Oxygenation in Encapsulated Islets

Jake YeungDecember 4, 2012Piret Lab Meeting

1

Native islets in pancreas are well vascularized

2(Suckale, 2011)

Encapsulated islets present new challenges

3(Vaithilingam & Tuch, 2011)

Outline

Oxygen supply and its effects on islet performance

Oxygen transfer modelling in encapsulated islets

Generating alginate beads for improved oxygen transfer

Conclusion and future work

4

Oxygen levels of transplanted rat islets

5

(Carlsson et al., 2001)

- Nondiabetic rat

- Diabetic rat

Oxygen levels and number of islets transplanted

6

(Carlsson et al., 2001)

Oxygen levels in peritoneal space

7

Method Animal pO2 (mmHg) Source Comments

Implanted tonometers

Rabbit 30 - 60(Klossner et al., 1974)

15 days in 10 rabbits

Polarographic oxygen electrode

Mouse 20 - 60(Bourdel et al., 2007)

1 hour in 6 mice

Galvanic electrode

Rabbit 15 - 55(Towell et al., 1976)

14 days in at least 7 rabbits

19F-NMR Mice 31 - 79(Goh et al., 2011)

16 days in 2 mice

Effect of hypoxia on insulin secretion

8

(Papas et al., 1996)

Outline

Oxygen supply and its effects on islet performance

Oxygen transfer modelling in encapsulated islets

Generating alginate beads for improved oxygen transfer

Conclusion and future work

9

Model geometry

10

(Greenemeier, 2008)

i Islet radius=75μm

Bead radius=250μm

Mass transport

Pressure gradient:

Four boundary conditions:a𝑡 𝑟 = 𝑅2, 𝑃2 = 𝑃𝑒𝑥𝑡

a𝑡 𝑟 = 𝑅1, 𝑃1 = 𝑃2

a𝑡 𝑟 = 𝑅1, 𝛼𝐷 1𝑑𝑃1

𝑑𝑟= 𝛼𝐷 2

𝑑𝑃2

𝑑𝑟

a𝑡 𝑟 = 𝑅0,𝑑𝑃1

𝑑𝑧= 0

11

(𝛼𝐷)𝑖

1

𝑟2𝑑

𝑑𝑟𝑟2𝑑𝑃𝑖𝑑𝑟

= 𝑉𝑖

Parameters for the model

Parameter Symbol Value Units

Oxygen consumption rate 𝑉𝑚𝑎𝑥 4 × 10−8 mol/cm3/s

Critical pO2 for cell death 𝑃𝑐 0.1 mmHg

Oxygen permeability in tissue 𝛼𝐷 1 1.24 × 10−14 mol/cm/mmHg/s

Oxygen permeability in 5%(v/v)

alginate𝛼𝐷 2 3.27 × 10−14 mol/cm/mmHg/s

12

(Johnson et al., 2009)

Oxygen and insulin secretion rate within beads

13

0 50 100 150 200 2500

20

40

60O

xyge

n p

arti

al p

ress

ure

(m

m H

g)

0 50 100 150 200 2500

0.2

0.4

0.6

0.8

1

Radial distance from centre (um)

Frac

tio

nal

insu

lin s

ecr

eti

on

rat

e

Alginate

Islets

Necrotic core

Pext=15mmHg

Pext=40mmHg

Pext=60mmHg

Pext=60mmHg

Pext=40mmHg

Pext=15mmHg

Islet diameter=150μmBead diameter=500μm

0 20 40 60 800

0.2

0.4

0.6

0.8

1

External partial pressure of oxygen (mm Hg)

Frac

tio

n o

f n

orm

al in

suli

n s

ecr

eti

on

rat

e

Fraction of normal insulin secretion rate

14

Islet diameter=150μmBead diameter=500μm

0 20 40 60 80 1000

20

40

60

80

External partial pressure of oxygen (mm Hg)

Ne

cro

tic

core

rad

ius

(um

)

Necrotic core size

15

Islet diameter=150μmBead diameter=500μm

Effect of bead size on islet performance

16

Islet diameter=150μmPext=40mmHg

200 300 400 500 600 700 800 900 10000

5

10

15

20

25

30

Bead diameter (um)

Ne

cro

tic

core

rad

ius

(um

)

200 300 400 500 600 700 800 900 10000

0.2

0.4

0.6

0.8

1

Frac

tio

n o

f n

orm

al in

suli

n s

ecr

eti

on

rat

e

Pext=40mmHgIslet diameter=150μm

Outline

Oxygen supply and its effects on islet performance

Oxygen transfer modelling in encapsulated islets

Generating alginate beads for improved oxygen transfer

Conclusion and future work

17

Microfluidic device: top view

18

Alginate

Mineral oil

Acidified oil Outlet

Courtesy of René Pedroza

Microfluidic device: side view

19

Generating alginate beads

20

Bead size distribution: t=0 to 20 minutes

21

0 3 2

72

149

247 2 0 0 0

0

5

10

15

20

25

30

0

40

80

120

160

0-1

00

10

0-2

00

20

0-3

00

30

0-4

00

40

0-5

00

50

0-6

00

60

0-7

00

70

0-8

00

80

0-9

00

90

0-1

00

0

>10

00

Nec

roti

c co

re r

adiu

s (u

m)

Bea

ds

Co

un

ted

Bead diameter (µm)

