continuous glucose monitoring for diabetes, obesity and metabolism research in rodents

Continuous Glucose Monitoring for Diabetes, Obesity and Metabolism Research in Rodents

Christian Schnell

Lab Head in-vivo Pharmacology, Oncology ResearchNovartis AG

Ralf Dechend, MDGroup Leader, ECRC, Charité Campus BuchMax Delbruck Center for Molecular Medicine, Berlin, Germany

2015 R&D 100 Finalists - PhysioTel HD-XG

InsideScientific is an online educational environment designed for life science researchers. Our goal is to aid in

the sharing and distribution of scientific information regarding innovative technologies, protocols, research tools

and laboratory services.

http://insidescientific.com/


Data Sciences International is seeking partners who are looking to change the way blood glucose monitoring is carried out in the research community.

Particularly, we wish to collaborate with researchers focused on the scientific and animal welfare benefits of the HD-XG implantable device.

Such research could include study designs to target:

• Increasing statistical power leading to a reduction in animal use

• Uncovering new and beneficial scientific evaluations that are currently not feasible

• Improving animal welfare and glucose measurements by decreasing animal stress

Learn More Here: www.crackit.org.uk

http://www.datasci.com/


http://www.crackit.org.uk/




http://www.crackit.org.uk/continuous-glucose-telemetry-enhance-research-and-improve-animal-welfare

Learn more about PhysioTel HD-HX…

Click to watch Product and Calibration Videos

http://www.datasci.com/glucose/resources/videos




Continuous Glucose Measurement in a Novel Rat Model of Type II Diabetes: Implication for pregnancy and fetal programming

Ralf Dechend, MDGroup Leader,

Experimental and Clinical Research Center, Charite Campus Buch

Max Delbruck Center for Molecular Medicine, Berlin, Germany

Copyright 2015 R. Dechend, Data Sciences Int’l & InsideScientific. All Rights Reserved.

A Novel Diabetes Model in Rats

• Tetracyclin induces knockdown of the insulin receptor

• Specific characteristics:

diabetes can be induced reversibly

induces typ II diabetes (insulin resistance)

• 2 models: preexistant diabetes and gestational diabetes

Click to access publication online

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2659743/

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2659743/

Dose Dependent Knock-Down of Insulin-Receptor

Characterization of the Diabetic Mother

** p≤0.001 ** p≤0.0001

A complete picture of insulin resistance…

Santos SH et. al., Oral administration of angiotensin-(1-7) ameliorates type 2 diabetes in rats. J Mol Med (Berl). 2014 Mar;92(3):255-65.

The Model: A First Intervention

http://www.ncbi.nlm.nih.gov/pubmed/24162089

Getahun et al. Am J Obstet Gynecol; 2008Bellamy et al. Lancet 2009

Gestational Diabetes: An Enormous Problem

http://www.ajog.org/article/S0002-9378(07)02128-X/abstract

http://www.ncbi.nlm.nih.gov/pubmed/19465232

A process whereby an insultapplied at a sensitive period of development results in long term changes in the structure or function of the organism.

Fetal Programming (Barker Hypothesis)

Gestational Diabetes: An Enormous Problem

Diabetes and Pregnancy...

The Mating Protocol

Non-diabetic pregnancy with…

• healthy fetus (group 1)

• Diabetic fetus (group 2)

Diabetic pregnancy with…

• healthy fetus (group 3)

• Diabetic fetus (group 4)

Healthy fetus…

• After non-diabetic pregnancy (group 1)

• After diabetic pregnancy (group 3)

Diabetic fetus…

• After non-diabetic pregnancy (group 2)

• After diabetic pregnancy (group 3)

Implantable Glucose Telemetry

• The glucose sensor is placed directly in free flowing blood

• provides an accurate signal for at least 28 days

• reduces blood draws, less stressed animals; more accurate data with reduced variability

• also provides continuous temperature and activity

Implantable Glucose Telemetry

TetO Rats Doxycyclin Treatment vs. ControlTetO Rats Doxycyclin treatment vs. Control

10.0 11.0 12.0 13.0 14.0 15.0 16.0 17.0 18.0 19.0 20.0 21.0 22.0 23.0 24.0

80.0

160.0

240.0

320.0

400.0

480.0

560.0

640.0

mg

/dL

19.08.2014 17:27:00

P72TETONr804RN309.Glucose.Glucose P59TETONr781RN300.Glucose.GlucoseDays

Printed: 22 01 2015 13:16:56

Example of Diabetic vs. Control Rat (Non-Pregnant)

Black = Control [P72TetONr804RN309.Glucose.Glucose]

Red = Dox. Treated [P59TetONr781RN300.Glucose.Glucose]

