Conditions: Type 1 Diabetes Hyperglycemia, Postprandial
Keywords: glucagon post-prandial glucose type 1 diabetes
Allocation: RANDOMIZED | Intervention model: CROSSOVER
Enrollment: 1
Healthy volunteers: no

Brief summary:
Glucagon regulation and response in persons with T1D at the basal state and in response to various stimuli remains unclear. Dr. Philip Cryer has previously reported that, in T1D young adults with a course of the disease of 16+9 years, the absence of endogenous insulin secretion results in increased glucagon secretion after a mixed meal, concluding that endogenous insulin reciprocally regulates the alpha-cell glucagon secretion and also suggesting that glucagon dysregulation may play an important role in post-prandial hyperglycemia in T1D. Interestingly, recent research on human islets have shown that insulin inhibits counter-regulatory glucagon secretion by a paracrine effect mediated by SGLT2-dependent stimulation of somatostatin release. An important gap in our knowledge is whether the timing of prandial insulin doses affects the glucagon response to a hyperglycemic stimulus in patients with T1D who have undetectable C-peptide.

Whether appropriately timed exogenous insulin can modify the glucagon response to glucose fluctuations has not been studied. As such, this pilot study aims to characterize the glucagon response to meal-time hyperglycemia and to compare the difference in glucagon secretion when mealtime bolus insulin is given before the meal versus after the meal with the objective of understanding factors that contribute to the peak post-prandial blood glucose and AUC of blood glucose after a mixed meal in this target population.

Detailed description:
Glucagon regulation and response in persons with T1D at the basal state and in response to various stimuli remains unclear. Dr. Philip Cryer has previously reported that, in T1D young adults with a course of the disease of 16+9 years, the absence of endogenous insulin secretion results in increased glucagon secretion after a mixed meal , concluding that endogenous insulin reciprocally regulates the alpha-cell glucagon secretion and also suggesting that glucagon dysregulation may play an important role in post-prandial hyperglycemia in T1D. Interestingly, recent research on human islets have shown that insulin inhibits counter-regulatory glucagon secretion by a paracrine effect mediated by SGLT2-dependent stimulation of somatostatin release. An important gap in our knowledge is whether the timing of prandial insulin doses affects the glucagon response to a hyperglycemic stimulus in patients with T1D who have undetectable C-peptide.

T1D course. In general, loss of 80% of beta cell function is needed to develop severe hyperglycemia. Endogenous insulin secretion is almost entirely lost within 3 to 5 years after the diagnosis . In children with new diagnosis of T1D, glucagon levels were highly associated with post prandial blood glucose, but not with HbA1c level. It has also been reported that postprandial hyperglucagonemia worsens significantly while C-peptide secretion declines during the first year of the disease. Also, in children with greater than five years of T1D, postprandial glucagon levels increased 160% from one to sixty months after diagnosis (8) and in those with disease duration of 6.9 + 4 years, glucagon levels were strongly correlated with post-prandial glucose and mildly correlated with HbA1C levels. A study in T1D adults performed in China showed that post-prandial glucagon levels were higher in T1D patients in the first year of the disease compared to those with longer course of the disease, and that hypoglycemic events were lower in newly diagnosed patients, suggesting that glucagon secretory function may become impaired with longer duration of the disease. These studies suggest that glucagon dysregulation could impact glucose control with exogenous insulin, and that this dysregulation may change with disease duration.

Post-prandial hyperglycemia adds to glucose variability, difficulty with insulin dosing and overall instability of glucose control. A recent analysis from the T1D Exchange study, that included 4768 participants younger than 26 years with a clinical diagnosis of T1D for at least 1 year showed that 21% of the participants reported administering insulin several minutes before, 44% immediately before, 10% during, and 24% after meal. Interestingly, participants who reported administering insulin during or after a meal were more likely to report missing ≥1 mealtime insulin dose per week compared with those who administered insulin before meals. In addition, a sub-analysis showed that participants who dose mealtime insulin in the postprandial period have poorer glycemic control and greater frequency of severe hypoglycemia. However, despite of these recent results, patients are often advised to give insulin after eating because caloric intake is uncertain. Understanding the mechanisms behind the higher post-prandial peak in glucose with delayed insulin administration may help clinicians guide appropriate timing of insulin administration for T1D patients with long course of the disease.