Bead size distribution: t=20 to 40 minutes

22

0

14 12

26

89

47

155 3 4 2

0

5

10

15

20

25

30

0

20

40

60

80

100

0-1

00

10

0-2

00

20

0-3

00

30

0-4

00

40

0-5

00

50

0-6

00

60

0-7

00

70

0-8

00

80

0-9

00

90

0-1

00

0

>10

00

Nec

roti

c co

re r

adiu

s (u

m)

Bea

ds

Co

un

ted

Bead diameter (µm)

Bead size distribution: t=40 to 60 minutes

23

0 4 10

165

67 69

32

7 3 0 10

5

10

15

20

25

30

0

40

80

120

160

200

0-1

00

10

0-2

00

20

0-3

00

30

0-4

00

40

0-5

00

50

0-6

00

60

0-7

00

70

0-8

00

80

0-9

00

90

0-1

00

0

>10

00

Nec

roti

c co

re r

adiu

s (u

m)

Bea

ds

Co

un

ted

Bead diameter (µm)

Viability of MIN6 cells versus time in syringe

24

0%

20%

40%

60%

80%

100%

0 20 40 60 80 100

Ce

ll vi

abili

ty (

%)

Time spent in syringe (minutes)

Bead size: microfluidic chip versus spinner flask

25

From microfluidic chip From spinner flask

0.00

0.05

0.10

0.15

0.20

0.25

0.30

0-1

00

10

0-2

00

20

0-3

00

30

0-4

00

40

0-5

00

50

0-6

00

60

0-7

00

70

0-8

00

80

0-9

00

90

0-1

00

0

>1

00

0

Vo

lum

etri

c Fr

acti

on

Bead diameter (µm)

0-100 100-200

200-300

300-400

400-500

500-600

600-700

700-800

800-900

900-1000

>1000

Bead diameter (µm)

Microfluidic chip versus spinner flask

+ Narrower distribution

+ Reduced shear stress

+ No entrapped bubbles or oil

– Coalescence of beads

– Losses inside tubing

– Transient effects on viability

26

Outline

Oxygen supply and its effects on islet performance

Oxygen transfer modelling in encapsulated islets

Generating alginate beads for improved oxygen transfer

Conclusion and future work

27

Conclusion

28

(Greenemeier, 2008)

i Islet radius=75μm

Bead radius=250μm

Literature review Modelling Microfluidics

Future work

29

Analyzing islets after transplantation

Process optimization

Courtesy of Blanche Lo

Acknowledgements

Piret lab

Ali Nazari

Blanche Lo

Chris Sherwood

René Pedroza

30

Cheung lab

Linfen Yu

Works cited

31

Bourdel, N., Matsuzaki, S., Bazin, J.-E., Pouly, J.-L., Mage, G., and Canis, M. (2007). Peritoneal tissue-oxygen tension during a carbon

dioxide pneumoperitoneum in a mouse laparoscopic model with controlled respiratory support. Hum. Reprod. 22, 1149–1155.

Suckale, J. (2011). Mouse pancreatic islet: Wikipedia.

Carlsson, P.-O., Palm, F., Andersson, A., and Liss, P. (2001). Markedly Decreased Oxygen Tension in Transplanted Rat Pancreatic Islets

Irrespective of the Implantation Site. Diabetes 50, 489–495.

Goh, F., Long, R., Simpson, N., and Sambanis, A. (2011). Dual perfluorocarbon method to noninvasively monitor dissolved oxygen

concentration in tissue engineered constructs in vitro and in vivo. Biotechnology Progress 27, 1115–1125.

Greenemeier, L. (2008). Pigs Could Be the Salvation of Diabetes Sufferers: Scientific American.

Klossner, J., Kivisaari, J., and Niinikoski, J. (1974). Oxygen and carbon dioxide tensions in the abdominal cavity and colonic wall of the

rabbit. The American Journal of Surgery 127, 711–715.

Liu, E.H., Rother, K.I., and Harlan, D.M. (2009). Islet Transplantation and the Challenges of Treating Type 1 Diabetes. Discovery

Medicine 5, 43–49.

Papas, K.K., Long Jr., R.C., Constantinidis, I., and Sambanis, A. (1996). Effects of oxygen on metabolic and secretory activities of βTC3

cells. Biochimica Et Biophysica Acta (BBA) - General Subjects 1291, 163–166.

Towell, M.E., Lysak, I., Layne, E.C., and Bessman, S.P. (1976). Tissue oxygen tension in rabbits measured with a galvanic electrode. J

Appl Physiol 41, 245–250.

Vaithilingam, V., and Tuch, B.E. (2011). Islet Transplantation and Encapsulation: An Update on Recent Developments. Rev Diabet Stud 8,

51–67.

Batch effects?

32

0

200

400

600

800

1000

1200

0 100 200 300 400

Bea

d d

iam

eter

(u

m)

Bead rank

t=0 to 20 min

t=20 to 40 min

t=40 to 60 min

2 4 6 8 100

10

20

30

40

50

60

70

Percentage of Alginate (%v/v)

Ne

cro

tic

core

siz

e (

um

)

2 4 6 8 100

0.2

0.4

0.6

0.8

1

Frac

tio

n o

f n

orm

al in

suli

n s

ecr

eti

on

rat

e

Effect of alginate concentration

33

Pext=40mmHgIslet diameter=150μmBead diameter=500 μm