool of cardiovascular risk assessment.

Citations

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Cystatin C in Patients of Metabolic Syndrome and its Correlation with the Individual Components of Metabolic Syndrome
Sunita Aghade, Jayshree S Bavikar, Pragati S Kadam, Reshakiran J Shendye
Indian Journal of Medical Biochemistry.2019; 23(2): 293. CrossRef
Cystatin C as a Predictor for Diabetes according to Glycosylated Hemoglobin Levels in Korean Patients
Eon Ju Jeon, Ji Hyun Lee
Diabetes & Metabolism Journal.2016; 40(1): 32. CrossRef
Association of Obesity with Serum Cystatin C in Korean Adults
Yang Ho Kang
The Korean Journal of Obesity.2015; 24(4): 199. CrossRef

Review

Oxidative Stress and Cell Dysfunction in Diabetes: Role of ROS Produced by Mitochondria and NAD(P)H Oxidase.: Sang Soo Kim, Seok Man Son; Korean Diabetes J. 2008;32(5):389-398. Published online October 1, 2008; DOI: https://doi.org/10.4093/kdj.2008.32.5.389

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Abstract PDF

Oxidative stress has been considered to be a major contributor to the pathogenesis of the diabetic macrovascular and microvascular complications. In the absence of an appropriate antioxidant defense mechanism, increased oxidative stress leads to the activation of stress-sensitive intracellular signaling pathways and the formation of gene products that cause damage and contribute to the late complications ofdiabetes. The source of reactive oxygen species (ROS) in the pancreatic beta cells and insulin sensitive cells has postulated to be the mitochondrial electron transport chain. NAD(P)H oxidase-dependent ROS production is also important as the source both in pancreatic beta cells and other cells. NAD(P)H oxidase mediated ROS can alter parameters of signal transduction, insulin secretion, insulin action, cell proliferation and cell death. Additionally, oxidative stress as the pathogenic mechanism linking insulin resistance with dysfunction of both pancreatic beta cells and endothelial cells, eventually leads to diabetes and its complications. Further investigation of the mechanisms and its therapeutic interventions based on focusing NAD(P)H oxidase associated ROS production in the islet cells and other islet cells are needed

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Original Articles

Migration of Vascular Smooth Muscle Cells by High Glucose is Reactive Oxygen Dependent.: Yong Seong An, Ji Hae Kwon, Yang Ho Kang, In Ju Kim, Yong Ki Kim, Seok Man Son; Korean Diabetes J. 2008;32(3):185-195. Published online June 1, 2008; DOI: https://doi.org/10.4093/kdj.2008.32.3.185

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Abstract PDF: BACKGROUND
Oxidative stress contributes to vascular diseases in patients with diabetes. As the mechanism of development and progression of diabetic vascular complications is poorly understood, this study was aimed to assess the potential role of hyperglycemia-induced oxidative stress and to determine whether the oxidative stress is a major factor in hyperglycemia-induced migration of vascular smooth muscle cells (VSMCs). METHODS: We treated primary cultured rat aortic smooth muscle cells for 72 hours with medium containing 5.5 mM D-glucose (normal glucose), 30 mM D-glucose (high glucose) or 5.5 mM D-glucose plus 24.5 mM mannitol (osmotic control). We measured the migration of VSMCs and superoxide production. Immunoblotting of PKC isozymes using phoshospecific antibodies was performed, and PKC activity was also measured. RESULTS: Migration of VSMCs incubated under high glucose condition were markedly increased compared to normal glucose condition. Treatment with diphenyleneiodonium (DPI, 10 micromol/L) and superoxide dismutase (SOD, 500 U/mL) significantly suppressed high glucose-induced migration of VSMCs. Superoxide production was significantly increased in high glucose condition and was markedly decreased after treatment with DPI and SOD. High glucose also markedly increased activity of PKC-delta isozyme. When VSMCs were treated with rottlerin or transfected with PKC-delta siRNA, nitro blue tetrazolium (NBT) staining and NAD(P)H oxidase activity were significantly attenuated in the high glucose-treated VSMCs. Furthermore, inhibition of PKC-delta markedly decreased VSMC migration by high glucose. CONCLUSION: These results suggest that high glucose-induced VSMC migration is dependent upon activation of PKC-delta, which may responsible for elevated intracellular ROS production in VSMCs, and this is mediated by NAD(P)H oxidase.