Glucagon Regulation:

Different models have been used to study glucagon regulation. Since both hyperglycemia and hypoglycemia stimulate alpha cells to produce glucagon, both the lack of intra-islet insulin and this U-shaped glucagon response contribute to glucose dysregulation in T1D . It has been reported that in T1D patients with a duration of the disease of 26.4+7.5 years, glucagon levels increased in response to the OGTT under euglycemic and hyperglycemic conditions, however the relative contribution of hyperglucagonemia to post-prandial glucose rise in the presence or absence of preprandial insulin has not been clearly elucidated. Whether appropriately timed exogenous insulin can modify the glucagon response to glucose fluctuations has not been studied. As such, this pilot study aims to characterize the glucagon response to meal-time hyperglycemia and to compare the difference in glucagon secretion when mealtime bolus insulin is given before the meal versus after the meal with the objective of understanding factors that contribute to the peak post-prandial blood glucose and AUC of blood glucose after a mixed meal in this target population.

Hypothesis

1. Patients with higher glucagon responses to a mixed meal stimulus will have higher peak post-prandial glucoses and greater AUC of post-prandial glucose.
2. Bolus dose insulin given pre-meal (20 min before meal) will result in lower peak post-prandial glucose, lower AUC and lower post-prandial glucagon levels
3. Bolus dose insulin given post-meal (20 min after meal) will result in higher peak post-prandial glucoses, higher AUC of post-meal glucose and higher glucagon levels
4. There will be a correlation between peak post-prandial glucagon response and post-meal glucose AUC, 0 to 180 minutes.

Research Plans The study will be conducted in the (Intensive Research Unit, part of the CTSA) Diabetes Center at Barnes Jewish Hospital/Washington University in St Louis.

Study Visits

* Screening will be done over the phone and with review of medical records if the required labs have been done in the past 6 months
* Visits A and B will be performed in random order
* Demographics, review of diabetes history, medications and allergies will be recorded at the first study visit. Insulin doses and effectiveness of current carbohydrate ratio will be evaluated and recorded. Mealtime insulin doses will be established at the first study visit.
* Mealtime insulin will be given via injection by a study nurse to standardize the administration procedure.
* Vital signs, height and weight will be done at each visit
* C-peptide will be checked with the fasting labs at the first study visit
* Baseline glucose should be 80 to 140 mg/dl. In case baseline glucose is not in the target range, the study visit will be rescheduled.
* Whole blood will be collected in vacutainers via an indwelling peripheral venous catheter with nursing supervision.
* Samples will be deidentified before sending to the laboratory.
* No genetic testing will be performed.

Visit A

* Glucose levels done fasting, 30, 60, 120, and 180 minutes
* Glucagon levels done fasting, 30, 60, 120, and 180 minutes
* Usual insulin dose will be administered 20 minutes prior to a mixed meal challenge with Ensure Plus\*

Visit B

* Glucose done fasting, 30, 60, 120, and 180 minutes
* Glucagon levels done fasting, 30, 60, 120, and 180 minutes
* Usual insulin dose will be administered 20 minutes after the mixed meal challenge with

Ensure Plus\*

\*Ensure Plus (Abbott Nutrition) at 6cc/kg (max 360 cc), content per 100 ml: carbohydrate 21.5 grams, protein 5.5 grams, with nursing supervision.