Example of Single Point Calibration of Diabetic Rat

0 25 50 75 100 125 1500

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lutz

ucker

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/dL

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t [min]

Blu

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2,05g/kg Glucose 2,7 g/kg Glucose

IP-GTTs in Normoglycemic Rats

Continuous Glucose Measurement (Diabetic Groups)

Continuous Glucose Measurement (Non-Diabetic Groups)

Day/Night Variability

Glucose in Normoglycemic Rats Pregnant vs. Non-Pregnant

• High variability

• Significant lower glucose levels in pregnancy

• Effect biggest at end of pregnancy

Glucose in Diabetic Rats Pregnant vs. Non-Pregnant

• Higher variability

• Tendency towards higher glucose levels in pregnancy

Within-Animal Variability For a Single Day (Non-Diabetic Groups)

Within-Animal Variability For a Single Day (Diabetic Groups)

Preliminary Summary of Results – a work in progress...

• Significant lower glucose levels in pregnancy in normoglycemic rats

• Tendency towards higher glucose levels in pregnancy in diabetic rats

Reduction in Sample Size By Using Continuous Monitoring

• Simulation based on 1000 simulation runs with 100 animals per run

• sample sizes necessary to detect a 10% mean change with 80% power is computed

• Minimum of 28 days gives the ability to see results in both acute and chronic studies.

• The ability to see a complete, accurate, blood glucose profile from freely moving, unstressed animals.

• Complete iPTT or oGTT can be performed

• Direct connection to data-management-systems

• Risk of operation

• Adhesion, fibrosis, encapsulation, risk of infection of the electrodes

• Quality of data depends on calibration, several calibration needed

• Price…

Summary

• Current blood glucose monitoring use blood samples taken from restrained animals experiencing stress. The number of samples may be limited, and collection requires manual labor: The end result is an incomplete profile of blood glucose management.

Assessment of Drug Related Blood Hyperglycemia Profile Using Real-Time Continuous Glucose Monitoring via Radio-telemetry in Rats

Christian Schnell

Lab Head in-vivo Pharmacology,

Oncology ResearchNovartis AG

Copyright 2015 C. Schnell, Data Sciences Int’l & InsideScientific. All Rights Reserved.

The traditional way....

• In rats, mostly in isoflurane anaesthetized animals using the sub-lingual blood collection approach and a glucometer

2 | Glucose radio-telemetry | C. Schnell

Monitoring of blood glucose levels in rats:

(Courtesy of DSI)

(0.7 mm Ø)


The new radio-telemetry approach using the HD-XG transmitter from DSI


(Courtesy of DSI)

Monitoring of blood glucose levels in BN rats:

• Ventral abdominal incision

• carefully separate the aorta from the vena cava just caudal to the point where the left renal vein crosses over the aorta and cranial to the iliac bifurcation

• placement of the cranial and caudal occlusion sutures (4-0 or 5-0)

The New Radio-Telemetry Approach: Abdominal Aorta Cannulation


(Courtesy of DSI)

← Abdominal Artery

Monitoring of blood glucose levels in BN rats:The New Radio-Telemetry Approach: Abdominal Aorta Cannulation

• Using the bent 25-gauge needle pierce the artery 1-2 mm cranial to the iliac bifurcation and insert the sensor upstream toward the heart

• Once the sensor is inserted into the vessel, withdraw the sensor introducer.


(Courtesy of DSI)


• Advance the sensor cranially so that the maximum amount most of sensor is within the vessel

• Do not advance the sensor past the cranial occlusion suture

• Dry the aorta at the sensor entry site with cotton tip applicators


(Courtesy of DSI)


• apply a very small amount of VetbondTM

tissue adhesive all the way around the entire sensor entry site

• Verify proper sensor placement by monitoring the live signal via a telemetry system

• Anchor the sensor in place with two small fiber patches


(Courtesy of DSI)

Monitoring of blood glucose levels in BN rats:The New Radio-Telemetry Approach: Intraperitoneal placement of the transmitter

• Close the abdominal wall using 4-0 or 5-0 non-absorbable suture with a simple interrupted pattern

• Incorporate the suture rib on the transmitter into the closure

• Close the skin incision using 4-0 or 5-0 absorbable suture


(Courtesy of DSI)

Monitoring of blood glucose levels in BN rats:

Frequency: 1 Hz Data sampling every minute

The New Radio-Telemetry Approach: Hardware…

Time (days)

-8 -7 -6 -5 -4 -3 -2 -1 0 1 2 3 4 5 6 7

-8

-7

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eig

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(g)

Implantation

Single housing

Values are mean +/- SEM (n=6; initial body weight: 185 ± 3 g)