Oxidative Stress Causes Vascular Insulin Resistance in OLETF Rat Through Increased IRS-1 Degradation.: Jung Lae Park, Young Sil Lee, Bo Hyun Kim, Yang Ho Kang, In Ju Kim, Yong Ki Kim, Seok Man Son; Korean Diabetes J. 2007;31(1):22-32. Published online January 1, 2007; DOI: https://doi.org/10.4093/jkda.2007.31.1.22

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Abstract PDF

BACKGROUND
Insulin resistance and oxidative stress have been reported to play essential pathophysiological roles in diabetic cardiovascular complication. The relationship between insulin resistance and oxidative stress in vasculature remains unclear. The study was conducted to assess whether oxidative stress induce vascular insulin resistance in OLETF rat, a model of type 2 diabetes METHODS: We used OLETF rats (20/30/40 weeks, n = 5/5/5), as models of type 2 DM, and LETO rats (20/30/40 weeks, n = 5/5/5) as controls. Aortas of each rats were extracted. Superoxide anion production was detected by NBT assay and lucigenin assay. 8-hydroxyguanosine (OHdG) and nitrotyrosine were detected as markers of oxidative stress in 20 and 40 weeks groups. The glucose uptake of aortas was measured by detecting 2-deoxyglucose uptake in both groups. The expression of IR, IRS-1, PI3-K and Akt/PKB were detected by immuno precipitation and immunoblotting in 20, 30 and 40 weeks groups RESULTS: Superoxide anion production and markers of oxidative stress (8-OHdG, nitrotyrosine) were significantly increased in aortas of OLETF rats compared with controls. Aortas of OLETF rats exhibited decreased IRS-1 content and increased phosphorylation of IRS-1 at Ser307 compared with LETO rats. There were no significant differences in expressions of IR, PI3-K and Akt/PKB between two groups CONCLUSION: These results suggest that oxidative stress induces insulin resistance in vasculature of OLETF rat specifically through increasing serine phosphorylation of IRS-1 and its degradation by a proteasome-dependent pathway, providing an alternative mechanism that may explain the association with insulin resistance and diabetic vascular complications.

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Anti-diabetic effects of benfotiamine on an animal model of type 2 diabetes mellitus
Kang Min Chung, Wonyoung Kang, Dong Geon Kim, Hyun Ju Hong, Youngjae Lee, Chang-Hoon Han
Korean Journal of Veterinary Research.2014; 54(1): 21. CrossRef

High Glucose Modulates Vascular Smooth Muscle Cell Proliferation Through Activation of PKC-sigma-dependent NAD(P)H oxidase.: Bo Hyun Kim, Chang Won Lee, Jung Lae Park, Yang Ho Kang, In Ju Kim, Yong Ki Kim, Seok Man Son; Korean Diabetes J. 2006;30(6):416-427. Published online November 1, 2006; DOI: https://doi.org/10.4093/jkda.2006.30.6.416

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Abstract PDF

BACKGROUND
Oxidative stress is thought to be one of the causative factors contributing to macrovascular complications in diabetes. However, the mechanisms of development and progression of diabetic vascular complications are poorly understood. We hypothesized that PKC-sigma isozyme contributes to ROS generation and determined their roles in the critical intermediary signaling events in high glucose-induced proliferation of vascular smooth muscle (VSM) cells. METHODS: We treated primary cultured rat aortic smooth muscle cells for 72 hours with medium containing 5.5 mmol/L D-glucose (normal glucose), 30 mmol/L D-glucose (high glucose) or 5.5 mmol/L D-glucose plus 24.5 mmol/L mannitol (osmotic control). We then measured cell number, BrdU incorporation, cell cycle and superoxide production in VSM cells. Immunoblotting of PKC isozymes using phoshospecific antibodies was performed, and PKC activity was also measured. RESULTS: High glucose increased VSM cell number and BrdU incorporation and displayed significantly greater percentages of S and G2/M phases than compared to 5.5 mmol/L glucose and osmotic control. The nitroblue tetrazolium (NBT) staining in high glucose-treated VSM cell was more prominent compared with normal glucose-treated VSM cell, which was significantly inhibited by DPI (10 micrometer), but not by inhibitors for other oxidases. High glucose also markedly increased activity of PKC-sigma isozyme. When VSM cells were treated with rottlerin, a specific inhibitor of PKC-sigma or transfected with PKC-sigma siRNA, NBT staining and NAD(P)H oxidase activity were significantly attenuated in the high glucose-treated VSM cells. Furthermore, inhibition of PKC-sigma markedly decreased VSM cell number by high glucose. CONCLUSION: These results suggest that high glucose-induced VSM cell proliferation is dependent upon activation of PKC-sigma, which may responsible for elevated intracellular ROS production in VSM cells, and this is mediated by NAD(P)H oxidase.