Laboratory Measures:

Core Laboratory for Clinical Studies at Washington University (https://research.wustl.edu/core-facilities/core-laboratory-clinical-studies, CLCS) will measure glucose on the Roche cobas c510 platform using Roche's coupled enzymatic assay (hexokinase, glucose-6-phosphoate dehydrogenase). The rate of NADPH formation is directly proportional to glucose concentration and measured photometrically. The interday imprecision is typically \<2.0% CV. CLCS will also measure C-peptide on the Roche cobas e601platform using Roche's sandwich electrochemiluminescence immunoassay. In this assay a biotinylated monoclonal C-peptide-specific antibody and a monoclonal C-peptide-specific antibody labeled with a ruthenium complex bind to C-peptide to form the sandwich. Then, using streptavidin-coated microparticles, the complex is drawn into the measuring cell and remains while unbound material is washed away. Application of a voltage to the electrode induces chemiluminescent emission that is measured by the instrument. Between run precision typically ranges from 1.9 - 2.7% CV.

Glucagon is measured using a radioimmunoassay kit from Millipore. This is a competitive immunoassay which uses I-125 labeled glucagon and a glucagon-specific antibody. Its stated inter-assay precision is 12% CV. Typical within-run precision is 2.0 - 3.0 % CV in our hands.

Methods of Data Analysis Data analysis will be performed using STATA 15.1 for Windows (StataCorp LP, College Station, Texas, United States). The sample size calculation assumes that the peak postprandial glucose will increase by 80 +/-20 mg/dl in the pre-meal insulin group and 140 +/-20 mg/dl in the post-meal group. So, the expected mean peak of postprandial glucose will be up to 220 mg/dL for Visit A and mean peak postprandial glucose will be up to 280 mg/dL for Visit B. The sample size will have 80% power to detect a difference between groups at p\<0.05. We have assumed correlation between visits A and B of 0.5. The sample size was corrected by an anticipated loss of follow up of 10% of the participants. The calculated sample size is 10 participants with full data sets available for 8 participants. The study randomization is designed as follows: 5 participants will first attend Visit A, and then Visit B. The 5 remaining participants will first attend Visit B and then Visit A. AUC of post-prandial glucose and glucagon and maximum postprandial peak of glucose and glucagon will be analyzed by using paired T student. Relationships between variables were assessed by a Spearman's correlation test.

Arms:
arm 1: VISIT 1: Pre-prandial insulin: will give short acting insulin (Humalog, Lispro, etc) with the same regimen patient was using at home 20 min prior the meal.

Then VISIT 2: Post-prandial insulin: will give short acting insulin (Humalog, Lispro, etc) with the same regimen patient was using at home 20 min post the meal. arm 2: VISIT 1: Post-prandial insulin: will give short acting insulin (Humalog, Lispro, etc) with the same regimen patient was using at home 20 min post the meal.

Then VISIT 2: Pre-prandial insulin: will give short acting insulin (Humalog, Lispro, etc) with the same regimen patient was using at home 20 min prior the meal.

Interventions: intervention 1: Insulin
Intervention descriptions: intervention 1: to give pre-prandial and post-prandial insulin (will use patient insulin dose patient use at home) to patients with T1D and evaluate the response of post-prandial glucagon and post-prandial hyperglycemia.

Protocol document:
Prot_SAP_000.pdf: 
  
 
 
 
 
 
 
Study Protocol 
Title: Glucagon Response to Prandial Insulin 
Administration in Persons with Type 1 Diabetes 
NCT04079881 
Protocol Date: 25 June 2020 
 
 


Glucagon T1DM (Version 2.1, 25JUN2020)  
 
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Summary of Changes 
Version 2.0 to 2.1 
Removing Dr. Zelada Castro as Primary Investigator and adding 
Dr. Janet McGill  
Version 1.0 to 2.0 
Adding an additional blood sample for future research 
 
Changes to blood draw volumes 
 
Clarification of location data & sample storage 
 
Clarification of location of lab for processing samples 


Glucagon T1DM (Version 2.1, 25JUN2020)  
 
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Glucagon Response to Prandial Insulin Administration in Persons 
with Type 1 Diabetes 
 
 
Principle Investigator:  
Janet McGill, MD 
 
Investigator:  
 
Henry Zelada Castro, MD  
 
Supervisors:   
 
