Diabetes
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Purchase Article
Right arrow View Shopping Cart
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow Request Permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Deacon, C. F.
Right arrow Articles by Holst, J. J.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Deacon, C. F.
Right arrow Articles by Holst, J. J.
Social Bookmarking
Add to CiteULike   Add to Del.icio.us   Add to Digg   Add to Reddit   Add to Technorati  
What's this?
Diabetes 50:1588-1597, 2001
© 2001 by the American Diabetes Association, Inc.

Dipeptidyl Peptidase IV Inhibition Reduces the Degradation and Clearance of GIP and Potentiates Its Insulinotropic and Antihyperglycemic Effects in Anesthetized Pigs

Carolyn F. Deacon1, Patricia Danielsen1, Letty Klarskov2, Mette Olesen2, and Jens J. Holst1

1 Medical Physiology and
2 Experimental Medicine, The Panum Institute, University of Copenhagen, Copenhagen, Denmark


   ABSTRACT
TOP
 ABSTRACT
INTRODUCTION
 RESEARCH DESIGN AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
Glucose-dependent insulinotropic peptide (GIP) is known to be degraded by dipeptidyl peptidase IV (DPP IV), forming an inactive metabolite, but the extent of the enzyme’s role in regulating the biological activity of GIP in vivo is still largely unknown. In nonfasted anesthetized pigs given an intravenous infusion of GIP, the intact peptide (determined by a novel NH2-terminally directed radioimmunoassay) accounts for only 14.5 ± 2.5% of total immunoreactivity. This is increased (to 40.9 ± 0.9%, P < 0.0001) by coadministration of valine-pyrrolidide (a specific DPP IV inhibitor) at a dose that completely inhibits plasma DPP IV activity. The plasma t1/2 of intact GIP is prolonged by the inhibitor (from 3.3 ± 0.3 to 8.1 ± 0.6 min; P < 0.001), whereas the t1/2 for COOH-terminal immunoreactivity is unaffected (13.2 ± 0.5 and 11.5 ± 0.8 min, pre- and postinhibitor). Measurement of arteriovenous concentration differences revealed that the liver, kidney, and extremities are the main sites of removal of exogenous intact GIP (organ extractions, 28.0 ± 2.2, 26.3 ± 5.7, and 21.8 ± 3.0%, respectively). These organ extractions are reduced (P < 0.02) but not eliminated (kidney and extremities) by valine-pyrrolidide (to 6.5 ± 4.6, 14.1 ± 3.1, and 13.9 ± 2.4%, respectively). Valine-pyrrolidide potentiates the insulinotropic effect of GIP (P < 0.02), resulting in an enhanced glucose disappearance rate (k, from 8.0 ± 0.5 to 15.5 ± 2.2%/min; P < 0.01) and a reduction in the glucose excursion after an intravenous glucose load (area under the curve, from 133 ± 23 to 75 ± 9 min · mmol/l; P < 0.05). These results suggest that DPP IV plays an important role in GIP metabolism but is not the sole enzyme responsible for its NH2-terminal degradation. Nevertheless, DPP IV inhibition increases the proportion of intact peptide sufficiently to enhance its insulinotropic and antihyperglycemic effects.


   INTRODUCTION
TOP
 ABSTRACT
 INTRODUCTION
RESEARCH DESIGN AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
Glucose-dependent insulinotropic peptide (GIP), also known as gastric inhibitory polypeptide, is secreted from the K-cells in the upper small intestine, in response to the presence of nutrients, especially fats (1). Together with the structurally related peptide, glucagon-like peptide 1 (GLP-1), released from the intestinal L-cell, GIP is considered an incretin hormone (2).

In vitro studies have shown GIP to be a substrate for the enzyme dipeptidyl peptidase IV (DPP IV) (3,4), resulting in the formation of a truncated inactive metabolite, GIP(3–42) (5). However, because all previously described GIP assays crossreact with both intact GIP and the metabolite, measurement of circulating concentrations of the biologically active form of the peptide has been hampered, so that the relevance of DPP IV for regulation of GIP activity in vivo has been difficult to investigate fully. The recent development of a novel assay directed toward the intact NH2-terminus of GIP has identified GIP(3–42) as a major component of endogenous GIP immunoreactivity in human plasma (6), suggesting an important role for the enzyme in GIP metabolism.

Only a few studies have focused on identifying sites of GIP degradation. In vitro studies using isolated perfused rat livers demonstrated a negligible hepatic extraction of exogenous immunoreactive GIP (7). Similar findings were obtained by comparing the portal-to-peripheral venous ratios (7) or arteriovenous concentration differences (8) for endogenous GIP in conscious dogs. Measurement of peripheral GIP in uremic subjects or patients with chronic renal failure pointed to the kidney as being important for the removal of GIP (9,10), as did the finding that GIP clearance was reduced in nephrectomized rats (11). More direct evidence of renal involvement was obtained from determination of arteriovenous concentration differences in man (11) and dog (9). However, because all these studies relied on antisera that were directed toward the central or COOH-terminal sequence in GIP, no conclusions can be drawn regarding clearance of intact biologically active GIP.

Based on the pivotal role of DPP IV in inactivating GLP-1, DPP IV inhibitors have been proposed as a new therapy for the treatment of diabetes (12,13). Several animal studies have been published showing that this approach does lead to an improvement in glucose tolerance (14,15,16), but—although a role for GLP-1 has been demonstrated—the contribution of GIP has only been speculated.

The present study was undertaken to assess the effect of DPP IV in regulating the biological activity of exogenously infused GIP and to see whether inhibition of the enzyme will enhance the insulinotropic effect of the peptide. In addition, simultaneous blood samples were collected during the infusions to allow determination of arteriovenous concentration differences across different organs using radioimmunoassays (RIAs) specific for each end of the molecule to identify the sites of elimination of circulating GIP. The ability of this approach to reveal metabolic degradation of GIP has been validated previously by high-performance liquid chromatography (HPLC) (6).


   RESEARCH DESIGN AND METHODS
TOP
 ABSTRACT
 INTRODUCTION
 RESEARCH DESIGN AND METHODS
RESULTS
 DISCUSSION
 REFERENCES
 
Anesthetized pig.
Nonfasted Danish LYY-strain pigs (33–40 kg) were used. After premedication with midazolam (0.5 mg/kg, Dormicum; Roche, Basel, Switzerland) and ketamine (10 mg/kg, Ketaminol; Veterinaria AG, Zurich, Switzerland), animals were anesthetized with intravenous administration of {alpha}-chloralose (66 mg/kg; Merck, Darmstadt, Germany) and ventilated with intermittent positive pressure. Catheters were placed in the right carotid artery for sampling of arterial blood, in a vein in the left ear for peptide infusion, and in a vein in the right ear for administration of glucose and valine-pyrrolidide. In addition, catheters were placed in the pulmonary artery and in the renal, hepatic, femoral, and portal veins as described previously (17). After surgical preparation, animals were heparinized and left undisturbed for 30 min. Anesthesia was maintained with administration of additional chloralose as necessary.