Citations

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High Glucose and/or Free Fatty Acid Damage Vascular Endothelial Cells via Stimulating of NAD(P)H Oxidase-induced Superoxide Production from Neutrophils
Sang Soo Kim, Sun Young Kim, Soo Hyung Lee, Yang Ho Kang, In Ju Kim, Yong Ki Kim, Seok Man Son
Korean Diabetes Journal.2009; 33(2): 94. CrossRef

Cell Cycle Progression of Vascular Smooth Muscle cell Through Modulation of p38 MAPK and GSK-3beta Activities Under High Glucose Condition.: Yang Ho Kang, In Ju Kim, Yong Ki Kim, Seok Man Son; Korean Diabetes J. 2005;29(5):418-431. Published online September 1, 2005

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Abstract PDF: BACKGOUND: Macroangiopathy, with atherosclerosis, is the leading cause of mortality and morbidity in diabetic patients. Vascular smooth muscle cells play a crucial role in atherosclerosis, as they proliferate, migrate and express genes that encode inducible growth factors. However, the mechanisms induced by hyperglycemia that accelerate the proliferative change of vascular smooth muscle cells in diabetes remain unclear. This study was aimed at clarifying the respective roles of hyperglycemia in the acceleration of vascular complications in diabetes, examine the effects of hyperglycemia on vascular smooth muscle cell proliferation and the possible underlying mechanisms, including cell cycle progression. METHODS: Primary cultured rat aortic RASMs were exposed to normal glucose(5 mmol/L D-glucose), high glucose(30 mmol/L D-glucose) or an osmotic control (5mmol/L D-glucose plus 24.5 mmol/L mannitol) for 72 hours. The effect of high glucose on cell proliferation was determined by assessing the cell count and BrdU incorporation. Proteins involved in the cell proliferation pathway (PDK1, Akt/PKB, p42/44 MAPK, p38 MAPK, GSK-3beta) and those in cell cycle progression (cdk4, cyclin D, cdk2, cyclin E and ppRb phosphorylation) were determined by Western blot analysis. cdk4 kinase and PKC activity assays were also performed. RESULTS: A high level of glucose increased both the cell count(P<0.01) and BrdU incorporation(P<0.01). The PDK1, Akt/PKB and p42/44 MAPK activities were not significantly increased. A high level of glucose significantly increased the activities of p38 MAPK (P<0.01) and GSK-3beta(P<0.05) and the expressions of cdk4, cyclin D and ppRb phosphorylation. The cdk4 (P<0.01) and PKC (P<0.05) activities were also significantly increased. The inhibition of protein kinase C with GF109203X markedly reduced the phosphorylations of p38 MAPK and GSK-3betaand the expressions of cdk4 and cyclin D. In addition, pretreatment with GF109203X decreased the cell number in response to a high glucose level. CONCLUSION: These findings suggest that a high level of glucose increases vascular smooth muscle cell proliferation, with the possible mechanism further increases the G1 to S phase cell cycle progression via the activation of PKC, p38 MAPK and GSK-3beta.