Janet McGill, MD, MA 
Jing Hughes, MD, PhD 
Philip Cryer MD 
 
Collaborators:       
 
Samantha Adamson MD, PhD 
 
 
OVERVIEW 
Glucagon regulation and response in persons with T1D at the basal state and in response to 
various stimuli remains unclear. Dr. Philip Cryer has previously reported that, in T1D young 
adults with a course of the disease of 16+9 years, the absence of endogenous insulin secretion 
results in increased glucagon secretion after a mixed meal, concluding that endogenous insulin 
reciprocally regulates the alpha-cell glucagon secretion (1,2) and also suggesting that glucagon 
dysregulation may play an important role in post-prandial hyperglycemia in T1D. Interestingly, 
recent research on human islets have shown that insulin inhibits counter-regulatory glucagon 
secretion by a paracrine effect mediated by SGLT2-dependent stimulation of somatostatin 
release (3).  
 
T1D course: (4) 
In general, loss of 80% of beta cell function is needed to develop severe hyperglycemia.  
Endogenous insulin secretion is almost entirely lost within 3 to 5 years after the diagnosis (5). In 
children with new diagnosis of T1D, glucagon levels were highly associated with post prandial 
blood glucose, but not with HbA1c level (6). It has also been reported that postprandial 
hyperglucagonemia worsens significantly while C-peptide secretion declines during the first year 
of the disease (7). Also, in children with greater than five years of T1D, postprandial glucagon 
levels increased 160% from one to sixty months after diagnosis (8) and in those with disease 
duration of 6.9 + 4 years, glucagon levels were strongly correlated with post-prandial glucose 
and mildly correlated with HbA1C levels (9). A study in T1D adults performed in China showed 
that post-prandial glucagon levels were higher in T1D patients in the first year of the disease 
compared to those with longer course of the disease, and that hypoglycemic events were lower 
in newly diagnosed patients (10), suggesting that glucagon secretory function may become 
impaired with longer duration of the disease. These studies suggest that glucagon dysregulation 
could impact glucose control with exogenous insulin, and that this dysregulation may change 
with disease duration.   
 
Insulin Timing and Hyperglycemia in T1D: 
Post-prandial hyperglycemia adds to glucose variability, difficulty with insulin dosing and overall 
instability of glucose control. A recent analysis from the T1D Exchange study, that included 
4768 participants younger than 26 years with a clinical diagnosis of T1D for at least 1 year 
showed that 21% of the participants reported administering insulin several minutes before, 44% 
immediately before, 10% during, and 24% after meal. Interestingly, participants who reported 
administering insulin during or after a meal were more likely to report missing ≥1 mealtime 
insulin dose per week compared with those who administered insulin before meals (11). In 
addition, a sub-analysis showed that participants who dose mealtime insulin in the postprandial 
period have poorer glycemic control and greater frequency of severe hypoglycemia (12). 
However, despite these recent results, patients are often advised to give insulin after eating 


Glucagon T1DM (Version 2.1, 25JUN2020)  
 
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because caloric intake is uncertain.  
 
Glucagon Regulation:   
Different models have been used to study glucagon regulation 
(13,14,15,16). Since both 
hyperglycemia and hypoglycemia stimulate alpha cells to produce glucagon, both the lack of 
intra-islet insulin and this U-shaped glucagon response contribute to glucose dysregulation in 
T1D (17,18). It has been reported that in T1D patients with a duration of the disease of 26.4+7.5 
years, glucagon levels increased in response to the OGTT under euglycemic and hyperglycemic 
conditions (19), however the relative contribution of hyperglucagonemia to post-prandial glucose 
rise in the presence or absence of pre-prandial insulin has not been clearly elucidated. Whether 
appropriately timed exogenous insulin can modify the glucagon response to glucose fluctuations 
has not been studied.  
 
Research Objective: 
 
To characterize the glucagon response to meal-time hyperglycemia and to compare the 
difference in glucagon secretion when mealtime bolus insulin is given before the meal versus 
after the meal with the objective of understanding factors that contribute to the peak post-
prandial blood glucose and AUC of blood glucose after a mixed meal in this target population. 
 