Six animals were used, each receiving two intravenous infusions of GIP, one before and one after administration of valine-pyrrolidide. Synthetic porcine GIP (Peninsula Laboratories Europe, St. Helens, U.K.), dissolved in 0.9% NaCl containing 1% human serum albumin (Behringwerke, Marburg, Germany) was given as a bolus dose of 0.75 nmol followed by an infusion at a rate of 2 pmol · kg–1 · min–1 for 57 min using a syringe pump, commencing at time 0. An intravenous infusion of glucose (0.2 g/kg; 50% solution) was administered between minutes 38 and 47. Arterial blood samples (4 ml) were collected at -10, 0, 10, 20, 30, 32, 35, 37, 42, 47, 49, 52, 54, and 57 min from the start of the infusion. After 57 min, the GIP infusion was stopped, and further blood samples were collected at 5, 10, 15, 20, 30, 45, 60, and 90 min. To allow determination of arteriovenous GIP concentration differences, simultaneous blood samples (2 ml) were collected from the pulmonary artery and the renal, hepatic, femoral, and portal veins at minutes 30 and 35. At 90 min after the cessation of the first GIP infusion, valine-pyrrolidide (300 µmol/kg, dissolved in 0.9% NaCl; a gift from Drs. R. D. Carr and L. Christiensen, Novo Nordisk, Bagsværd, Denmark) was given as a bolus intravenous injection over 2 min. Blood samples were collected at 15 and 30 min, after which the second GIP infusion was started and the protocol was repeated for blood sampling and glucose infusion. The volume of blood collected over the entire procedure amounted to 204 ml, which, for a 40-kg pig, is 5.1% of the blood volume. This blood loss has previously been shown not to affect blood pressure or heart rate (17).

Due to the long survival time of the inhibitor, it is impossible to use a conventional crossover design whereby some animals received GIP together with valine-pyrrolidide as the first infusion. This raises the possibility that the first GIP/glucose infusion may influence the response to the second GIP/glucose infusion. Therefore, three additional animals were given two successive GIP/glucose infusions as above, but without the infusion of valine-pyrrolidide, and blood samples were collected from the carotid artery.

Blood glucose was measured immediately after collection of samples (One Touch II; Lifescan, Lyngby, Denmark). Blood samples were collected into chilled tubes containing EDTA (7.4 mmol/l final concentration), aprotinin (500 kallikrein inhibitory equivalents ([KIE]/ml blood; Novo Nordisk, Bagsværd, Denmark), and valine-pyrrolidide (0.01 mmol/l final concentration) for hormonal analysis and into heparinized tubes for DPP IV activity determination. Samples were kept on ice until centrifugation at 4°C. Plasma was separated and stored at -20°C until analysis. To verify that the GIP in the infusate remained intact, samples from the infusate were collected before and after the experiment for later HPLC analysis (6).

DPP IV activity determination.
Plasma DPP IV activity was determined by Dr. M. Wilken (Novo Nordisk A/S, Bagsværd, Denmark) using a bioassay method that is based on the fact that DPP IV is the sole enzyme responsible for NH2-terminal degradation of GLP-1 in plasma (4,18). Briefly, GLP-1(7–37) (5 µl, 100 fmol) was added to plasma samples (95 µl) and incubated for 1 h at 37°C, after which the amount of GLP-1 that remained undegraded was determined by an enzyme-linked immunosorbent assay method using a monoclonal antibody (Mab26.1) that is specific for the intact peptide. As a reference, GLP-1 (5 µl, 100 fmol) was added to heat-inactivated plasma (95 µl), which contained 0.01 mmol/l valine-pyrrolidide and 500 KIE/ml aprotinin. By defining the DPP IV activity in the reference as zero and comparing the amount of intact GLP-1 in the samples with the reference, the DPP IV activity was calculated relative to the reference and expressed as a percentage. The minimum detectable difference in DPP IV activity was 3.2%.

Hormonal analysis.
Total GIP was measured using the COOH-terminally directed antiserum R65 (19,20), which reacts fully with intact GIP and the NH2-terminally truncated metabolite GIP(3–42) but not with the so-called 8-kDa GIP, of which the chemical nature and relation to GIP secretion is uncertain. The assay has a detection limit of 1.2 pmol/l. Intact, biologically active GIP was measured using a newly developed assay (6). Briefly, antibodies were raised by immunizing rabbits with the synthetic sequence GIP(1–10)-cys (Genosys Biotechnologies (Europe), Cambridge, U.K.), coupled to keyhole limpet hemocyanin using m-maleimidobenzoyl-N-hydroxysuccinimide ester, as described by Dyrberg and Kofod (21). Antiserum 98171 could be used in a final dilution of 1:40,000 and endows the assay with a detection limit of ~5 pmol/l and an ED50 of 48 pmol/l. The assay is specific for the intact NH2-terminus of GIP and crossreacts less than 0.1% with GIP(3–42) or with the structurally related peptides GLP-1(7–36)-amide, GLP-1(9–36)-amide, GLP-2(1–33), GLP-2(3–33), or glucagon at concentrations of up to 100 nmol/l. For both assays, human GIP (Peninsula Laboratories Europe, St. Helens, Merseyside, U.K.) was used as standard; radiolabeled GIP was obtained from Amersham Pharmacia Biotech (Little Chalfont, Buckinghamshire, U.K.). Separation of bound peptide from free peptide was achieved using plasma-coated charcoal (22). Plasma samples were extracted with ethanol (70% vol/vol; final concentration), resulting in recoveries of 85% and intra-assay variations of <6%.

Insulin immunoreactivity was measured in unextracted plasma using antiserum 2004 (23), and glucagon immunoreactivity was determined after ethanol extraction, using the COOH-terminally directed antiserum 4305, which measures glucagon of pancreatic origin (23).

Calculations and statistical analysis.
The plasma t1/2 of GIP was calculated by loge-linear regression analysis of peptide concentrations (after subtraction of endogenous values) in samples collected after the end of the infusion. The metabolic clearance rate (MCR) of GIP was calculated from the actual infusion rate for each animal divided by the plateau plasma concentration, after subtraction of basal values. Results are expressed per kilogram body weight. The incremental area under the curve (AUC) for glucose and insulin were calculated using the trapezoidal method, after subtraction of the basal concentrations measured in samples before the start of each GIP infusion. Additionally, for the period immediately after the glucose load, the AUCs were calculated after subtraction of the concentrations in the 35-min samples collected before administration of glucose. The disappearance rate (k) for glucose was calculated using the formula k = 0.693/t1/2.

For each animal, the extraction of exogenous GIP across the hind limb, kidney, and portal bed was calculated as a ratio, defined as

([GIP-IR]carotid artery – [GIP-IR]vein)/[GIP-IR]carotid artery

where [GIP-IR]carotid artery is the concentration of GIP immunoreactivity in the carotid artery and [GIP-IR]vein is the concentration of GIP immunoreactivity in the vein.

