/ / 2 実験方法 1. 対象 ( 1) 75 g (OGTT) HbA1c 2 2 HbA1c 5.4% HbA1c % OGTT (No. 187) 2. 高血糖グルコースクランプ ( STG-22 ) (BD Insyte TM, 20G 30 mm,
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1 J Kanazawa Med Univ , 要約 : 2 (incr) (IRI) 2 incr ( 11.1 mmol/l 5.6 mmol/l ) 60 (0.5 g/kg) IRI C total GIP total GLP , 11.2 mmol/l IRI 1 IRI incr IRI (AUC) [7.2 vs nmol/l (0-45 )] AUC (48.2 vs. 60.4%) AUC IRI GIP 2 incr incr キーワード : (GIP) -1 (GLP-1) 2 諸言 (1) 2 (glucosedependent insulinotropic polypeptide: GIP) -1 (glucagon-like peptide-1: GLP-1) dipeptydil peptidase-4 (DPP-4) (2) 2 2 DPP-4 GLP GIP GLP-1 (3, 4) GLP-1 (5) (isoglycemic )(1, 6) 2 (7-15) 2 (1, 9, 16-20) Isoglycemic 7
2 / / 2 実験方法 1. 対象 ( 1) 75 g (OGTT) HbA1c 2 2 HbA1c 5.4% HbA1c % OGTT (No. 187) 2. 高血糖グルコースクランプ ( STG-22 ) (BD Insyte TM, 20G 30 mm, Becton Dickinson Medical Singapore) 45 20% 13 (glucose infusion rate: GIR) (21) 11.1 mmol/l 5.6 mmol/l g/kg (20% ) EDTA 2Na 4 1,900 g 測定項目 GIR GIR 5 GIR C ( Lumipulse Presto TM, Lumipulse Presto TM C- ( ) EDTA (enzyme-linked immunosorbent assay: ELISA) total Total GIP total GLP-1 Human GIP (Total) ELISA KIT (Millipore, St. Charles, MO, USA) EDI TM Total GLP-1 ELISA Kit (Epitope Diagnostics, San Diego, CA, USA) 1 2 / /9.4% /14.8% 4. 解析 C (20, 22) C C C 10 5 #0 Total GIP total GLP F t Mann-Whitney U p 0.05 結果 1. グルコースクランプ状況と血中インスリン Cペプチド濃度 ( 1 1A) ( 1.5) mmol/l ( - p ) GIR ( 1B) GIR ( - p 8
3 性別 ( 男 / 女 ) 年齢 ( 歳 ) BMI 糖尿病歴 ( 年 ) [ 中央値 ( 範囲 )] 降圧薬使用者脂質改善薬使用者 HbA1c (%) 空腹時血糖 (mmol/l) 空腹時インスリン (pmol/l) HOMA-IR HOMA-β 表 1. 糖尿病患者 6 / ± ±2.9 2 (0.3 2) 1 (ARB + CCB) 2 ( スタチン ) 6.5± ± ± ± ±25.0 非糖尿病対照者 6 / ± ±1.2-1 (ARB + CCB) 2 ( スタチン ) 5.5± ± ± ± ±20.9 ARB, アンジオテンシン II 受容体拮抗薬 ; CCB, カルシウム拮抗薬 ; 統計処理は 2 群が等分散の場合は対応のない t 検定, 異分散の場合は Mann- Whitney U 検定 ( ) を用いた p 値 図 1. ( ) ( )(A) (GIR; B) (IRI; C) C (CPR; D) 6 C D 1 C 4 9
4 / / ) ( 1 1C) Grubbs-Smirnov ( 1C, D 2 4A, B 3 4C-F 5 ) 2 ( 1 ) ( - p ) C ( 1D) 2. 時間経過補正後のグルコースクランプ状況と血中インスリン C ペプチド増加量 C ( 2A) GIR ( 2B) GIR 0 30 ( 2C) C ( 2D) #25 #120 OGTT 30 図 2. ( ) ( ) (A) (GIR; B) ( IRI; C) C ( CPR; D)
5 図 3. (AUC) (IRI; A, B, E, F) C (CPR; C, D, G, H) A-D AUC E-H AUC AUC A, C, E, G 45 B, D, F, H t Mann-Whitney U ( ) 図 4. ( ) ( ) total GIP(A, C, D) total GLP-1(B, E, F) A, B C-F AUC 2 C, E 45 D, F 75 1 (A, B) (C-F) 11
6 / / 図 5. ( 45 ) total GIP (A, D) total GLP-1 (B, E) ( IRI; C, F) A-C AUC D-F AUC GIR # 増加量 AUC 絶対値および増加量 AUC/ 総分泌量 AUC 相対比による経口糖負荷後インスリン C ペプチド反応の比較 ( 3A, B, E, F) C ( 3C, D, G, H) (AUC) AUC AUC ( 3A-D) AUC ( 3E-H) 2 ( 3A, C, E, G) ( 3B, D, F, H) 2 4. 高血糖クランプ下経口糖負荷後のGIP GLP-1 反応 total GIP / / pmol/l / pmol/l ( p ) total GLP ( , 4.6)/ ( , 3.7) pmol/l ( , 3.3)/ ( , 4.8) pmol/l total GIP ( 4A, C, D) total GLP-1 ( 4B, E, F) ( 4A, B) GIP GLP-1 GLP-1 ( - p ; / p /p 0.13) AUC ( 4C-F) GIP GLP-1 12