 Research Hypothesis: 
 
1. Patients with higher glucagon responses to a mixed meal stimulus will have higher peak 
post-prandial glucoses and greater AUC of post-prandial glucose. 
2. Bolus dose insulin given pre-meal (20 min before meal) will result in lower peak post-
prandial glucose, lower AUC and lower post-prandial glucagon levels 
3. Bolus dose insulin given post-meal (20 min after meal) will result in higher peak post-
prandial glucoses, higher AUC of post-meal glucose and higher glucagon levels 
4. There will be a correlation between peak post-prandial glucagon response and post-
meal glucose AUC, 0 to 180 minutes 
 
Potential Contribution: 
  
Glucagon regulation and response in persons with T1D at the basal state and in response to 
various stimuli remains unclear. An important gap in our knowledge is whether the timing of 
prandial insulin doses affects the glucagon response to a hyperglycemic stimulus in patients 
with T1D who have undetectable C-peptide. Understanding the mechanisms behind the higher 
post-prandial peak in glucose with delayed insulin administration may help clinicians guide 
appropriate timing of insulin administration for T1D patients with long course of the disease 
 
 
METHODS 
 
Recruitment:   
 
Subjects will be recruited from the investigators’ private patient population.  Screening will be 
done over the phone and with review of medical records. 
 
 
 
 


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Inclusion criteria: 
 Persons of all races, ethnicity and genders will be included, Age >18 
 Type 1 Diabetes for >5 years  
 HbA1c <9.5% 
 Using either MDI or insulin pumps  
 Patients using CGM may continue the use during the study, however glucoses will be 
measured by laboratory methods. 
 Hemoglobin:  Female 10.8 -17.05 g/dL and Males 11.7-19.25 g/dl 
 Hematocrit:    Female 32.8-48.7% and  Males 37-55.3% 
 
Exclusion criteria: 
 Type 2 diabetes, monogenic diabetes, pancreatic diseases 
  eGFR <60 ml/min/1.73m2. 
 Use of daily steroids, any route, for any purpose  
 Pregnancy, prisoners, other vulnerable populations or persons unable to understand the 
protocol and provide written informed consent  
 
Data Collection: 
See Appendix A 
 
LOCATION OF STUDY VISITS 
The study will be conducted in the Washington University-Diabetes Center, Center for 
Advanced Medicine 13B and/or the Intensive Research Unit, Center for Outpatient Health 5th 
floor. 
 
PROCEDURES  
 
Study Visits (Visits will take approximately 4 hours) 
 Informed Consent may be performed on a visit prior to Visits A/B – subjects will be 
provided informed consent 
 Visits A and B will be performed in random order, these visits will take place on 2 
separate days within 1 week of the first visit 
 Demographics, Obtain medical and medication history, including review of diabetes 
history, medications and allergies will be recorded at the first study visit. Insulin doses 
and effectiveness of current carbohydrate ratio will be evaluated and recorded. Mealtime 
insulin doses will be established at the first study visit. 
 Baseline glucose should be 80 to 180 mg/dl. In case baseline glucose is not in the target 
range, the investigator will determine if the subject should return and the study visit 
rescheduled. 
 Urine pregnancy test for women of childbearing potential 
 Brief physical exam performed at first study visit 
 Vital signs, height and weight performed at each visit 
 Meal tolerance tests with timed samples collected as noted below performed at Visit A & 
B.  Administration of liquid meal and Mealtime insulin to be given via injection by a study 
nurse to standardize the administration procedure. 
 C-peptide may be checked with the fasting labs at the first study visit 
 A sample for future research will be obtained, about 5ml (1tsp.) 
 Whole blood will be collected in vacutainers via an indwelling peripheral venous catheter 
with nursing supervision from the diabetes center at Washington University. Maximum 
blood collection will be about 20ml (4 tsp). 
 


