oral glucose administration leads to greater insulin release from pancreatic islets than intravenous glucose loading that yields equivalent glucose levels . gut hormonal substances released in response to glucose include the incretins , gastric inhibitory polypeptide ( gip ) and glucagonlike peptide1 ( glp1 ) , which are responsible for 5060% of postprandial insulin secretion . gip is secreted on meal ingestion from kcells in the proximal small intestine , whereas glp1 is secreted from lcells in the distal small intestine and colon , and binds to their respective receptors ( gip receptor [ gipr ] and glp1 receptor ) on the surface of pancreatic cells to stimulate insulin secretion by increasing the intracellular adenosine 3,5monophosphate ( camp ) concentration . the incretin effect has been shown to be reduced in type 2 diabetic subjects compared with those with normal glucose tolerance ( ngt ) in previous studies , suggesting that a reduced incretin effect might be associated with hyperglycemia after food intake and glucose loading in type 2 diabetes . when intravenous infusion of gip or glp1 was carried out in type 2 diabetic subjects , glp1 potentiated insulin secretion from pancreatic cells , but gip did not , showing that the gipr signal is downregulated in cells in type 2 diabetes . in studies using rodent models , it was reported that gipr mrna and protein expression levels in islets are decreased in the diabetic state . in contrast , in the nondiabetic obese state the gip signal might be enhanced as a result of increased gipr sensitivity of cells to gip or increased gip secretion from kcells in the nondiabetic obese state . indeed , gip concentrations are reported to be increased in obese rodent models and human subjects compared with those in lean rodents and human subjects , respectively . furthermore , we have previously shown the hypersensitivity of gipr to gip in cells of high fatinduced obese mice . plasma glp1 concentrations in type 2 diabetic patients are reported to be reduced after meal ingestion and glucose loading . however , in other studies it was reported that glp1 concentrations did not differ in ngt and type 2 diabetic subjects . thus , the measurement of gip and glp1 concentrations in various metabolic states is important to evaluate the effects of incretin on insulin secretion . insulin sensitivity in asian subjects has been shown to be higher than in mexican americans and caucasians in previous reports , which is partly as a result of the fact that asians , including japanese , are generally less obese . furthermore , insulin secretion rather than insulin sensitivity is the more important factor in progression from ngt to diabetes in japanese subjects . we have reported that earlyphase insulin secretion is considerably decreased , even in japanese ngt subjects with 1h plasma glucose ( pg ) levels during oral glucose tolerance test ( ogtt ) of more than 180  mg / dl . thus , it is especially important to evaluate incretin secretion and determine the factors associated with incretin secretion in japanese ngt subjects , because gip and incretin is responsible for more than 50% of postprandial insulin secretion after glucose ingestion . the factors responsible for incretin secretion have been reported in caucasian subjects , but have not been thoroughly elucidated in japanese subjects . in the present study , we evaluated gip and glp1 levels during ogtt and determined the factors involved in gip and glp1 secretion ( area under the curve [ auc ] of gip and glp1 during ogtt ) in japanese ngt subjects . the subjects had no history of hypertension , hyperlipidemia or kidney and liver diseases , and did not take any drugs 2  weeks before the study . the study was designed in compliance with the ethics regulations of the helsinki declaration and kyoto university . blood samples for the measurement of liver and kidney function , hba1c , serum triglyceride ( tg ) , total cholesterol and highdensity lipoprotein ( hdl)cholesterol levels were drawn after an overnight fast . all subjects received ogtt . after the subjects fasted overnight for 1016  h , standard ogtt with 75  g glucose was given according to the national diabetes data group recommendations . blood samples were collected at 15 , 0 , 10 , 20 , 30 , 60 , 90 , 120 , 150 and 180  min after glucose loading and were centrifuged at 1800  g at 4c for 10  min . after collecting supernatant of the samples , plasma and serum plasma gip , glp1 levels and the various parameters ( pg , serum immunoreactive insulin [ iri ] , serum cpeptide reactivity [ cpr ] , tg , serum free fatty acid [ ffa ] and plasma glucagon ) were measured at the indicated times ( plasma gip and glp1 levels were measured at 15 , 0 , 10 , 30 , 60 , 90 , 120 and 180  min after glucose loading , and plasma glucagon levels were measured at 15 , 0 , 30 , 60 , 90 , 120 and 180  min after glucose loading ) . the pg levels