Monitoring of blood glucose levels in BN rats:The New Radio-Telemetry Approach: Recovery post-implantation…

• Rats are highly social animals

• When ever possible, group housing conditions should be privileged

• Body weight change was recorded daily in radio-telemetered rats post-implantation


Time (days)

-8 -7 -6 -5 -4 -3 -2 -1 0 1 2 3 4 5 6 7

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(g)


Implantation

Single housing Group housing

Values are mean +/- SEM (n=6; initial body weight: 185 ± 3 g)

Monitoring of blood glucose levels in BN rats:The New Radio-Telemetry Approach: Recovery post-implantation…

• Body weight change was recorded daily in radio-telemetered rats post-implantation

• Body weight nearly normalized 8 days following implantation in single housed animals

• However, a clear tendency of enhanced body weight gain was observed when a pair mate was added into the cage


Time of the day (hours)

0 2 4 6 8 10 12 14 16 18 20 22 24

3.5

4.0

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Mo

tor

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ity

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unts

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ea

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M)

Blo

od

Glu

co

se l

ev

els

(mm

ol/L

, m

ea

n ±

SE

M)

= inactive = active

Monitoring of blood glucose levels in BN rats:The New Radio-Telemetry Approach: Diurnal rhythms (n=12)

• For the first time, circadian rhythms of glucose and motor activity could be measured in unstressed freely moving Brown-Norway rats

• Values were significantly lower (P< 0.005) during the day (inactive phase, 6 AM to 6 PM) than during the night (active phase, 6 PM to 6 AM)


0 2 4 6 8 10 12 14 16 18 20 22 24

36.0

36.2

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tor

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tiv

ity (

co

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/min

, m

ea

n ±

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M)

Bo

dy T

em

pe

ratu

re

(°C

, m

ea

n ±

SE

M)

= inactive = active



• Circadian rhythms of glucose and motor activity could also be concomitantly assessed in unstressed freely moving Brown-Norway rats

• Values were significantly lower (P< 0.005) during the day (inactive phase, 6 AM to 6 PM) than during the night (active phase, 6 PM to 6 AM)



0 2 4 6 8 10 12 14 16 18 20 22 24

3.5

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tor

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unts

/min

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ea

n ±

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M)

Blo

od

Glu

co

se l

ev

els

(mm

ol/L

, m

ea

n ±

SE

M)

= inactive = active

• Blood glucose baseline values during the inactive phase were around 5 ± 0.3 mmol/L

• Historical data collected sublingually in the Lab (following a short isoflurane anesthesia) were more in the range of 8 to 10 mmol/L

• How can we explain this difference?


Time (min)

-15 -10 -5 0 5 10 15 20 25 30 35 40 45 50

35.0

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(mm

ol/L

)


Isofluran →

Monitoring of blood glucose levels in BN rats:The New Radio-Telemetry Approach: Acute effect of anaesthesia

• One possible explanation could be the stress effect of isoflurane anaesthesia on blood glucose levels

• To address this question, we measured baseline blood glucose levels in a rat via radio-telemetry in his home cage

• The rat was then anaesthetized with isoflurane for 5 min

Time (min)

-15 -10 -5 0 5 10 15 20 25 30 35 40 45 50

35.0

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M)

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Glu

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(mm

ol/L

)


Isofluran →


• Following these 5 min of anaesthesia, the rat was returned to his cage and blood glucose / body temperature (BT) recorded continuously

• Blood glucose values very rapidly raised to levels of up to 8 mmol/L while BT decreased by 1.5 °C

• It took nearly 20 to 25 min until the blood glucose levels returned to initial values

Time (min)

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min

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(mm

ol/L

)


Isofluran →

Time (min)

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)

Isofluran →Isofluran →



Is o flu r a n e T e le m e tr y

0

5

1 0

1 5

2 0

Blo

od

glu

co

se

le

ve

ls (

mm

ol/

L)

me

an

S

D

*( 8 .6 )

(5 .0 )

(n=42) (n=12)

Monitoring of blood glucose levels in BN rats:The New Radio-Telemetry Approach: Base line assessment

• Comparison of baseline blood glucose values in rats measured sublingually following isoflurane anesthesia (left) or via radio-telemetry (right). *: p< 0.0001, statistical significance, two tailed t-Test.