For the liver, the hepatic blood supply was calculated on the assumption that 75% originates from the portal vein and 25% is of arterial origin. The extraction across the lungs was defined as

([GIP-IR]pulmonary artery – [GIP-IR]carotid artery)/[GIP-IR]pulmonary artery

where [GIP-IR]pulmonary artery is the concentration of GIP immunoreactivity in the pulmonary artery.

Data are expressed as means ± SE and were analyzed using GraphPAD InStat software (version 1.13; San Diego, CA) and Statistica software (StatSoft, Tulsa, OK). Two-factor analysis of variance (ANOVA) for repeated measures with post-hoc analysis was used to analyze time-course curves before and after DPP IV inhibition. The t1/2, k, AUC, arteriovenous concentration differences, and MCR were analyzed using ANOVA (when appropriate) and two-tailed Student’s t tests for paired and nonpaired data. One-sample two-tailed tests were used to evaluate organ extractions. P < 0.05 was considered significant.


   RESULTS
TOP
 ABSTRACT
 INTRODUCTION
 RESEARCH DESIGN AND METHODS
 RESULTS
DISCUSSION
 REFERENCES
 
Comparison of two successive GIP infusions.
Similar concentrations of intact and total GIP were measured during the first and second infusions of GIP alone (Fig. 1), and the percentage of GIP that remained intact was unchanged (13.2 ± 0.4 and 11.9 ± 0.5%, first and second infusions, respectively). Similarly, there were no significant differences in blood glucose or insulin profiles after the first GIP/glucose infusion compared with the second (Fig. 1; glucose k, 10.5 ± 2.8 vs. 9.8 ± 0.9%/min).



View larger version (23K):
[in this window]
[in a new window]
  		FIG. 1. Plasma GIP immunoreactivity, measured with COOH-terminally and NH2-terminally directed RIAs, blood glucose, and plasma insulin concentrations during and after two successive intravenous infusions of GIP (2 pmol · kg–1 · min–1, 57 min). Animals received the first infusion of GIP ({square}), and the second infusion ({blacksquare}) was started 90 min after the end of the first infusion. Glucose infusions (0.2 g/kg) were administered during minutes 38–47 of each GIP infusion. Data are means ± SE; n = 3. The horizontal arrows indicate the periods of the infusions. There are no significant differences between the two infusions for any of the parameters measured.

 
DPP IV inhibition.
Administration of valine-pyrrolidide (300 µmol/kg) completely inhibited plasma DPP IV activity for the remainder of the experiment.

GIP pharmacokinetics.
HPLC analysis of infusate samples confirmed that the GIP eluted as a single peak with the same retention time as intact GIP(1–42) and that no degradation took place in the infusate during the course of the experiment.

Plasma GIP concentrations measured by NH2-terminal assay (intact peptide) were lower (P < 0.001) at all points than concentrations determined with the COOH-terminal assay (intact + NH2-terminally degraded peptide). Treatment with the DPP IV inhibitor had no effect on COOH-terminal immunoreactivity, but NH2-terminal GIP levels increased significantly (P < 0.001) after valine-pyrrolidide (Fig. 2). During infusion of GIP alone, the intact peptide accounted for 14.5 ± 0.9% of total immunoreactivity (AUC), and this was increased to 40.9 ± 2.5% (P < 0.0001) in the presence of valine-pyrrolidide.



View larger version (16K):
[in this window]
[in a new window]
  		FIG. 2. Plasma GIP immunoreactivity in blood samples from the carotid artery, measured with COOH-terminally (•) and NH2-terminally ({square}) directed RIAs. Animals received a 57-min intravenous infusion of GIP (2 pmol · kg–1 · min–1). Valine-pyrrolidide (val-pyd; 300 µmol/kg) was given 90 min after the end of the first GIP infusion, and after an additional 30 min, the second GIP infusion was started. Glucose infusions (0.2 g/kg) were given during minutes 38–47 of each GIP infusion. Data are means ± SE; n = 6. The horizontal arrows indicate the periods of the infusions. NH2-terminal immunoreactivity is lower than COOH-terminal immunoreactivity at all time points (P < 0.001). The COOH-terminal GIP profiles before (minutes 0–127) and after (minutes 180–327) the inhibitor are not significantly different, but the NH2-terminal GIP profile is increased after the inhibitor (P < 0.001).

 
When GIP was infused alone, the plasma t1/2 of the intact peptide (determined by NH2-terminal RIA) was shorter than that determined by COOH-terminal assay (3.3 ± 0.3 vs. 13.2 ± 0.5 min; P < 0.001). However, in the presence of valine-pyrrolidide, the NH2-terminal t1/2 was prolonged to 8.1 ± 0.6 min (P < 0.001 compared with NH2-terminal t1/2 for GIP alone). The COOH-terminal t1/2 during DPP IV inhibition (11.5 ± 0.8 min) did not differ from the COOH-terminal t1/2 before administration of valine-pyrrolidide.

There was a significant difference in the MCR determined with the two assays when GIP was infused alone (7.19 ± 0.93 and 51.45 ± 8.91 ml · kg–1 · min–1, COOH-terminal and NH2-terminal, respectively; P < 0.001), but this difference disappeared when GIP was infused in the presence of valine-pyrrolidide (8.31 ± 1.01 and 19.83 ± 2.73 ml · kg–1 · min–1, COOH-terminal and NH2-terminal, respectively; P > 0.05). The COOH-terminal MCR was unaffected by coinfusion of the inhibitor, whereas the NH2-terminal MCR was significantly shortened (P < 0.001) compared with GIP alone.

GIP extraction.
Plasma concentrations measured during the GIP infusion without and with DPP IV inhibition are shown in Table 1, and the regional extraction is shown in Fig. 3. No extraction across the lungs was detected with either assay when either GIP was infused alone or in the presence of valine-pyrrolidide. Significant extraction across the hind limb was detected with both NH2-terminal and COOH-terminal assays when GIP was infused alone, and there was a trend (P = 0.07) for this to be higher when determined with the NH2-terminal assay. There was no effect on COOH-terminal hind-limb extraction when valine-pyrrolidide was coinfused, but NH2-terminal extraction was significantly (P < 0.02) reduced. Similarly, both assays detected significant extraction across the renal bed when GIP was infused alone, but in this case, the extraction detected by both assays was reduced by DPP IV inhibition (P < 0.02). Extraction of GIP across the splanchnic bed was detected only by the COOH-terminal assay in the absence of DPP IV inhibition, but when the inhibitor was included, no extraction was detected with either assay. The significant hepatic extraction detected by the NH2-terminal assay was abolished by coinfusion of valine-pyrrolidide (P < 0.002). In contrast, no significant hepatic extraction could be detected with the COOH-terminal assay when GIP was infused alone, but a small extraction was detected after DPP IV inhibition.


View this table:
[in this window]
[in a new window]
  		TABLE 1 Plasma concentrations of exogenous GIP during intravenous infusion

 


View larger version (23K):
[in this window]
[in a new window]
  		FIG. 3. Regional extraction of GIP during intravenous infusion determined by COOH-terminal (R65) and NH2-terminal (171) RIAs before and after administration of valine-pyrrolidide (val-pyd). The extraction is the ratio calculated by dividing the arteriovenous concentration difference by the arterial concentration. The hepatic blood supply was calculated on the assumption that 75% originates from the portal vein and 25% is of arterial origin. Data are means ± SE of individual extractions across each organ calculated for each animal; n = 6. * P < 0.05, ** P < 0.01, *** P < 0.001, difference from 0. A positive balance indicates a net extraction of GIP.

 
GIP pharmacodynamics.
GIP concentrations determined with the COOH-terminal assay were still decreasing (because of the previous infusion) when the inhibitor was administered and continued to decrease after valine-pyrrolidide was administered (39 ± 2 pmol/l before the inhibitor, 35 ± 1 pmol/l at 15 min, and 31 ± 3 pmol/l at 30 min after the inhibitor). In contrast, valine-pyrrolidide stopped and even reversed the decrease in NH2-terminal GIP concentrations (8 ± 1 pmol/l before the inhibitor, 9 ± 1 pmol/l at 15 min, and 11 ± 1 pmol/l at 30 min after the inhibitor), reflecting elevated levels of intact endogenous GIP (Fig. 2). This was accompanied by a small reduction in blood glucose (5.4 ± 0.4 mmol/l before the inhibitor, decreasing to 4.8 ± 0.4 mmol/l at 30 min after the inhibitor, P < 0.005 [Fig. 4]; AUC -6 ± 1 min · mmol/l, P < 0.01 vs. 0), a transient trend toward an increase in insulin (58 ± 14 pmol/l before the inhibitor, peaking at 77 ± 21 pmol/l at 15 min after the inhibitor, P > 0.05 [Fig. 5]; AUC 321 ± 180 min · pmol/l, P > 0.05 vs. 0), and a decrease in plasma glucagon (14 ± 2 pmol/l before the inhibitor, decreasing to 10 ± 2 pmol/l at 15 min after the inhibitor, P < 0.01 [Fig. 6]).



View larger version (13K):
[in this window]
[in a new window]
  		FIG. 4. Blood glucose concentrations measured before, during, and after intravenous infusions of GIP (2 pmol · kg–1 · min–1) given alone (minutes 0–150) or during (minutes 150–327) DPP IV inhibition. Valine-pyrrolidide (val-pyd; 300 µmol/kg) was given 90 min after the end of the first GIP infusion, and after a additional 30 min, the second GIP infusion was started. Glucose infusions (0.2 g/kg) were given during minutes 38–47 of each GIP infusion. Data are means ± SE; n = 6. The horizontal arrows indicate the periods of the infusions. The blood glucose profile obtained after the glucose load before valine-pyrrolidide treatment (minutes 42–147) is significantly higher (P < 0.05) than that obtained after administration of valine-pyrrolidide (minutes 222–327).

 


View larger version (14K):
[in this window]
[in a new window]
  		FIG. 5. Plasma insulin concentrations measured before, during, and after intravenous infusions of GIP (2 pmol · kg–1 · min–1) given alone (minutes 0–150) or during (minutes 150–327) DPP IV inhibition. Valine-pyrrolidide (val-pyd; 300 µmol/kg) was given 90 min after the end of the first GIP infusion, and after an additional 30 min, the second GIP infusion was started. Glucose infusions (0.2 g/kg) were given during minutes 38–47 of each GIP infusion. Data are means ± SE; n = 6. The horizontal arrows indicate the periods of the infusions. The plasma insulin profile obtained before valine-pyrrolidide treatment (minutes 0–147) is significantly lower (P < 0.02) than that obtained after administration of valine-pyrrolidide (minutes 180–327).

 


View larger version (13K):
[in this window]
[in a new window]
  		FIG. 6. Plasma glucagon concentrations measured before, during, and after intravenous infusions of GIP (2 pmol · kg–1 · min–1) given alone (minutes 0–150) or during (minutes 150–327) DPP IV inhibition. Valine-pyrrolidide (val-pyd; 300 µmol/kg) was given 90 min after the end of the first GIP infusion, and after an additional 30 min, the second GIP infusion was started. Glucose infusions (0.2 g/kg) were given during minutes 38–47 of each GIP infusion. Data are means ± SE; n = 6. The horizontal arrows indicate the periods of the infusions. There are significant differences (P < 0.02) in the plasma glucagon profiles obtained before (minutes 0–147) and after (minutes 180–327) valine-pyrrolidide treatment.

 
Comparison of two successive GIP infusions in the absence and presence of DPP IV inhibition.
In comparison with the first infusion of GIP alone, the glucose excursion in response to intravenous administration of glucose was reduced (P < 0.05) when GIP was infused in the presence of valine-pyrrolidide, resulting in a smaller AUC in the period after the glucose load (Fig. 4; Table 2). This was mainly due to the increased disappearance rate for glucose in the presence of the inhibitor (GIP alone, 8.0 ± 0.5%/min; GIP + valine-pyrrolidide, 15.5 ± 2.2%/min; P < 0.01). The presence of the inhibitor resulted in a small increase in insulin secretion even before the start of the glucose infusion, and this was greatly potentiated (P < 0.02) after the glucose load was administered (Fig. 5, Table 2). Infusion of GIP alone had no effect on plasma glucagon before the glucose load, but there was a modest suppression in response to the glucose (from 10.3 ± 1.5 to 8.2 ± 1.6 pmol/l; P < 0.001). Plasma glucagon concentrations decreased after valine-pyrrolidide administration (from 13.5 ± 1.9 to 10.0 ± 1.6 pmol/l; P < 0.05), but this decrease was reversed after the GIP infusion started, reaching 16.2 ± 1.7 pmol/l. DPP IV inhibition potentiated the decrease in glucagon in response to the glucose challenge (to 6.5 ± 0.5 pmol/l; P < 0.01 [Fig. 6]), but after the second GIP infusion was ended, plasma glucagon levels rebounded and exceeded basal values (P < 0.05).


View this table:
[in this window]
[in a new window]
  		TABLE 2 Incremental AUCs for glucose and insulin attained during infusions of GIP alone or with inhibition of DPP IV

 

   DISCUSSION
TOP
 ABSTRACT
 INTRODUCTION
 RESEARCH DESIGN AND METHODS
 RESULTS
 DISCUSSION
REFERENCES
 
In the present study, valine-pyrrolidide was used to examine the role of DPP IV in regulating the degradation and biological activity of GIP. The results show that inhibition of the enzyme reduces the clearance of the peptide and results in increased levels of intact, biologically active GIP. This is associated with enhancement of its insulinotropic activity and results in a reduction in the glucose excursion after an intravenous glucose load.

Although it has been known since 1993 that GIP is susceptible to degradation by DPP IV in vitro (3), producing a metabolite, GIP(3–42), which lacks significant insulinotropic activity (5), only a few studies have provided specific data pointing to the relevance of the enzyme for in vivo regulation of GIP bioactivity. Thus, Kieffer et al. (24), using HPLC to follow degradation of radiolabeled GIP in rats, showed that the peptide was rapidly cleaved at the NH2-terminus in normal animals but not in a DPP IV–deficient strain, suggesting a role for the enzyme in vivo. Further support came from a study using a DPP IV–resistant GIP analog, which resulted in improved glycemic control compared with native GIP (25,26). However, direct assessment of the extent of GIP degradation has been hampered by the lack of specific analyses, because all reported assays also crossreact with the NH2-terminally truncated metabolite GIP(3–42). The recent development of an NH2-terminally directed GIP assay (6) coupled with the use of a specific DPP IV inhibitor has, for the first time, made it possible to follow GIP degradation in vivo and fully assess the degree to which DPP IV influences the peptide’s biological activity.

During infusion of GIP alone, the intact peptide, as determined using the NH2-terminal assay, accounts for only ~15% of the overall immunoreactivity. This suggests that greater NH2-terminal degradation may occur in pigs, because a recent study in humans, using the same assays, showed that intact GIP accounts for 37% of total immunoreactivity after intravenous infusion (6). The plasma t1/2 for both NH2-terminal and COOH-terminal immunoreactivity were also lower in the pig (3 and 13 min, respectively) compared with humans (7 and 17 min, respectively) (6), which may indicate that GIP is cleared more quickly in pigs than in humans. Inclusion of the DPP IV inhibitor significantly increased the proportion of intact GIP, but it is worth noting that full protection was not attained. This is in contrast to the situation for GLP-1, in which valine-pyrrolidide completely prevented NH2-terminal degradation of exogenous GLP-1 (27), but the reason for this difference is not clear. The same inhibitor was used in both studies, and inhibition of the enzyme was complete for the duration of the experiment. The possibility exists, therefore, that GIP may be a substrate for other enzymes that cleave the peptide in the NH2-terminal portion and are not inhibitable by valine-pyrrolidide, an inhibitor known to be highly specific for DPP IV.

Sites of GIP extraction were identified from differences in arteriovenous concentrations across the various organ beds. Changes in these differences after DPP IV inhibition were interpreted to indicate a role of the enzyme at these sites. However, it cannot be totally excluded that DPP IV inhibition per se could lead to a change in blood flow across the different organ beds.

Determination of the arteriovenous concentration differences revealed that the kidney is a major site of GIP extraction, in agreement with previously reported findings in humans (9,10,11), dogs (9), and rats (11). This was detected with both assays, consistent with the peptide being filtered and undergoing proteolysis within the renal tubules. However, DPP IV inhibition significantly reduced renal clearance detected with both NH2-terminal and COOH-terminal assays. It is likely that GIP is degraded by DPP IV in the renal capillary endothelium. This would be prevented by the inhibitor, thereby explaining the reduced clearance detected by the NH2-terminal assay. The reduced clearance detected with the COOH-terminal assay can be interpreted as indicating that NH2-terminal cleavage is necessary before a later stage of degradation can occur; thus, preservation of the NH2-terminus of the peptide possibly affords protection from other subsequent enzymatic attacks.

The other major site of GIP removal is the liver, where the NH2-terminal assay revealed that cleavage in the NH2-terminal part of the peptide occurs. However, no significant extraction is detected with the COOH-terminal assay, confirming the negative findings of earlier studies in rats (7) and dogs (7,8), in which assays of similar specificity were used. In the presence of the enzyme inhibitor, NH2-terminal extraction decreased to undetectable levels, indicating the involvement of DPP IV. This is fully consistent with the location of the enzyme in high concentration on hepatocytes (28). However, with DPP IV inhibition, a modest COOH-terminal extraction was detected. This is more difficult to explain, but because direct comparison of COOH-terminal extraction before and after the inhibitor revealed no effect of valine-pyrrolidide, it may simply be a coincidental finding.

Extraction across the hind limb, representing degradation by connective, supportive, and muscular tissues and their capillary supply, was detected by both assays. This was reduced by valine-pyrrolidide, indicating a role of DPP IV in sites such as the capillary endothelium (29). However, significant NH2-terminal extraction was still detectable, which again suggests that other enzymes are involved. It has been suggested that neutral endopeptidase 24.11 may play a role in GLP-1 degradation (17,30), but this seems less likely in the case of GIP because GIP was reported to be a poor substrate for the endopeptidase (30).

Thus, the main sites of GIP extraction were identified as the kidney, liver, and extremities. If one takes into account the relative cardiac output (passing through the lungs and being distributed with 25% to the kidneys, 33% to the liver, and 25% to the extremities), then the liver is more important in terms of whole-body clearance of exogenous GIP, followed by the kidneys and the extremities. However, because the endogenous peptide travels first to the liver before being distributed to the remainder of the body, the liver becomes quantitatively much more important in terms of extraction of endogenous GIP.

The use of a DPP IV inhibitor clearly confirms that the enzyme does play an important role in the in vivo regulation of GIP activity, as is also seen with the other incretin hormone, GLP-1 (27), with inhibition of the enzyme leading to a marked improvement in the insulinotropic action of GIP. It cannot be totally excluded that the experimental design per se may contribute to this effect, in that increased insulin secretion occurred with the second GIP/glucose infusions (in the presence of the inhibitor). However, this seems unlikely, because the glucose and insulin responses to two successive infusions of GIP alone were of a similar magnitude.

Valine-pyrrolidide treatment resulted in an increase in the proportion of intact biologically active GIP and was accompanied by a transient increase in insulin secretion even before the glucose load. This could be due to the increased levels of intact GIP having a modest stimulatory effect on the ß-cell at a time when blood glucose levels may have been just above the threshold below which GIP ceases to be insulinotropic (31), but it is also likely that enhanced levels of intact endogenous GLP-1 contribute to the effect. After the glucose load, the presence of valine-pyrrolidide significantly enhanced insulin secretion and led to a reduction in the glucose excursion, with a doubling of the disappearance rate for glucose. GIP also lowers blood glucose by directly influencing pancreatic secretion (32,33), but there is evidence that it also has extrapancreatic effects. GIP receptors have been identified in several tissues (34,35), and there are reports that GIP promotes glucose uptake in muscle (36) and fat (37,38,39). GIP has also been indicated to increase the affinity of the insulin receptor (40), giving rise to the suggestion that GIP can modulate the effects of insulin by directly altering target tissue sensitivity to insulin (38), thereby contributing to the effective uptake of glucose postprandially (39,41). GIP has been shown to enhance insulin-mediated glucose disposal in sheep (42), rats (25), and mice (26), although a study in humans failed to show an effect (43). Therefore, it is possible that the increased glucose clearance seen in the present study in pigs after DPP IV inhibition may be due to the increased levels of intact GIP stimulating insulin secretion and directly or indirectly improving peripheral glucose uptake.

Several studies have now examined the effects of DPP IV inhibitors on glucose homeostasis and demonstrated that they result in improved glucose tolerance (14,15,16). Although the role of GLP-1 in mediating this effect has been shown by specific analysis of intact and total GLP-1, revealing that DPP IV inhibition protects the peptide from NH2-terminal degradation (15,27), the lack of suitable methodology has, until now, meant that the contribution of GIP could only be a matter of speculation. As the present study shows, DPP IV inhibition does prevent at least part of the NH2-terminal degradation of GIP, resulting in increased concentrations of the bioactive form of the peptide and improving both insulin secretion and glucose excursion. It can, therefore, be concluded that the mechanism of action of DPP IV inhibitors involves protection of the intact forms of both incretin hormones.

GLP-1 inhibits glucagon secretion both in vivo and in isolated pancreata (44), and basal levels of GLP-1 have been reported to play a role in the tonic inhibition of the pancreatic {alpha}-cell, either directly or indirectly via somatostatin or insulin (45). In contrast, GIP has a glucagonotropic effect in isolated {alpha}-cells (46), isolated pancreatic islets (47,48), perfused rat pancreata (49), and porcine pancreata (J. J. Holst, unpublished observations) and after intravenous infusion in rats (50) and humans (51). In the present study, infusion of GIP alone had no effect on plasma glucagon concentrations, possibly because concentrations of the intact peptide were insufficient. Moreover, a fall in glucagon concentrations was seen when the DPP IV inhibitor was given. This was also seen in our previous study with GLP-1 (27) and could be attributed to the increased concentrations of endogenous intact GLP-1 arising after DPP IV inhibition having an inhibitory effect on {alpha}-cell secretion. However, once the second GIP infusion was started, the increased levels of intact GIP arising from inhibition of DPP IV activity reverse the decrease by stimulating glucagon secretion. The apparent paradox, whereby GIP potentiates the reduction in glucagon levels in response to the glucose load, could be a consequence of an indirect effect caused by the inhibitory effect of GIP-induced insulin secretion (52).

In summary, our results demonstrate that GIP is degraded in a tissue-specific manner, with the liver, kidney, and extremities being the main sites of extraction. DPP IV is not the sole enzyme capable of inactivating GIP, but the use of a DPP IV inhibitor confirms that the enzyme does play a major role in regulating the insulinotropic and antihyperglycemic effects of GIP in vivo.


   ACKNOWLEDGMENTS
 
This study was supported by grants from the Danish Medical Research Council, the Danish Biotechnology Program, and Grosserer Ernst Fischers Mindelegat.

We thank Lene Albæk and Anne-Grethe Juul for their technical assistance.


   FOOTNOTES
 
Address correspondence and reprint requests to Dr. Carolyn F. Deacon, Department of Medical Physiology, The Panum Institute, Blegdamsvej 3, DK-2200 Copenhagen N, Denmark. E-mail: deacon{at}mfi.ku.dk.

Received for publication 7 September 2000 and accepted in revised form 11 January 2001.

ANOVA, analysis of variance; AUC, area under the curve; DPP IV, dipeptidyl peptidase IV; GIP, glucose-dependent insulinotropic peptide; GLP-1, glucagon-like peptide 1; HPLC, high-performance liquid chromatography; KIE, kallikrein inhibitory equivalents; MCR, metabolic clearance rate; RIA, radioimmunoassay.


   REFERENCES
TOP
 ABSTRACT
 INTRODUCTION
 RESEARCH DESIGN AND METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 

  1. Krarup T, Holst JJ, Larsen KL: Responses and molecular heterogeneity of IR-GIP after intraduodenal glucose and fat. Am J Physiol 249:E195–E200, 1985[Abstract/Free Full Text]
  2. Nauck MA, Bartels E, Ørskov C, Ebert R, Creutzfeldt W: Additive insulinotropic effects of exogenous synthetic human gastric inhibitory polypeptide and glucagon-like peptide-1-(7–36) amide infused at near-physiological insulinotropic hormone and glucose concentrations. J Clin Endocrinol Metab 76:912–917, 1993[Abstract]
  3. Mentlein R, Gallwitz B, Schmidt WE: Dipeptidyl peptidase IV hydrolyses gastric inhibitory polypeptide, glucagon-like peptide-1 (7–36)amide, peptide histidine methionine and is responsible for their degradation in human serum. Eur J Biochem 214:829–835, 1993[Medline]
  4. Pauly RP, Rosche F, Wermann M, McIntosh CH, Pederson RA, Demuth HU: Investigation of glucose-dependent insulinotropic polypeptide-(1–42) and glucagon-like peptide-1-(7–36) degradation in vitro by dipeptidyl peptidase IV using matrix-assisted laser desorption/ionization-time of flight mass spectrometry: a novel kinetic approach. J Biol Chem 271:23222–23229, 1996[Abstract/Free Full Text]
  5. Jörnvall H, Carlquist M, Kwauk S, Otte SC, McIntosh CH, Brown JC, Mutt V: Amino acid sequence and heterogeneity of gastric inhibitory polypeptide (GIP). FEBS Lett 123:205–210, 1981[Medline]
  6. Deacon CF, Nauck MA, Meier J, Hücking K, Holst JJ: Degradation of endogenous and exogenous gastric inhibitory polypeptide (GIP) in healthy and in type 2 diabetic subjects as revealed using a new assay for the intact peptide. J Clin Endocrinol Metab 85:3575–3581, 2000[Abstract/Free Full Text]
  7. Hanks JB, Andersen DK, Wise JE, Putnam WS, Meyers WC, Jones RS: The hepatic extraction of gastric inhibitory polypeptide and insulin. Endocrinology 115:1011–1018, 1984[Abstract]
  8. Chap Z, O’Dorisio TM, Cataland S, Field JB: Absence of hepatic extraction of gastric inhibitory polypeptide in conscious dogs. Dig Dis Sci 32:280–284, 1987[Medline]
  9. O’Dorisio TM, Sirinek KR, Mazzaferri EL, Cataland S: Renal effects on serum gastric inhibitory polypeptide (GIP). Metabolism 26:651–656, 1977[Medline]
  10. Sirinek KR, O’Dorisio TM, Gaskill HV, Levine BA: Chronic renal failure: effect of hemodialysis on gastrointestinal hormones. Am J Surg 148:732–735, 1984[Medline]
  11. Jorde R, Burhol PG, Gunnes P, Schulz TB: Removal of IR-GIP by the kidneys in man, and the effect of acute nephrectomy on plasma GIP in rats. Scand J Gastroenterol 16:469–471, 1981[Medline]
  12. Deacon CF, Nauck MA, Toft-Nielsen M, Pridal L, Willms B, Holst JJ: Both subcutaneously and intravenously administered glucagon-like peptide-1 are rapidly degraded from the NH2-terminus in type II diabetic patients and in healthy subjects. Diabetes 44:1126–1131, 1995[Abstract]
  13. Holst JJ, Deacon CF: Inhibition of the activity of dipeptidyl peptidase IV as a treatment for type 2 diabetes. Diabetes 47:1663–1670, 1998[Abstract]
  14. Pederson RA, White HA, Schlenzig D, Pauly RP, McIntosh CH, Demuth HU: Improved glucose tolerance in Zucker fatty rats by oral administration of the dipeptidyl peptidase IV inhibitor isoleucine thiazolidide. Diabetes 47:1253–1258, 1998[Abstract]
  15. Balkan B, Kwasnik L, Miserendino R, Holst JJ, Li X: Inhibition of dipeptidyl peptidase IV with NVP-DPP728 increases plasma GLP-1 (7–36 amide) concentrations and improves oral glucose tolerance in obese Zucker rats. Diabetologia 42:1324–1331, 1999[Medline]
  16. Pauly RP, Demuth HU, Rosche F, Schmidt J, White HA, Lynn F, McIntosh CH, Pederson RA: Improved glucose tolerance in rats treated with the dipeptidyl peptidase IV (CD26) inhibitor Ile-thiazolidide. Metabolism 48:385–389, 1999[Medline]
  17. Deacon CF, Pridal L, Klarskov L, Olesen M, Holst JJ: Glucagon-like peptide-1 undergoes differential tissue-specific metabolism in the anesthetized pig. Am J Physiol 271:E458–E464, 1996[Abstract/Free Full Text]
  18. Deacon CF, Johnsen AH, Holst JJ: Degradation of glucagon-like peptide-1 by human plasma in vitro yields an N-terminally truncated peptide that is a major endogenous metabolite in vivo. J Clin Endocrinol Metab 80:952–957, 1995[Abstract]
  19. Krarup T, Madsbad S, Moody AJ, Regeur L, Faber OK, Holst JJ, Sestoft L: Diminished immunoreactive gastric inhibitory polypeptide response to a meal in newly diagnosed type I (insulin-dependent) diabetics. J Clin Endocrinol Metab 56:1306–1312, 1983[Abstract]
  20. Krarup T, Holst JJ: The heterogeneity of gastric inhibitory polypeptide in porcine and human gastrointestinal mucosa evaluated with five different antisera. Regul Pept 9:35–46, 1984[Medline]
  21. Dyrberg T, Kofod H: Synthetic peptides as antigen. In Animal Virus Pathogenesis: A Practical Approach . Oldstone MBA, Ed. Oxford, IRL Press, 1990, p. 163–171
  22. Ørskov C, Holst JJ, Seier Poulsen S, Kirkegaard P: Pancreatic and intestinal processing of proglucagon in man. Diabetologia 30:974–981, 1987
  23. Ørskov C, Jeppesen J, Madsbad S, Holst JJ: Proglucagon products in the plasma of non-insulin dependent diabetics and nondiabetic controls in the fasting state and following oral glucose and intravenous arginine. J Clin Invest 87:415–423, 1991
  24. Kieffer TJ, McIntosh CHS, Pederson RA: Degradation of glucose-dependent insulinotropic polypeptide (GIP) and truncated glucagon-like peptide 1 (tGLP-1) in vitro and in vivo by dipeptidyl peptidase IV. Endocrinology 136:3585–3596, 1995[Abstract]
  25. O’Harte FPM, Mooney MH, Flatt PR: NH2-terminally modified gastric inhibitory polypeptide exhibits amino-peptidase resistance and enhanced antihyperglycemic activity. Diabetes 48:758–765, 1999[Abstract]
  26. O’Harte FPM, Mooney MH, Kelly CMN, Flatt PR: Improved glycaemic control in obese diabetic ob/ob mice using N-terminally modified gastric inhibitory polypeptide. J Endocrinol 165:639–648, 2000[Abstract]
  27. Deacon CF, Hughes TE, Holst JJ: Dipeptidyl peptidase IV inhibition potentiates the insulinotropic effect of glucagon-like peptide 1 in the anesthetized pig. Diabetes 47:764–769, 1998[Abstract]
  28. Elovson J: Biogenesis of plasma membrane glycoproteins: purification and properties of two rat liver membrane glycoproteins. J Biol Chem 255:5807–5815, 1980[Abstract/Free Full Text]
  29. Lojda Z: Studies on dipeptidyl(amino)peptidase IV (glycyl-proline-naphthylamidase). II. Blood vessels. Histochemistry 59:153–166, 1979[Medline]
  30. Hupe-Sodmann K, McGregor GP, Bridenbaugh R, Goke R, Goke B, Thole H, Zimmermann B, Voigt K: Characterisation of the processing by human neutral endopeptidase 24.11 of GLP-1(7–36) amide and comparison of the substrate specificity of the enzyme for other glucagon-like peptides. Regul Pept 58:149–156, 1995[Medline]
  31. Schmid R, Schusdziarra V, Aulehner R, Weigert N, Classen M: Comparison of GLP-1 (7–36amide) and GIP on release of somatostatin-like immunoreactivity and insulin from the isolated rat pancreas. Z Gastroenterol28:280–284, 1990
  32. Dupre J, Ross SA, Watson D, Brown JC: Stimulation of insulin secretion by gastric inhibitory polypeptide in man. J Clin Endocrinol Metab 37:826–828, 1973[Medline]
  33. Pederson RA: Gastric inhibitory polypeptide. In Gut Peptides: Biochemistry and Physiology . Walsh J, Dockray G, Eds. New York, Raven Press, 1994, p. 217–259
  34. Usdin TB, Mezey E, Button DC, Brownstein MJ, Bonner TI: Gastric inhibitory polypeptide receptor, a member of the secretin-vasoactive intestinal peptide receptor family, is widely distributed in peripheral organs and the brain. Endocrinology 133:2861–2870, 1993[Abstract]
  35. Yip RG, Boylan MO, Kieffer TJ, Wolfe MM: Functional GIP receptors are present on adipocytes. Endocrinology 139:4004–4007, 1998[Abstract/Free Full Text]
  36. O’Harte FPM, Gray AM, Flatt PR: Gastric inhibitory polypeptide and effects of glycation on glucose transport and metabolism in isolated mouse abdominal muscle. J Endocrinol 156:237–243, 1998[Abstract]
  37. Hauner H, Glatting G, Kaminska D, Pfeiffer EF: Effects of gastric inhibitory polypeptide on glucose and lipid metabolism of isolated rat adipocytes. Ann Nutr Metab 32:282–288, 1988[Medline]
  38. Oben J, Morgan L, Fletcher J, Marks V: Effect of the entero-pancreatic hormones gastric inhibitory polypeptide and glucagon-like peptide-1(7–36)amide on fatty acid synthesis in explants of rat adipose tissue. J Endocrinol 130:267–272, 1991[Abstract]
  39. Morgan LM: The role of gastrointestinal hormones in carbohydrate and lipid metabolism and homeostasis: effects of gastric inhibitory polypeptide and glucagon-like peptide-1. Biochem Soc Trans 26:216–222, 1998[Medline]
  40. Starich GH, Bar RS, Mazzaferri EL: GIP increases insulin receptor affinity and cellular sensitivity in adipocytes. Am J Physiol 249:E603–E607, 1985[Abstract/Free Full Text]
  41. Yip RGC, Wolfe MM: GIP biology and fat metabolism. Life Sci 66:91–103, 2000[Medline]
  42. Rose MT, Itoh F, Takahashi Y: Effect of glucose-dependent insulinotropic polypeptide on whole-body glucose utilization in sheep. Exp Physiol 83:783–792, 1988
  43. Service FJ, Heiling VJ, Go VL, Rizza RA: Lack of effect of gastric inhibitory polypeptide on hepatic and extrahepatic insulin action. J Clin Endocrinol Metab 70:1398–1402, 1990[Abstract]
  44. Ørskov C, Holst JJ, Nielsen OV: Effect of truncated glucagon-like peptide-1 [proglucagon-(78–107)amide] on endocrine secretion from pig pancreas, antrum and non-antral stomach. Endocrinology 123:2009–2013, 1988[Abstract]
  45. Schirra J, Sturm K, Leicht P, Arnold R, Göke B, Katschinski M: Exendin(9–39)amide is an antagonist of glucagon-like peptide-1(7–36)amide in humans. J Clin Invest 101:1421–1430, 1998[Medline]
  46. Ding WG, Renstrom E, Rorsman P, Buschard K, Gromada J: Glucagon-like peptide I and glucose-dependent insulinotropic polypeptide stimulate Ca2+-induced secretion in rat alpha-cells by a protein kinase A-mediated mechanism. Diabetes 46:792–800, 1997[Abstract]
  47. Bailey CJ, Wilkes LC, Conlon JM, Armstrong PH, Buchanan KD: Effects of gastric inhibitory polypeptide, vasoactive intestinal peptide and peptide histidine isoleucine on the secretion of hormones by isolated mouse pancreatic islets. J Endocrinol 125:375–379, 1990[Abstract]
  48. Opara EC, Go VL: Influence of gastric inhibitory polypeptide (GIP) and glucose on the regulation of glucagon secretion by pancreatic alpha cells. Regul Pept 32:65–73, 1991[Medline]
  49. Pederson RA, Brown JC: Interaction of gastric inhibitory polypeptide, glucose, and arginine on insulin and glucagon secretion from the perfused rat pancreas. Endocrinology 103:610–615, 1978[Abstract]
  50. Taminato T, Seino Y, Go Y, Inoue Y, Kadowaki S: Synthetic gastric inhibitory polypeptide: stimulatory effect on insulin and glucagon secretion in the rat. Diabetes 26:480–484, 1977[Abstract]
  51. Dupre J, Caussignac Y, McDonald TJ, Van Vliet S: Stimulation of glucagon secretion by gastric inhibitory polypeptide in patients with hepatic cirrhosis and hyperglucagonemia. J Clin Endocrinol Metab72:125–129, 1991
  52. Asplin CM, Paquette TL, Palmer JP: In vivo inhibition of glucagon secretion by paracrine beta cell activity in man. J Clin Invest 68:314–318, 1981

Add to CiteULike CiteULike   Add to Del.icio.us Del.icio.us   Add to Digg Digg   Add to Reddit Reddit   Add to Technorati Technorati    What's this?


This article has been cited by other articles:


Home page
 DiabetesHome page
B. J. Lamont and D. J. Drucker
Differential Antidiabetic Efficacy of Incretin Agonists Versus DPP-4 Inhibition in High Fat Fed Mice
Diabetes, January 1, 2008; 57(1): 190 - 198.
[Abstract] [Full Text] [PDF]

Home page
 Diabetes CareHome page
D. J. Drucker
Dipeptidyl Peptidase-4 Inhibition and the Treatment of Type 2 Diabetes: Preclinical biology and mechanisms of action
Diabetes Care, June 1, 2007; 30(6): 1335 - 1343.
[Full Text] [PDF]

Home page
 Eur J EndocrinolHome page
J. Ryskjaer, C. F Deacon, R. D Carr, T. Krarup, S. Madsbad, J. Holst, and T. Vilsboll
Plasma dipeptidyl peptidase-IV activity in patients with type-2 diabetes mellitus correlates positively with HbAlc levels, but is not acutely affected by food intake
Eur. J. Endocrinol., September 1, 2006; 155(3): 485 - 493.
[Abstract] [Full Text] [PDF]

Home page
 Am. J. Physiol. Endocrinol. Metab.Home page
C. F. Deacon, A. Plamboeck, M. M. Rosenkilde, J. de Heer, and J. J. Holst
GIP-(3-42) does not antagonize insulinotropic effects of GIP at physiological concentrations
Am J Physiol Endocrinol Metab, September 1, 2006; 291(3): E468 - E475.
[Abstract] [Full Text] [PDF]

Home page
 J. Biol. Chem.Home page
I. Alana, J. C. Parker, V. A. Gault, P. R. Flatt, F. P. M. O'Harte, J. P. G. Malthouse, and C. M. Hewage
NMR and Alanine Scan Studies of Glucose-dependent Insulinotropic Polypeptide in Water
J. Biol. Chem., June 16, 2006; 281(24): 16370 - 16376.
[Abstract] [Full Text] [PDF]

Home page
 Am. J. Physiol. Heart Circ. Physiol.Home page
L. A. Nikolaidis, D. Elahi, Y.-T. Shen, and R. P. Shannon
Active metabolite of GLP-1 mediates myocardial glucose uptake and improves left ventricular performance in conscious dogs with dilated cardiomyopathy
Am J Physiol Heart Circ Physiol, December 1, 2005; 289(6): H2401 - H2408.
[Abstract] [Full Text] [PDF]

Home page
 J. Pharmacol. Exp. Ther.Home page
B. F. Burkey, X. Li, L. Bolognese, B. Balkan, M. Mone, M. Russell, T. E. Hughes, and P. R. Wang
Acute and Chronic Effects of the Incretin Enhancer Vildagliptin in Insulin-Resistant Rats
J. Pharmacol. Exp. Ther., November 1, 2005; 315(2): 688 - 695.
[Abstract] [Full Text] [PDF]

Home page
 J. Biol. Chem.Home page
C.-H. Chien, L.-H. Huang, C.-Y. Chou, Y.-S. Chen, Y.-S. Han, G.-G. Chang, P.-H. Liang, and X. Chen
One Site Mutation Disrupts Dimer Formation in Human DPP-IV Proteins
J. Biol. Chem., December 10, 2004; 279(50): 52338 - 52345.
[Abstract] [Full Text] [PDF]

Home page
 DiabetesHome page
C. F. Deacon
Therapeutic Strategies Based on Glucagon-Like Peptide 1
Diabetes, September 1, 2004; 53(9): 2181 - 2189.
[Abstract] [Full Text] [PDF]

Home page