The Effect of High Glucose on the Proliferation and Migration of Vascular Smooth Muscle Cells.: Mi Kyoung Kim, Yang Ho Kang, Seok Man Son, In Ju Kim, Yong Ki Kim; Korean Diabetes J. 2004;28(5):407-415. Published online October 1, 2004

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Abstract PDF: BACKGROUND
Oxidative stress contributes to vascular diseases for patients with diabetes by promoting vascular smooth muscle cell (VSMC) proliferation, monocyte/macrophage infiltration, and vascular tone alteration. As the mechanism of development and progression of diabetic vascular complications is poorly understood, this study was aimed to assess the potential role of hyperglycemia-induced oxidative stress and to determine whether thise oxidative stress is a major factor in hyperglycemia-induced migration and proliferation of VSMCs. METHODS: Rat aortic VSMCs were incubated for 48 hours in either a normal glucose (NG, 5.5 mM) or a high glucose (HG, 30 mM) condition. We then measured the proliferation and migration of VSMCs and the superoxide production. RESULTS: The migration and proliferation of VSMCs incubated under a HG condition were markedly increased compared to the NG condition. Treatment with diphenyleneiodonium (DPI, 10 M) and superoxide dismutase (SOD, 500 U/mL) significantly suppressed the HG-induced migration and proliferation of VSMCs. Superoxide production was significantly increased in the HG condition, and it was markedly decreased after a treatment with DPI and SOD. CONCLUSION: These data suggest that HG-induced VSMC migration and proliferation are related to the production of superoxide anion that is derived from NAD(P)H oxidase.

Mechanism of Impaired Endothelium-dependent Vasodilation in Otsuka Long-Evans Tokushima Fatty (OLETF) Rats .: Kook Jin Chun, Seok Man Son, In Ju Kim, Chi Dae Kim, Seok Dong Yoo, Yong Ki Kim; Korean Diabetes J. 2002;26(1):47-57. Published online February 1, 2002

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Abstract PDF: BACKGROUND
Impaired vascular endothelium-dependent relaxation and augmented contractile responses have been reported in several long-term animals hyperglycemia models and human diabetic patients. Since oxidative stress has been implicated as a contributor to impaired vascular function, the mechanism of an impaired endothelium-dependent vasodilation in Otsuka Long-Evans Tokushima Fatty (OLETF) rats was investigated. METHODS: This present study was undertaken to characterize both the vascular production and the enzymatic source of the superoxide anion in the type 2 diabetic rats. RESULTS: In the thoracic aortas of OLETF rats, endothelium-dependent relaxation was markedly attenuated compared to that of the control rats (LETO, Long-Evans Tokushima Otsuka) in association with a significant increase in superoxide production (2421.39+/-07.01 nmol/min/mg). There was no difference in eNOS expression between the OLETF rats and LETO rats. The increased production of superoxide anion was significantly attenuated by diphenyleneiodonium (DPI, 10 mol/L), NAD (P)H oxidase inhibitor. In line with these results, studies using various enzyme inhibitors such as DPI, allopurinol, rotenone and L-NMMA suggest that the main source of superoxide anions in the aorta is NAD (P)H oxidase. CONCLUSION: These results suggest that enhanced NAD(P)H oxidase activity and reduced nitric oxide (NO) availability through an interaction between NO and superoxide anion contribute to the impaired endothelium-dependent vasodilation in OLETF rats.

Study on the Mechanism of Neutrophil Adhesion to Retinal Capillary Endothelial Cells under High Glucose Condition.: Seok Man Son, Young Sil Lee, Chang Won Lee, Seok Dong Yoo, In Ju Kim, Yong Ki Kim; Korean Diabetes J. 2001;25(1):35-49. Published online February 1, 2001