• Telemetry reduces blood glucose basal levels and variability (3R’s)

Time (min)

-15 -10 -5 0 5 10 15 20 25 30 35 40 45 50 55 60 65

35.0

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(mm

ol/L

)


← Vehicle p.o. • Furthermore, we wanted to assess the stress effect of oral administration on blood glucose levels

• A very minor effect on blood glucose levels (+ 0.5 mmol/L) and BT could be evidenced

• Blood glucose levels and BT normalized after 20 min

Monitoring of blood glucose levels in BN rats:The New Radio-Telemetry Approach: Effect of vehicle p.o. (n=5)

Time (hours)

Day 1 Day 2 Day 3 Day 4 Day 5

Blo

od

Glu

co

se L

evels

( m

mol/L )

4.0

4.5

5.0

5.5

6.0

6.5

7.0

Active

Time (hours)


Bo

dy T

em

pera

ture

( d

egre

e C

els

ius )

36.5

37.0

37.5

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38.5

Active

Time (hours)


Lo

co

mo

tor

Acti

vit

y (

counts

/min

)

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Active


Monitoring of blood glucose levels in BN rats:The New Radio-Telemetry Approach: Base line assessment (n=9)

Blood Glucose Locomotor Activity Body Temperature

Conclusions:

• At day 1 of treatment, hyperglycemia could be managed by insulin production

• At steady state (Day 4-5), a clear hyperglycemia profile was observed

• Telemetry monitoring allows the assessment of degree and duration of pathway inhibition

Dosed at 10 a.m. before the inactive period (n=6)

Time (hours)

Day 0 Day 1 Day 2 Day 3Blo

od

Glu

co

se L

evels

(

mm

ol/L,

mean ±

SE

M)

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BYL719Vehicle BYL719 BYL719

Day 4 Day 5 Day 6 Day 7

BYL719 BYL719 Active


Monitoring of blood glucose levels in BN rats:The New Radio-Telemetry Approach: BYL719 50 mg/kg p.o.

Time post treatment (hours)

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ted

Pla

sm

a c

on

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trati

on

(µm

ol/L

, m

ea

n ±

SE

M)

BYL719 50 mg/kg p.o.

Blo

od

Glu

co

se lev

els

(mm

ol/L

, m

ea

n ±

SE

M)

• Values are mean +/- SEM at day 4 (n=6)

• BYL719 treatment in BN rats induced a transient glucose level increase suggestive of glucose metabolism impairment

• A clear PK/PD relationship could be demonstrated


Monitoring of blood glucose levels in BN rats:The New Radio-Telemetry Approach: BYL719 50 mg 10 a.m.

1

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16

Time post-implantation (days)

1 - 10 11 - 20 21 - 30 31 - 40 41 - 50 51 - 60 61 - 70 71 - 80 81 - 90


DSI success rate ≥ 28 days → 80%

Monitoring of blood glucose levels in BN rats:Radio-Telemetry : Success Rate ≥ 28 days → 92%

.

1 → Battery

2

3

4 → signal quality

5

6

7

8

9

10

11 → Battery

12

13 → signal quality

14

15

16

Time post-implantation (days)

1 - 10 11 - 20 21 - 30 31 - 40 41 - 50 51 - 60 61 - 70 71 - 80 81 - 90


DSI success rate ≥ 28 days → 80% : Hardware related failure

Monitoring of blood glucose levels in BN rats:Radio-Telemetry : Success Rate ≥ 60 days → 85%

.

• The HD-XG glucose telemetry implant offers a novel solution to obtain continuous, real-time, blood glucose measurements in laboratory animals in preclinical research

• We can now collect data for up to 84 days while reducing animal stress and measurement variability commonly associated with glucose test strip (3R’s)

• For the first time, a continuous 24h profile on blood glucose for NVP-BYL719 could be obtained. This profile is reproducible over time and a clear PK/PD relationship could be demonstrated

• It can be emphasized that the use of such a novel technology will enable advances in the understanding of the physiology of glucose metabolism regulation following PI3K inhibition possibly leading to improved therapies in the clinic


Monitoring of blood glucose levels in BN rats:Radio-Telemetry : Conclusions…

Data Sciences International is seeking partners who are looking to change the way blood glucose monitoring is carried out in the research community.

Particularly, we wish to collaborate with researchers focused on the scientific and animal welfare benefits of the HD-XG implantable device.

Such research could include study designs to target:

• Increasing statistical power leading to a reduction in animal use

• Uncovering new and beneficial scientific evaluations that are currently not feasible

• Improving animal welfare and glucose measurements by decreasing animal stress

Learn More Here: www.crackit.org.uk







http://www.crackit.org.uk/continuous-glucose-telemetry-enhance-research-and-improve-animal-welfare

Thank You!

Christian [email protected]

For additional information on Data Sciences International PhysioTel HD-XG implant for continuous glucose recordings and additional implantable telemetry solutions for physiological monitoring please visit:

http://www.datasci.com

Dr. Ralf [email protected]