were measured by glucose oxidase method . total gip and total glp1 levels were measured using human gip elisa kit ( linco research , st charles , mo , usa ; range of detection from 8.2  pg / ml to 2000  pg / dl ) and human glp1 elisa kit ( meso scale discovery , gaithersburg , md , usa ; range of detection from 2.4  pg / ml to 1,000,000  pg / dl ) , respectively , as previously described . the auc of pg , iri , cpr , tg , ffa , glucagon , total gip ( aucgip ) and total glp1 ( aucglp1 ) were calculated . we then analyzed the relationship between the auc of gip ( gip secretion ) and glp1 ( glp1 secretion ) and age , body mass index ( bmi ) and the parameters during ogtt . basal insulin secretion and sensitivity were evaluated by homeostasis model assessment ( homa ) cell function and homeostasis model assessment of insulin resistance ( homair ) , respectively . earlyphase insulin secretion and systemic insulin sensitivity during ogtt were evaluated by insulinogenic index and insulin sensitivity index ( isi ) composite . the calculations of the four indices were as follows : equation 
 equation 
 equation 
 equation 
  all analyses were carried out using statistical analysis software ( spss version 17.0 , ibm , somers , ny , usa ) system . statistical analysis was carried out by anova with fisher s plsd test for changing levels of gip , glp1 , and the parameters during ogtt and differences between the two groups were assessed by unpaired ttest . we used simple regression analysis to determine the relationship between aucgip or aucglp1 and the age , bmi and the parameters during ogtt , and we carried out multiple regression analysis to determine the factors most strongly associated with aucgip and aucglp1 , and the indices of insulin secretion and sensitivity . the subjects had no history of hypertension , hyperlipidemia or kidney and liver diseases , and did not take any drugs 2  weeks before the study . the study was designed in compliance with the ethics regulations of the helsinki declaration and kyoto university . blood samples for the measurement of liver and kidney function , hba1c , serum triglyceride ( tg ) , total cholesterol and highdensity lipoprotein ( hdl)cholesterol levels were drawn after an overnight fast . all subjects received ogtt . after the subjects fasted overnight for 1016  h , standard ogtt with 75  g glucose was given according to the national diabetes data group recommendations . blood samples were collected at 15 , 0 , 10 , 20 , 30 , 60 , 90 , 120 , 150 and 180  min after glucose loading and were centrifuged at 1800  g at 4c for 10  min . after collecting supernatant of the samples , plasma and serum plasma gip , glp1 levels and the various parameters ( pg , serum immunoreactive insulin [ iri ] , serum cpeptide reactivity [ cpr ] , tg , serum free fatty acid [ ffa ] and plasma glucagon ) were measured at the indicated times ( plasma gip and glp1 levels were measured at 15 , 0 , 10 , 30 , 60 , 90 , 120 and 180  min after glucose loading , and plasma glucagon levels were measured at 15 , 0 , 30 , 60 , 90 , 120 and 180  min after glucose loading ) . total gip and total glp1 levels were measured using human gip elisa kit ( linco research , st charles , mo , usa ; range of detection from 8.2  pg / ml to 2000  pg / dl ) and human glp1 elisa kit ( meso scale discovery , gaithersburg , md , usa ; range of detection from 2.4  pg / ml to 1,000,000 the auc of pg , iri , cpr , tg , ffa , glucagon , total gip ( aucgip ) and total glp1 ( aucglp1 ) were calculated . we then analyzed the relationship between the auc of gip ( gip secretion ) and glp1 ( glp1 secretion ) and age , body mass index ( bmi ) and the parameters during ogtt . basal insulin secretion and sensitivity were evaluated by homeostasis model assessment ( homa ) cell function and homeostasis model assessment of insulin resistance ( homair ) , respectively . earlyphase insulin secretion and systemic insulin sensitivity during ogtt were evaluated by insulinogenic index and insulin sensitivity index ( isi ) composite . the calculations of the four indices were as follows : equation 
 equation 
 equation 
 equation 
  all analyses were carried out using statistical analysis software ( spss version 17.0 , ibm , somers , ny , usa ) system . statistical analysis was carried out by anova with fisher s plsd test for changing levels of gip , glp1 , and the parameters during ogtt and differences between the two groups were assessed by unpaired ttest . we used simple regression analysis to determine the relationship between aucgip or aucglp1 and the age , bmi and the parameters during ogtt , and we carried out multiple regression analysis to determine the factors most strongly associated with aucgip and aucglp1 , and the indices of insulin secretion and sensitivity . mean age was 31.7    1.3  years and mean bmi was 23.1    0.9  kg / m . hdl , highdensity lipoprotein ; homa , homeostasis