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Visit A 
• 
Glucose levels done fasting, 30, 60, 120, and 180 minutes 
• 
Glucagon levels done fasting, 30, 60, 120, and 180 minutes 
• 
Usual insulin dose will be administered 20 minutes prior to a mixed meal 
challenge with Ensure Plus* 
Visit B 
• 
Glucose done fasting, 30, 60, 120, and 180 minutes 
• 
Glucagon levels done fasting, 30, 60, 120, and 180 minutes 
• 
Usual insulin dose will be administered 20 minutes after the mixed meal 
challenge with Ensure Plus* 
 
*Ensure Plus (Abbott Nutrition) at 6cc/kg (max 360 cc), content per 100 ml: carbohydrate 21.5 
grams, protein 5.5 grams, with nursing supervision. 
 
PROTECTION OF CONFIDENTIALITY  
Data Security  
Study data may be stored in REDCAP or on WUSM protected servers as well as collective data 
which will be stored in a locked cabinet in a locked office.  Study staff will be required to have 
their own logon/password to access electronic data and access to paper charts will be limited to 
authorized study staff. 
 
Blood Samples may be processed and stored in the Diabetes Center or in the Center for 
Outpatient Health clinical research unit (CRU) in a locked room with access restricted to study 
staff or delivered directly to the Washington University Core lab and the investigators lab for 
handling and analysis.  Blood samples will be coded before sending to the laboratory.  The 
samples may include a subject identification number and date of birth.  
Plasma and serum specimens will be stored in a Washington University School of Medicine 
research Laboratory for future studies of other metabolic parameters relating to pancreatic 
function and/or insulin signaling; 5 ml of plasma and serum will be collected at one of the visits. 
 
ASSESSMENT OF RISKS AND BENEFITS 
Potential risks include: 
- 
Mild pain from IV catheter insertion 
- 
Hypoglycemia after insulin administration. Patient will be monitored by a physician and a 
nurse during the whole visit. 
 
Methods 
Alpha cell function will be assessed by measuring fasting and post-prandial glucagon and 
calculation Area under the Curve ratio (AUC-Glucagon 0-180min/AUC-Glucose 0-180min). 
Some of the participants characteristics will include in the study are: Demographics, BMI, history 
of DKA, DKA at onset of diagnosis, diabetes comorbidities (Retinopathy, cardiovascular, 
nephropathy, neuropathy), concomitant autoimmune disease, type and dose of insulin, 
 
Methods of Data Analysis  
Data analysis will be performed using STATA 15.1 for Windows (StataCorp LP, College Station, 
Texas, United States). The sample size calculation assumes that the peak postprandial glucose 
will increase by 80 +/-20 mg/dl in the pre-meal insulin group and 140 +/-20 mg/dl in the post-
meal group. So, the expected mean peak of postprandial glucose will be up to 220 mg/dL for 
Visit A and mean peak postprandial glucose will be up to 280 mg/dL for Visit B. The sample size 
will have 80% power to detect a difference between groups at p<0.05. We have assumed 
correlation between visits A and B of 0.5. The sample size was corrected by an anticipated loss 
of follow up of 10% of the participants. The calculated sample size is 10 participants with full 


Glucagon T1DM (Version 2.1, 25JUN2020)  
 
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data sets available for 8 participants. The study randomization is designed as follows: 5 
participants will first attend Visit A, and then Visit B. The 5 remaining participants will first attend 
Visit B and then Visit A. AUC of post-prandial glucose and glucagon and maximum postprandial 
peak of glucose and glucagon will be analyzed by using paired T student. Relationships 
between variables were assessed by a Spearman’s correlation test.   


