from stem cells to beta cells possible cure for diabetes mellitus
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From Stem Cells to Beta Cells: Possible Cure for Diabetes Mellitus. By Ryan Scavinski. Diabetes Mellitus. Type 1 Diabetes is caused by the autoimmune destruction of β -cells within the pancreas. No β -cells, no insulin Diabetics need to monitor blood glucose and control it with insulin.

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Presentation Transcript
diabetes mellitus
Diabetes Mellitus
  • Type 1 Diabetes is caused by the autoimmune destruction of β-cells within the pancreas.
  • No β-cells, no insulin
  • Diabetics need to monitor blood glucose and control it with insulin.
past treatments
Past Treatments
  • Transplantation of pancreatic islet cells
  • Problems
    • Patient requires more than 600 islets/kg body weight = two deceased organ donors
    • Immunological rejection
using stem cells
Using Stem Cells
  • Two approaches used to differentiate Embryonic Stem Cells into β-cells or Insulin Producing Cells (IPCs)
    • Embryoid Body Formation
    • Definitive Endoderm Formation
embryoid body formation
Embryoid Body Formation
  • An embryoid is the arrangement of stem cells destined to differentiate into the ectoderm, mesoderm and endoderm.
  • With multiple treatment of growth factors, Embryonic stem cells give rise to Nestin cells, which in turn differentiate into IPCs.
problems
Problems
  • Low efficiency for producing IPCs
  • Also showed development of tumors in the kidney and spleen in some transplanted mice.
definitive endoderm approach
Definitive Endoderm Approach
  • This approach bypasses the Embryoid formation and generates the endoderm, in which produce the cells needed for insulin production.
differentiation steps
Differentiation Steps
  • 1. ESCs were placed onto a culture dish with a chemically defined medium (CDM) containing 50 ng/mL Activin A for 4 days
slide10
With and without Activin A
  • PCR of expression of endoderm genes
    • Gapd, sox17, pdx1, hlxb9, hnf4a and insulin

gapd sox17 pdx1 hlxb9 hnf4α insulin

continued
continued
  • 2. Then the cells were transferred onto a CDM with 10-6 M Retinoic acid (RA) for another 4 days
slide12
Without Activin A and RA

With Activin A and RA

small clusters of differentiated ES

slide13
PCR
  • A+/RA+
  • A-/RA-
  • A+/RA-
  • A-/RA+

gapd sox17 pdx1 hlxb9 hnf4α insulin

continued1
continued
  • 3. Then the CDM was changed to modified islet maturation medium containing bFGF- a pancreatic cell maturation factor for 3 days
  • 4. Finally the differentiated cells were switched to a islet maturation medium containing nicotinamide and the bFGF for another 3 days.
slide15
In this final stage, many differentiated cells formed spherical clusters
  • Also expressed the pancreatic β cell markers such as pdx1, INSULIN, glucokinase and glut2 shown in PCR
differentiated embryonic stem cells
Differentiated Embryonic Stem Cells

gapd sox17 pdx1 hlxb9 hnf4α insulin glut2 Amy SST Sur1 GCG GCK

Maturation

Control

slide17
To test cells for insulin release, cells were incubated in buffer containing 2.5 mM glucose for 15 min
  • Then incubated with 27.5 mM glucose for another 15 min
  • Tested for insulin release with a Rat/Mouse insulin ELISA
slide18

2.5 mM

27.5 mM

Insulin secretion (ng/mg)

Suspension Adhesion

transplantation
Transplantation
  • Differentiated cells were transplanted under the renal capsule (kidney) of diabetic mice.
  • 30% showed normal blood glucose levels for 6 weeks
  • They removed the cell transplanted kidney-mice regained hyperglycemia
slide20

30

25

20

15

10

5

0

Blood glucose (mM)

0 7 14 21 28 35 42 49 56

Days after transplantation

results and discussion
Results and Discussion
  • The combination of Activin A and Retinoic Acid is an effective method to induce Embryonic Stem Cells to differentiate into insulin producing cells
  • Further research in needed to see if the difference between human and mouse will impair the function of transplanted ESC derived cells
slide23
Reference:
  • Soria, B., Skoudy, A., and Martin, F. 2001. From stem cells to beta cells: new strategies in cell therapy of diabetes mellitus. Diabetologia 44 407-415
  • Raikwar, S. and Zavazava, N. 2009. Insulin producing cells derived from embryonic stem cells: are we there yet?. Journal of Cellular Physiology, 218 256-263
  • Jiang, W., Shi, Y., Zhao, D., Chen, S., Youg, J., Zhang, J., Qing, T., Sun, X., Zhang, P., Ding, M., Li, D., and Deng, H. 2007. In vitro derivation of functional insulin producing cells from human embryonic stem cells. Cell Research. 17 333-344
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