model assessment ; homair , homeostasis model assessment of insulin resistance ; isi , insulin sensitivity index . the levels of gip , glp1 , pg , iri , cpr , tg , ffa and glucagon after glucose loading were measured ( figure  1 ) . the subjects were diagnosed ngt according to who criteria with fasting plasma glucose and 2h glucose levels below 6.1 and 7.8  mmol / l , respectively . levels of pg , iri and cpr were significantly increased from 10  min after glucose loading compared with fasting level ( figure  1a  c ) . ffa levels were significantly decreased from 10  min after glucose loading ( figure  1d ) . glucagon levels were significantly decreased from 30  min after glucose loading ( figure  1f ) . total gip levels were significantly increased from 10  min during ogtt ( figure  1 g ) . total glp1 levels were significantly increased from 10  min during ogtt with peaks at 30 and 120  min ( figure  1h ) . concentrations of ( a ) plasma glucose , ( b ) serum immunoreactive insulin , ( c ) serum cpeptide reactivity ( cpr ) , ( d ) serum free fatty acid ( ffa ) , ( e ) serum triglyceride ( tg ) , ( f ) glucagon , ( g ) total gastric inhibitory polypeptide ( gip ) and ( h ) total glucagonlike peptide1 ( glp1 ) during oral glucose tolerance test in 17 japanese subjects . mean    se , * p  <  0.05 , * * p  <  0.01 , * * we analyzed the relationship between aucgip or aucglp1 and age , bmi and the several parameters ( auc of pg , iri , cpr , tg , ffa and glucagon ) . aucgip were positively related to bmi and auc of cpr , iri and glucagon , but aucglp1 was not related to these factors ( figure  2a  c ; auc data of iri during ogtt are not shown ; p  <  0.05 ) . in contrast , aucglp1 was inversely related to auc of pg ( figure  2d ) , but aucgip was not . simple regression analysis of gastric inhibitory polypeptide secretion ( aucgip ) and ( a ) body mass index ( bmi ) , ( b ) auc of serum cpeptide reactivity ( cpr ) and ( c ) glucagon . ( d ) simple regression analysis of glucagonlike peptide1 secretion ( aucglp1 ) and auc of plasma glucose ( pg ) . we then analyzed the relationship between aucgip or aucglp1 and indices of insulin secretion and insulin sensitivity . aucgip was positively related to insulinogenic index and homair , whereas aucglp1 was positively related to homa cell function ( figure  3a  c ) . isi composite was not related to either aucgip or aucglp1 ( figure  3d ) . in addition , multiple regression analysis was carried out to determine the factors strongly associated with aucgip and aucglp . the insulinogenic index was the most strongly associated factor in aucgip ( correlation coefficients 0.56 , standardized  0.56 , p  <  0.05 ) of the four indices ; homa cell function was the strongest factor in aucglp1 ( homa cell function : correlation coefficients 0.524 , standardized  0.870 , p  <  0.01 , isi composite : correlation coefficients 0.063 , standardized  0.581 , p  <  0.05 ) .  relationship between gastric inhibitory polypeptide secretion ( aucgip ) and glucagonlike peptide1 secretion ( aucglp1 ) and the indices of insulin secretion and insulin sensitivity . ( a ) insulinogenic index , ( b ) homeostasis model assessment ( homa ) cell function , ( c ) homeostasis model assessment of insulin resistance ( homair ) and ( d ) insulin sensitivity index ( isi ) composite . in the present study , we estimated the incretin level after glucose loading in japanese ngt subjects and found that plasma gip and glp1 levels during ogtt are related to different factors . incretin action of gip is reduced in the diabetic state as a result of decreased gip receptor expression on pancreatic cells , whereas gip signaling is enhanced and maintains glucose homeostasis by compensatory increased insulin secretion in the obese state . in some human studies in caucasians , plasma gip levels are increased in obese subjects and there is a positive relationship between aucgip and auc of ffa during ogtt . in the present study , aucgip after glucose loading was not associated with auc of ffa , but was positively associated with bmi , homair , and auc of iri and cpr after glucose loading . in fact , obese subjects are known to have hyperinsulinemia and insulin resistance , and bmi was strongly associated with auc of iri and cpr . thus , gip secretion from kcells may well be associated with insulin resistance to maintain postprandial hyperinsulinemia in japanese ngt subjects . it might be explained by the fact that gip secretion is associated with the amount of glucose loading , whereas blood glucose levels are maintained within normal levels by gipinduced compensatory insulin secretion in ngt subjects . glp1 secretions of type 2 diabetes subjects after glucose or meal ingestion are diverse in human studies . recently , it is reported that glp1 levels after ingestion of glucose and mix meal in japanese type 2 diabetic subjects were not decreased compared with those in ngt subjects , suggesting that glp1 secretion is not decreased in japanese type 2 diabetes . two studies of caucasian subjects found that aucglp1 during ogtt is positively associated with age and auc of glucagon , whereas auc of glp1 is negatively associated with bmi or bodyweight and auc of ffa . in the present study , aucglp1 was negatively related to auc of pg during ogtt , showing that the increase in glp1 secretion after glucose loading is associated with a decrease in postprandial glucose levels in japanese ngt subjects . it has been reported that glp1 levels after glucose loading are positively related to gastric empting in caucasian subjects . although we did not measure gastric empting of the subjects in the present study , increasing glp1 secretion after glucose loading might decrease postprandial glucose levels through gastric emptying . in the present study , however , because japanese subjects are less obese than caucasian subjects , the difference observed in the relationship between aucgip and glp1 , and auc of ffa might reflect this ethnic difference in caucasians and japanese . insulin secretion , rather than insulin sensitivity , is the more important factor in the progression from ngt to type 2 diabetes in japanese patients . because incretin is an intestinal hormone that induces postprandial insulin secretion , we hypothesize that gip and glp1 secretion is more crucial in japanese subjects than in caucasian subjects . indeed , glp1 mimetics and dpp4 inhibitors improve glycemic control better in japanese type 2 diabetic patients than in caucasian type 2 diabetic patients in clinical trials . we therefore evaluated the correlation between gip secretion ( aucgip ) and glp1 secretion ( aucglp1 ) , and the indices of insulin secretion and insulin sensitivity in japanese ngt subjects during ogtt . the values of homa cell , insulinogenic index , homair and isi composite were similar to those in previous studies of japanese subjects . aucgip was positively associated with the insulinogenic index and homair , and the insulinogenic index was strongly associated with aucgip , whereas aucglp1 was associated only with homa cell among the four indices . it has been reported that earlyphase insulin secretion is an important factor in the progression from ngt through impaired glucose tolerance ( igt ) to type 2 diabetes , and that basal insulin secretion ( homa cell ) and insulin resistance are important factors in the progression from ngt through impaired fasting glucose ( ifg ) to type 2 diabetes in japanese patients . thus , enhancing the gip and glp1 signals might be particularly useful in inhibiting the progression of type 2 diabetes in japanese patients . recently , variants at the gip receptor gene locus associated with 2h glucose levels during ogtt were identified by metaanalysis of genomewide association studies . in subjects who carry this gip receptor risk allele , these data seem to support our results that gip secretion is associated with insulinogenic index in japanese ngt subjects . in conclusion , we evaluated plasma gip and glp1 levels during ogtt in japanese ngt subjects . glp1 secretion was associated with pg during ogtt , and basal insulin secretion ( homa cell ) and gip secretion was associated with bmi and earlyphase insulin secretion ( insulinogenic index ) . thus , there might be different factors associated with gip and glp1 secretion during ogtt in japanese subjects .
abstractaims / introduction :   gastric inhibitory polypeptide ( gip ) and glucagonlike peptide1 ( glp1 ) are major incretins that potentiate insulin secretion from pancreatic cells . 
 the factors responsible for incretin secretion have been reported in caucasian subjects , but have not been thoroughly evaluated in japanese subjects . 
 we evaluated the factors associated with incretin secretion during oral glucose tolerance test ( ogtt ) in japanese subjects with normal glucose tolerance ( ngt).materials and methods :   we measured plasma gip and glp1 levels during ogtt in 17 japanese ngt subjects and evaluated the factors associated with gip and glp1 secretion using simple and multiple regression analyses.results :   gip secretion ( aucgip ) was positively associated with body mass index ( p  <  0.05 ) , and area under the curve ( auc ) of cpeptide ( p  <  0.05 ) and glucagon ( p  <  0.01 ) , whereas glp1 secretion ( aucglp1 ) was negatively associated with auc of plasma glucose ( p  <  0.05 ) . 
 the insulinogenic index was most strongly associated with gip secretion ( p  <  0.05 ) ; homeostasis model assessment cell was the most the strongly associated factor in glp1 secretion ( p  <  0.05 ) among the four indices of insulin secretion and insulin sensitivity.conclusions :   several distinct factors might be associated with gip and glp1 secretion during ogtt in japanese subjects . 
 ( j diabetes invest , doi : 10.1111/j.20401124.2010.00078.x , 2011 )