Glucagon T1DM (Version 2.1, 25JUN2020)  
 
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REFERENCES 
1.  
Cryer PE. Minireview: Glucagon in the pathogenesis of hypoglycemia and hyperglycemia 
in diabetes. Endocrinology. 2012;153(3):1039-1048. doi:10.1210/en.2011-1499 
2.  
Cooperberg BA, Cryer PE. Insulin reciprocally regulates glucagon secretion in humans. 
Diabetes. 2010;59(11):2936-2940. doi:10.2337/db10-0728 
3.  
Vergari E, Knudsen JG, Ramracheya R, et al. Insulin inhibits glucagon release by SGLT2-
induced stimulation of somatostatin secretion. Nat Commun. 2019;10(1):139. 
doi:10.1038/s41467-018-08193-8 
4.  
Yosten GLC. Alpha cell dysfunction in type 1 diabetes. Peptides. 2018;100:54-60. 
doi:10.1016/j.peptides.2017.12.001 
5.  
Schiffrin A, Suissa S, Weitzner G, Poussier P, Lalla D. Factors predicting course of beta-
cell function in IDDM. Diabetes Care. 1992;15(8):997-1001. doi:10.2337/diacare.15.8.997 
6.  
Porksen S, Nielsen LB, Kaas A, et al. Meal-stimulated glucagon release is associated with 
postprandial blood glucose level and does not interfere with glycemic control in children 
and adolescents with new-onset type 1 diabetes. J Clin Endocrinol Metab. 
2007;92(8):2910-2916. doi:10.1210/jc.2007-0244 
7.  
Brown RJ, Sinaii N, Rother KI. Too much glucagon, too little insulin: time course of 
pancreatic islet dysfunction in new-onset type 1 diabetes. Diabetes Care. 2008;31(7):1403-
1404. doi:10.2337/dc08-0575 
8.  
Fredheim S, Andersen M-LM, Porksen S, et al. The influence of glucagon on postprandial 
hyperglycaemia in children 5 years after onset of type 1 diabetes. Diabetologia. 
2015;58(4):828-834. doi:10.1007/s00125-014-3486-3 
9.  
Urakami T, Nagano N, Suzuki J, Yoshida A, Takahashi S, Mugishima H. Influence of 
plasma glucagon levels on glycemic control in children with type 1 diabetes. Pediatr Int Off 
J Jpn Pediatr Soc. 2011;53(1):46-49. doi:10.1111/j.1442-200X.2010.03184.x 
10.  Li K, Song W-J, Wu X, et al. Associations of serum glucagon levels with glycemic 
variability in type 1 diabetes with different disease durations. Endocrine. 2018;61(3):473-
481. doi:10.1007/s12020-018-1641-1 
11.  Datye KA, Boyle CT, Simmons J, et al. Timing of Meal Insulin and Its Relation to 
Adherence to Therapy in Type 1 Diabetes. J Diabetes Sci Technol. 2018;12(2):349-355. 
doi:10.1177/1932296817728525 
12.  Peters A, Van Name MA, Thorsted BL, Piltoft JS, Tamborlane WV. POSTPRANDIAL 
DOSING OF BOLUS INSULIN IN PATIENTS WITH TYPE 1 DIABETES: A. Endocr Pract 
Off J Am Coll Endocrinol Am Assoc Clin Endocrinol. 2017;23(10):1201-1209. 
doi:10.4158/EP171813.OR 
13.  Gopel SO, Kanno T, Barg S, Weng XG, Gromada J, Rorsman P. Regulation of glucagon 
release in mouse -cells by KATP channels and inactivation  of TTX-sensitive Na+ 
channels. J Physiol. 2000;528(Pt 3):509-520. doi:10.1111/j.1469-7793.2000.00509.x 
14.  MacDonald PE, De Marinis YZ, Ramracheya R, et al. A K ATP channel-dependent 
pathway within alpha cells regulates glucagon release from both rodent and human islets 
of Langerhans. PLoS Biol. 2007;5(6):e143. doi:10.1371/journal.pbio.0050143 