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Abstract PDF: BACKGROUND
Diabetic retinopathy is a leading cause of adult vision loss and blindness. Much of the retinal damage that characterizes the disease results from retinal vascular leakage and occlusion. Capillary occlusion is the result of microvascular thrombi in which erythrocytes, platelets and leukocytes each may play a major role. Thus, we investigated the pathogenesis of leukocyte stasis by exposing bovine retinal capillary endothelial cells (BRCECs) for high glucose concentration. METHODS: We examined the adhesion of neutrophils to BRCECs incubated in media containing 5.5-30 mmol/L D-glucose for 24 hours. We also measured the expression of E-selectin on endothelial cells and the activation of NF(nuclear transcription factor)-kappaB in nuclear fractions of endothelial cells by using electrophoretic mobility shift assay. RESULTS: We observed that 30 mmol/L D-glucose significantly increased the adhesion of neutrophils to BRCECs (12.5% vs. 3.0%, p<0.01) and migration of neutrophil across cultured BRCEC monolayers (41.0% vs. 21.0%, p<0.05) in respect to 5.5 mmol/L D-glucose. The expression of E-selectin was increased incubated with 30 mmol/L D-glucose compared with 5.5 mmol/L D-glucose (1.45 OD vs. 0.54 OD, p<0.01). Electrophoretic mobility shift assay of nuclear extracts of BRCECs exposed for 24 h to 30 mmol/L D-glucose revealed an intense NF-kappaB activation compared with cells cultured in 5.5 mmol/L D-glucose (8.72x104 countsxmm2 vs.1.88x104 countsxmm2, p<0.01). CONCLUSION: These results suggest that high glucose concentration promote neutrophil adhesion to the BRCECs through upregulation of cell surface expression of E-selectin, possibly depending on NF-kappaB activation and may have implications for the induction of microvasculopathy of diabetic retinopathy.

Solyble ICAM-1 and BCAM-1 in Patients with NIDDM.: Young Min Kim, Yong Gi Kim, Seok Man Son, In Ju Kim, Seok Dong Yoo, Young Keun Choi, Chang Won Lee, Jun Hyup Ahn; Korean Diabetes J. 1999;23(3):315-325. Published online January 1, 2001

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Abstract PDF: BACKGROUND
The development of vascular complications in diabetic patients changess their quality of life, as well as shortens their life expectancy. It has been recently discovered that the expressions of the cell adhesion molecules initiate vascular complications and have major effects on the progress of atherosclerosis. We measured soluble forms of intercelluar adhesion molecule-1 (sICAM-1) and vascular cell adhesion molecule-1 (sVCAM-1), the immunoglobulin superfamily members of the cell adhesion molecules concerning firm adhesion and transendothelial migration during leukocyte- endothelial cell interactions to clarify their concentrations and their relation with glycemic control and plasma lipoproteins as well as differences in concentration according to the presence of diabetic microvascular complcations in non-insulin dependent diabetes mellitus (NIDDM) patients. METHODS: Serum sICAM-1and sVCAM-1 levels were measured by commercial ELISA kits in 35 NIDDM patients without overt macrovascular complications of diabetes or acute inflammation and 10 normal controls matched with body mass index and plasma lipoprotein levels. The mean age of the patient group and control group was 55.82+3.43 years and 46.30+15.15 years, respectively. Clinical characteristics and laboratory parameters such as fasting plasma glucose, HbAplasma lipoproteins and status of diabetic microvascular complications were evaluated and their relations with the levels of sICAM-1 and sVCAM-1 were analyzed. RESULTS: 1) The level of sICAM-1 in NIDDM patients was significantly higher than that of normal controls (15.79+6.21 ng/mL vs. 11.98+2.35, p<0.05). sVCAM-1 showed the trend in elevation in NIDDM patients, but had no statistical significance (p=0.053). 2) The level of soluble ICAM-1 was positively correlated with HbAlc>, and plasma triglyceride levels (r=0.38, p<0.05, r=0.36, p<0.05, respectively) and negatively correlated with HDL (r=-0.44, p<0.01) in the patient group. There were no differences in their age, sex, and the presence of hypertension with the levels of sICAM-1 and no relation between sICAM-1 level and body mass index, plasma total cholesterol, Lp (a), fasting plasma glucose, fasting plasma C-peptide levels. Plasma LDL was partially correlated with the level of sICAM-1, but failed to reveal statistical significance. sVCAM-1 level was not correlated with any parameters discussed above, but had a tendency of correlation with HbAlc level (r=0.31, p=0.06). 3) No significant correlation was noted between the levels of sICAM-1 or sVCAM-1 and the duration of diabetes. 4) Both sICAM-1 and sVCAM-1 levels were significantly higher in patients with diabetic nephropathy when compared to patients without nephropathy (21.58+7.11 ng/mL vs. 14.06+4.84 ng/mL, p<0.05, 37.51+16.91 ng/mL vs. 22.26+8.89 ng/mL, p<0.05, respectively, but such differences were not noted when patients were classifed according to the presence of retinopathy or neuropathy. 5) Both sICAM-1and sVCAM-1 levels did not correlate in the patient group or in the normal control group. CONCLUSION: These findings suggest that enhanced expression of the the endothelial cell adhesion molecules in diabetic patients can be explained by endothelial dysfunction caused by persistent hyperglycemia and dyslipidemia. Furthermore, it can be suggested that endothelial dysfunction may be initiated by diabetes itself and can be deteriorated by combined dyslipidemia. From the result of the elevated concentrations of sICAM-1 and sVCAM-1 in patients with diabetic nephropathy, we can suggest that the elevation of these cell adhesion molecules may be useful as markers in diabetic nephropathy. More selective and prospective studies are necessary in order to reveal thesignificance of these cell adhesion molecules in the pathogenesis of diabetic vascular complications.

Effect of Oxidized LDL on Neutrophil Adhesion and Transendothelial Migration.: Seok Man Son, In Ju Kim, Yong Ki Kim; Korean Diabetes J. 1999;23(1):12-24. Published online January 1, 2001

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Abstract PDF: BACKGROUND
It is well recognized that oxidized low density lipoproteins (ox-LDL) play a critical role in the pathogenesis of atherosclerosis. The activation of circulating leukocytes and their adhesion to the vascular endothelium in response to acute stimuli characterize the first step in initiation of an acute inflammatory response. Through the action of degranulation products, adherent leukocytes induce vascular hyperpermeability and contribute to vascular injury. So, we have investigated the neutrophil adhesion to vascular endothelium, a constant feature of early atherogenesis and transendothelial migration of neutrophil induced by ox-LDL. METHOD: In a series of experiments, human umbilical vascular endothelial cells (HUVECs) were incubated for 24 h after addition of native human LDL (100 ug/mL) and of ox-LDL (100 ug/mL) to the medium. The adherence of 51Cr-labeled neutrophils to endothelial monolayers was measured by neutrophil adhesion assay. For diapedesis experiments, HUVECs were grown to confluence on 8.0um pore cell culture inserts. 51Cr-labeled neutrophils were added to the apical surface of HUVEC monolayers and allowed to migrate into the lower chamber for 3 h under the same preparations of native and oxidized LDLs. Reaults: The secretion of IL-8 depended on the concentration of IL-1a and LPS used to stimulate endothelial monolayers in vitro. In addition, ox-LDL triggered secretion of IL-8 from cultured HUVECs compared to that of n-LDL (867.6 pg/mL vs. 273.1 pg/mL, p<0.01). Increased adherence of neutrophils to HUVECs vs observed with ox-LDL preparation compared to native LDL preparation (36.8+1.5% vs. 25.9+1.7%, p<0.05). Similarly, neutrophil migration across cultured endothelial monolayers was also significantly increased by ox-LDL (48.7+3.8% vs. 34.4+2.9%, p<0.05). CONCLUSION: These results show that ox-LD1. can induce increased neutrophil adhesion and migration through IL-8, a potent effector of neutophil functions, secreted by stimulated endothelial cells. So, we suggest that ox-LDL may affect many components of the atherogenic process, including the early step in the initiation of m acute inflammation of vascular endothelial cells.

Effect of High Glucose Concentration on Expression of Adhesion Molecules in Endothelial Cells.: In Ju Kim, Seok Man Son, Min Ki Lee, Hee Jeong, Yong Ki Kim; Korean Diabetes J. 1998;22(3):280-289. Published online January 1, 2001

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Abstract PDF: BACKGROUND
Accelerated atherosclerotic vascular disease is the leading cause of mortality in patients with diabetes mellitus. Endothelial dysfunction, monocytes, and platelets are well known pathogenic factors in atherogenesis. Changes in the neutrophils and endothelial cells, an important early events in all inflammatory process, may contribute to the atherogenesis at early stage, but the significance of this process is not established yet. So we investigated the effects of glucose on the expression of adhesion molecules in endothelial cells, which retlects the change in endothelial-neutrophil adhesive interactions.METHODS: The human umbilical vein endothelial cells(HUVECs) are purchased from American Type Culture Collection. The cells were incubated upto 24 hours to evaluate the expression of E-selectin, PECAM-1, and P-selectin on the cell surface using whole cell ELISA method and soluble P-selectin under different glucose concentration(5.5, 15, and 30 mmol/L). Neutrophil adherence was also measured hy incubation of isolated human neutrophils with monolayers of HUVECs under same different glucose concentration. RESULTS: After 24h incubation with a various concentration of glucose, neutrophil adherence to high concentration of glucose(15 and 30mmol/L)- treated endothelium was significantly increased(5.0 +0.4 and 10.4+0.5%, respectively) compared with adhesion to low concentration of glucose(5.5mmol/ L)-treated endothelium(2.9.+0.4%). Incubation of HUVECs for 24 h in 30mmol/L glucose increased absorbance of E-selectin to 1.36+0.16(P<0.01) and reduced that of P-selectin to 0.56+0.04 compared with the results of respective control culture in 5.5mmol/L glucose(p<0.01), but not changed PECAM-1 expression. In addition, 24 h exposure of HUVECs to 30mmol/L glucose decreased soluble P-selectin concentration to 0.33+0.06ng/mL(P<0.01). CONCLUSION: The results of this study demonstrate that high concentration of glucose stimulates neutrophil adhesion to endothelial cells in association with increased expression of E-selectin. These results suggest that high glucose can directly affect interaction between neutrophil and endothelial cell through a adhesion molecule, especially E-selectin dependent mechanism. Further study should be necessary to investigate the significance of this phenomenon.

Effect of Glucose on Adherence of Neutrophils to Endothelial Cells.: Seok Man Son, Seok Dong Yoo, In Ju Kim, Yong Ki Kim, Hee Bag Park, Chi Dae Kim, Ki Whan Hong; Korean Diabetes J. 1997;21(3):262-270. Published online January 1, 2001

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Abstract PDF: BACKGROUND
Accelerated atherosclerotic vascular disease is the leading cause of mortality in patients with diabetes mellitus. To clarify the mechanisms that cause macrovascular dysfunction in diabetes, we examined the effect of high glucose on the adhesion of neutrophils to the endothelial cells and release of TNF-a from cultured rabbit aortic endotheIial cells. METHODS: Rabbit aortic endothelial cells in primary culture were prepared by the collagenase digestion method. Cells were incubated for various time upto 24 hours to evaluate TNF-a response to different glucose concentrations(0, 5.5, 11, 22mmol/L). Isolated rabbit neutrophils were incubated with monolayers of rabbit aortic endothelial cells under different glucose condition. RESULTS: After 24 hrs incubation with various concentrations of glucose, neutrophil adherence to high concentration of glucose(11 and 22mM)-treated endothelium was significantly increased(46+/-7 and 64 +/-6%, respectively) compared with adhesion to low concentration of glucose(0 or 5.5mM)-treated endothelium(3l +/-5 and 30+/-3%, respectively), In addition, when TNF-a imrnunoreactivity in the culture medium was measured by enzyme-linked immunoassay after 24 hours of incubation with various concentration of glucose, the secretion of TNF-a from endothelial cells was significantly increased in a concentration-dependent manner upon exposure to high concentration of glucose, CONCLUSION: The results of this study ciemonstrate tht high concentration of glucose stimulates neutrophil adhesion to endothelial cells in association with increased production of TNF-a from endothelial cells. These results suggest that glucose directly causes increased interaction between neutrophil and endothelial cell through a TNF-a-dependent mechaniasm,

Effect of high glucose on function of cultured rabbit vascular endothelial cells.: Seok Man Son, In Ju Kim, Yong Ki Kim, Chi Dae Kim, Ki Whan Hong; Korean Diabetes J. 1997;21(2):156-167. Published online January 1, 2001

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Abstract PDF: BACKGROUND
Vascular disease accounts for the majority of the clinical complications of diabetes mellitus. Changes in local control of vascular tone such as imbalanced production of relaxing and contracting factors by endothelium may be related to the initiation and maintenance of abnormal vascular reactivity characteristically seen in diabetic vascular complications. Cytokines and growth factors released from injured endothelial cells, T-cells, and macro-phages enhance atherogenesis. In this study, we examined NO and TNF-a released from cultured rabbit aortic endothelial cells(RAECs) under different glucose concentration to investigate the relationship between high glucose and endothelial cell dysfunction. METHODS: The thoracic and abdominal aortae of rabbit(23kg) were isolated and periadventitial connective tissue was carefully removed. Rabbit aortic endothelial cells in primary culture were prepared by the m.ethod of Schwartz with modification. RAECs were grown to confluence in 25 cm2 flask in DMEM supplemented with 20% FBS, 150pg/mL endothelial cell growth supplernent, 90pg/mL heparin, 100 U/mL penicillin and 100pg/mL streptomycin at 37'C in humidified 5% carbon dioxide in air. For experiments, confluent cells were replaced in 1 1 mm, 48 well plate containing same medium composition. Cells were then incubated in the presence or absence of FBS for various times up to 48 hours(time course) to eveluate the NO and TNF-a response to different glucose concentrations(0, 5.5, 11, 22, and 44 mmol/ L). Cells were also incubated with various concentration of ACH and ADP(10, 10', 10 and 10' mol/L) and 10' mol/L of ACH or ADP with different glucose concentrations for 24 hours to evaluate stimulated effect of ACH and ADP on NO release. RESULTS: 1) Total NO release from RAECs was significantly in a time-dependent. After 48 hours incubation, the total secretion of NO was significantly higher in culture medium with FRS than without FBS. 2) Glucose concentration resembling severe hyper-glycemic conditions(22 and 44 mmol/L) significantly inhibited NO release from RAECs, 3) Acetylcholine and ADP induced a clear dose-dependent NO release in RAECs. 4) Stimulation of acetylcholine and ADP on NO release according to different glucose concentration was not significantly higher than NO release in culture medium with glucose alone. 5) The increment in TNF-a levels was associated with a significant increase at higher glucose concentration, 6) There was a negative correlation between NO and TNF-a release in culture medium with FBS but not in culture medium without FBS. CONCLUSION: Our data show that decreased NO release and increased TNF-a release from RAECs were noted under high glucose concentration. Such interaction could play a significant role in the development of diabetic vascular complication in hyperglycernic conditions.

Study on Role of Neutrophil in Endothelial Cell Injury under High Glucose Condition.: Seok Man Son, In Ju Kim, Yong Ki Kim; Korean Diabetes J. 2000;24(6):652-665. Published online January 1, 2001

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Abstract PDF: BACKGROUND
High glucose level plays a major role in the injury of endothelium during the early event in diabetic vascular complication. It was speculated that high glucose level may cause endothelial cell injury by neutrophil activation. METHODS: The human umbilical vein endothelial cells (HUVEC) were obtained from American Type Culture Collection. The cells were incubated as long as 24 hours to evaluate the expression of E-selectin on the cell surface using whole cell ELISA method. The adherence of neutrophils to human umbilical endothelial cell monolayers and transendothelial migration of 51Cr-labeled neutrophils were evaluated under the condition of different concentrations of D-glucose (5.5, 15, and 30 mmol/L). L-glucose (30 mmol/L) was used as an osmotic control after 24h incubation. We also measured neutrophil-mediated endothelial cell cytotoxicity using a 51Cr-release assay and release of activating markers (lactoferrin and myeloeroxidase) from neutrophils under the same conditions. RESULTS: The expression of E-selectin was increased on endothelium when incubated with medium containing high glucose (30 mmol/L) compared to control (5.5 mmol/L) preparation (1.36 OD vs. 0.79 OD, p<0.05). Increased adherence of neutrophils to HUVEC was observed with high glucose when compared to control (10.4% vs. 2.9%, p<0.01). Similarly, neutrophil migration across the cultured endothelial monolayers were also significantly increased by high glucose (49.8% vs. 27.3%, p<0.05). 51Cr-release from endothelial cells by neutrophils stimulated with high glucose was greater than that with control (27.5% vs. 10.6%, p<0.05). Release of activating markers from neutrophils incubated with high glucose was greater than that from neutrophils incubated with control. CONCLUSION: These results indicate that high glucose increases the adherence and transendothelial migration of neutrophils and cause endothelial cell injury through neutrophil activation. Thus, it is concluded that high glucose level maybe contribute to manifestation of the diabetic vascular disease, including the early step in the initiation of an acute inflammation of vascular endothelial cells.

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