7 表 2. ( 45 ) 全 例 糖尿病患者 対照者 (n=12) (n=6) (n=5) (n=6) AUC[ΔIRI(#0 #45)] 絶対値を従属変数とした場合モデル 1 自由度調整決定係数 (p 値 ) (0.0019)* (0.011)* (0.10) - (0.71) 回帰係数 (p 値 ); AUC[ΔGIP(#0 #45)] AUC[ΔGLP-1(#0 #45)] ΔIRI(60') 6.29 (0.062) (0.98) 74.4 (0.0039)* 8.07 (0.059) 24.3 (0.22) 72.0 (0.029)* 3.19 (0.19) 22.1 (0.094) 5.30 (0.052) 3.04 (0.73) (0.79) 56.4 (0.41) モデル 2 自由度調整決定係数 (p 値 ) (0.0004)* (0.0026)* (0.30) (0.45) 回帰係数 (p 値 ); AUC[ΔGIP(#0 #45)] ΔIRI(60') 6.31 (0.043)* 74.5 (0.0017)* 9.48 (0.032)* 59.3 (0.021)* 3.83 (0.50) 39.4 (0.17) 4.61 (0.40) 57.5 (0.30) AUC[ΔIRI(#0 #45)]/ 総分泌量 AUC 相対比を従属変数とした場合モデル 3 自由度調整決定係数 (p 値 ) (0.20) (0.038)* (0.068) (0.32) 回帰係数 (p 値 ); AUC[ΔGIP(#0 #45)] AUC[ΔGLP-1(#0 #45)] ΔIRI(60') (0.054) (1.00) (0.70) (0.22) (0.023)* (0.12) (0.12) (0.038) (0.089) (0.39) (0.49) (0.58) モデル 4 自由度調整決定係数 (p 値 ) (0.093) (0.038)* (0.15) (0.15) 回帰係数 (p 値 ); AUC[ΔGIP(#0 #45)] AUC[ΔGLP-1(#0 #45)] (0.041) (0.91) (0.030)* (0.040)* (0.62) (0.10) (0.26) (0.44), 例外的な糖尿病患者 1 例を除いた場合 ; *, 有意なp 値 5. インクレチン効果とその規定因子候補間での回帰分析 1 GIP GLP-1 AUC AUC GIP C 1 GIP (r 0.71, p 0.014) AUC GIP r 0.72, p ( 2) AUC GIP p GIP GLP-1 4 (p 0.049) GIP (p 0.019) 考察
8 / / Andersen (22) (7-15, 20, 23) GIP GLP-1 (7-9, 13) GIR (7, 8, 10, 15) 2 Nauck (1) (16-19) BMI (31.5) HOMA-IR (4.79) 2 2 GIP GLP-1 DPP-4 total GIP total GLP-1 total GLP-1 2 kcal/ GLP-1 (24) GLP-1 GLP-1 (8, 13) GLP-1 GIP GLP-1 (5) 2 Nauck (1) (16-19) isoglycemic AUC AUC AUC AUC Nauck Elahi (20) Salehi (9) Woerle (14) C AUC AUC C AUC AUC AUC AUC 14
9 isoglycemic 2 Meier Nauck (25) GIP GIP GLP-1 (26) GIP (4) GIP 2 2 利益相反の開示 (DK) ( ) (B)(PR ; HI) KAKEN (K2011-6, K2012-6; AN) 文 1. Nauck M, Stöckmann F, Ebert R et al: Reduced incretin effect in type 2 (non-insulin-dependent) diabetes. Diabetologia 1986; 29: Yabe D, Seino Y: Two incretin hormones GLP-1 and GIP: comparison of their actions in insulin secretion and cell preservation. Prog Biophys Mol Biol 2011; 107: Toft-Nielsen MB, Damholt MB, Madsbad S et al: Determinants of the impaired secretion of glucagon-like peptide-1 in type 2 diabetic patients. J Clin Endocrinol Metab 2001; 86: Nauck MA, Heimesaat MM, Orskov C et al: Preserved incretin activity of glucagon-like peptide 1 [7-36 amide] but not of synthetic human gastric inhibitory polypeptide in patients with type-2 diabetes mellitus. J Clin Invest 1993; 91: Nauck MA, Vardarli I, Deacon CF et al: Secretion of glucagon-like peptide-1 (GLP-1) in type 2 diabetes: what is up, what is down? Diabetologia 2011; 54: Nauck MA, Homberger E, Siegel EG et al: Incretin effects of increasing glucose loads in man calculated from venous insulin and C-peptide responses. J Clin Endocrinol Metab 1986; 63: Salehi M, Vahl TP, D Alessio DA: Regulation of islet hormone release and gastric emptying by endogenous glucagon-like peptide 1 after glucose ingestion. J Clin Endocrinol Metab 2008; 93: Vollmer K, Gardiwal H, Menge BA et al: Hyperglycemia acutely lowers the postprandial excursions of glucagon-like peptide-1 and gastric inhibitory polypeptide in humans. J Clin Endocrinol Metab 2009; 94: Salehi M, Aulinger B, Prigeon RL et al: Effect of endogenous GLP-1 on insulin secretion in type 2 diabetes. Diabetes 2010; 59: Henry RR, Smith SR, Schwartz SL et al: Effects of saxagliptin on -cell stimulation and insulin secretion in patients with type 2 diabetes. Diabetes Obes Metab 2011; 13: Carrel G, Egli L, Tran C et al: Contributions of fat and protein to the incretin effect of a mixed meal. Am J Clin Nutr 2011; 94: Salehi M, Prigeon RL, D Alessio DA: Gastric bypass surgery enhances glucagon-like peptide 1-stimulated postprandial insulin secretion in humans. Diabetes 2011; 60: Salehi M, Aulinger B, D Alessio DA: Effect of glycemia on plasma incretins and the incretin effect during oral glucose tolerance test. Diabetes 2012; 61: Woerle HJ, Carneiro L, Derani A et al: The role of endogenous incretin secretion as amplifier of glucose-stimulated insulin secretion in healthy subjects and patients with type 2 diabetes. Diabetes 2012; 61: An Z, Prigeon RL, D Alessio DA: Improved glycemic control enhances the incretin effect in patients with type 2 diabetes. J Clin Endocrinol Metab 2013; 98: Knop FK, Vilsbøll T, Højberg PV et al: Reduced incretin effect in type 2 diabetes: cause or consequence of the diabetic state? Diabetes 2007; 56: Laferrère B, Heshka S, Wang K et al: Incretin levels and effect are markedly enhanced 1 month after Roux-en-Y gastric bypass surgery in obese patients with type 2 diabetes. Diabetes Care 2007; 30: Muscelli E, Mari A, Casolaro A et al: Separate impact of obesity and glucose tolerance on the incretin effect in normal subjects and type 2 diabetic patients. Diabetes 2008; 57: Mari A, Bagger JI, Ferrannini E et al: Mechanisms of the incretin effect in subjects with normal glucose tolerance and patients with type 2 diabetes. PLoS One 2013; 8: e Elahi D, Andersen DK, Muller DC et al: The enteric enhancement of glucose-stimulated insulin release. The role of GIP in aging, obesity, and non-insulin-dependent diabetes mellitus. Diabetes 1984; 33: DeFronzo RA, Tobin JD, Andres R: Glucose clamp technique: a method for quantifying insulin secretion and resistance. Am J Physiol 1979; 237: E 献 15
10 / / 22. Andersen DK, Elahi D, Brown JC et al: Oral glucose augmentation of insulin secretion. Interactions of gastric inhibitory polypeptide with ambient glucose and insulin levels. J Clin Invest 1978; 62: Henchoz E, D Alessio DA, Gillet M et al: Impaired insulin response after oral but not intravenous glucose in heart- and liver-transplant recipients. Transplantation 2003; 76: Marathe CS, Rayner CK, Bound M et al: Small intestinal glucose exposure determines the magnitude of the incretin effect in health and type 2 diabetes. Diabetes 2014; 63: Meier JJ, Nauck MA: Is the diminished incretin effect in type 2 diabetes just an epi-phenomenon of impaired beta-cell function? Diabetes 2010; 59: Vilsbøll T, Krarup T, Madsbad S et al: Both GLP-1 and GIP are insulinotropic at basal and postprandial glucose levels and contribute nearly equally to the incretin effect of a meal in healthy subjects. Regul Pept 2003; 114: Reduced Incretin Effect in Type 2 Diabetes is Mainly Attributable to Impaired Glucose Responsiveness of β-cells: Evaluation via an Oral Sugar Load under a Hyperglycemic Clamp Hiroki Ito, Atsushi Nakagawa, Daisuke Koya Department of Endocrinology and Metabolism, Kanazawa Medical University Graduate School of Medical Science, Uchinada, Ishikawa , Japan In this study, we aimed to ascertain whether the incretin effect is reduced in type 2 diabetes and to identify factors that regulate this effect. To evaluate the incretin effect, we administered an oral sugar load under the hyperglycemic glucose clamp technique, and the effect was defined in two ways: an absolute increase in insulin levels and the ratio of the increase in insulin to the total insulin secretion. After an overnight fast, the hyperglycemic clamp technique (the higher of either 5.6 mmol/l above the fasting level or 11.1 mmol/l) was performed in 6 type 2 diabetic and 6 nondiabetic men. Maltose (0.5 g/kg) was orally administered 60 minutes after initiation of glucose infusion. Blood samples were drawn every 5 or 10 minutes for 210 minutes, and insulin, C-peptide, total glucose-dependent insulinotropic polypeptide (GIP), and glucagon-like peptide-1 (GLP-1) levels were analyzed. Blood glucose was clamped at 12.8 and 11.2 mmol/l in the diabetic and control subjects, respectively. One diabetes patient was excluded from the diabetic group before comparison of the two groups because his excessive insulin responses were statistically judged to be outliers. Because the insulin levels continue to linearly increase throughout the hyperglycemic clamp procedure, the estimated moving basal levels of insulin were extrapolated from the pre-maltose concentrations, and the incretin effects were evaluated based on the increments from the estimated basal levels. The absolute value of the area under the curve (AUC) of incremental insulin concentrations was significantly lower in the diabetic group than that in the control group (7.2 vs min nmol/l [0 40 min]), whereas no significant difference was detected in the ratio of the incremental AUC to the total AUC of insulin (48.2 vs. 60.4%). There were no significant differences in the GIP and GLP-1 responses between groups. Univariate and multivariate regression analyses demonstrated significant correlations of the absolute incremental insulin AUCs with the insulin increments before the oral sugar load, and of the incremental AUC/total AUC ratios of insulin with the GIP increments, but not with the pre-load insulin. These findings suggest that the reduced incretin effect in type 2 diabetes is attributable mainly to impaired β-cell glucose responsiveness, and there is no specific impairment in the incretin effect in type 2 diabetes. Key Words: incretin effect, hyperglycemic glucose clamp, glucose-dependent insulinotropic polypeptide, glucagon-like peptide-1, type 2 diabetes 16
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