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15.  Vieira E, Salehi A, Gylfe E. Glucose inhibits glucagon secretion by a direct effect on mouse 
pancreatic alpha  cells. Diabetologia. 2007;50(2):370-379. doi:10.1007/s00125-006-0511-1 
16.  Liu Y-J, Vieira E, Gylfe E. A store-operated mechanism determines the activity of the 
electrically excitable  glucagon-secreting pancreatic alpha-cell. Cell Calcium. 
2004;35(4):357-365. doi:10.1016/j.ceca.2003.10.002 
17.  Gylfe E, Gilon P. Glucose regulation of glucagon secretion. Diabetes Res Clin Pract. 
2014;103(1):1-10. doi:10.1016/j.diabres.2013.11.019 
18.  Salehi A, Vieira E, Gylfe E. Paradoxical stimulation of glucagon secretion by high glucose 
concentrations. Diabetes. 2006;55(8):2318-2323. doi:10.2337/db06-0080 
19.  Kramer CK, Borgono CA, Van Nostrand P, Retnakaran R, Zinman B. Glucagon response 
to oral glucose challenge in type 1 diabetes: lack of impact of euglycemia. Diabetes Care. 
2014;37(4):1076-1082. doi:10.2337/dc13-2339 
 


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Data Collection – Appendix A 
Date: 
 
Patient Name: 
DOB: 
MRN: 
Evaluator: 
 
Visit A:  
BP: 
HR: 
RR: 
T:  
Weight: 
BMI: 
 
1. Demographic information: 
a. Age 
b. Sex: 
c. Married: 
d. Time of the disease (T1D) 
e. History of DKA  
f. DKA at onset of diagnosis 
g. Diabetes comorbidities (Retinopathy, cardiovascular, nephropathy, neuropathy) 
h. Concomitant autoimmune disease 
i. History of hypoglycemia, 
j. Hypoglycemia unawareness   
2. T1D: 
 Diabetes medications: 
a. Oral: 
b. Non-injectable insulin 
c. Insulin 
 ICR 
 Statins: 
3. Allergies: 


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4. Meal time prandial insulin dose to be used (in second visit) 
5. Physical Examination: 
a. General: 
b. HEENT: 
c. Cardio: 
d. Lungs: 
e. GI: 
f. Extremities: 
g. Skin: 
h. Neuro: 
6. Labs: 
 Fasting C-peptide 
 CBC (if required) 
 GFR (if required) 
 A1C (if required) 
7. Glucose Target prior intervention: (Baseline glucose should be 80 to 140 mg/dl. In case 
baseline glucose is not in the target range, the study visit will be rescheduled): Yes, No 
 
8. Intervention:  
Usual insulin dose will be administered 20 minutes prior to a mixed meal challenge with Ensure Plus 
 
• 
Glucose levels: 
• 
Fasting: 
• 
30 min: 
• 
60 min: 
• 
120 mins: 
• 
180 min: 
 
• 
Glucagon levels: 
• 
Fasting: 
• 
30 min: 
• 
60 min: 
• 
120 mins: 
• 
180 min: 
9. Was the intervention hold? Yes, No 
 
Reason: 
 
 
 
 
 
 


Glucagon T1DM (Version 2.1, 25JUN2020)  
 
Page 11 of 12 
 
Visit B:  
BP: 
HR: 
RR: 
T:  
Weight: 
BMI: 
 
1. Glucose Target prior intervention: (Baseline glucose should be 80 to 140 mg/dl. In case 
baseline glucose is not in the target range, the study visit will be rescheduled): Yes, No 
2. Physical Examination: 
a. General: 
b. HEENT: 
c. Cardio: 
d. Lungs: 
e. GI: 
f. Extremities: 
g. Skin: 
h. Neuro: 
 
 
3. Intervention 
 
Usual insulin dose will be administered 20 minutes post to a mixed meal challenge with Ensure Plus 
 
• 
Glucose levels: 
• 
Fasting: 
• 
30 min: 
• 
60 min: 
• 
120 mins: 
• 
180 min: 
 
• 
Glucagon levels: 
• 
Fasting: 
• 
30 min: 
• 
60 min: 
• 
120 mins: 
• 
180 min: 
 
4. Was the intervention hold? Yes, No  
Reason: