Distribution of Pancreatic Polypeptide in the Head of the Human Pancreas. By Andrew Kringas

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1 Distribution of Pancreatic Polypeptide in the Head of the Human Pancreas By Andrew Kringas

2 Diabetes (Diabetes mellitus) A condition when the body doesn t properly respond to insulin Type 1 Type 2 Gestational Diabetes Affecting our society today

3 Diabetes Diabetes comes along with a lot of comorbilities (Watkins 2003) Serious cardiovascular side affects come with Type 2 Diabetes Neuropathy, Retinopathy, and Nephropathy

4 Fabiola Souza and Harris, C-dihydrotetrabenzine ( 11 C-DTBZ) as a radioligand 11 C-DTBZ is very efficient for in vivo visualization of the vesicular monoamine transporter, type 2 (VMAT2)

5 Saisho et al VMAT2 is expressed in pancreatic beta cells and is a target for measurement of beta cell mass in vivo Researchers questioned VMAT2 and how much is actually expressed within the pancreas Most beta cells expressed VMAT2 and expression wasn t changed by the presence of diabetes

6 Research Plan This research plans to further evaluate insulin, VMAT2 and pancreatic polypeptide content in the pancreas by obtaining human pancreatic tissue. Properly stain the tissue for insulin, VMAT2 and pancreatic polypeptide. This research hopes to better define the ratio between the pancreatic tissue and the beta cells in the head, middle and tail of the pancreas

7 Methods Methods branching off Butler et al Pancreatic tissue was taken from deceased donors within hours of expiration to decrease the chances of tissue breakdown Small cubes of tissue (approximately 2 cm x 2 cm) were cut from the head, body and tail of the pancreas and placed in formalin solution Immunohistochemistry

8 Methods Primary antibodies Insulin (2 hours) VMAT2 (Overnight) Pancreatic Polypeptide (Overnight) Anti-body staining: Anti-mouse (Insulin, 2 hours) Anti-rabbit (VMAT2, overnight) Anti-goat (Pancreatic Polypeptide, overnight)

9 Methods DAB Staining One drop of the Buffer; ph 7.5 Two drops of the DAB One drop of hydrogen peroxide One drop of nickel solution Hematoxylin and Eosin stain

10 Data Collection Image Pro Plus 6.3 A. B. C. (Insulin stain)

11 Insulin Staining 0.03 Fractional Insulin Staining Area Head ( n=3) Body (n=5) Tail (n=5) The average fractional area for insulin staining in the head, body and tail of the pancreas was 1.16, 1.75 and 1.77%, respectively. There was no statistically significant difference between the means

12 Pancreatic Polypeptide Staining Fractional PPY Staining Area Head ( n=3) Body (n=4) Tail (n=5) The average fractional area for PPY staining in the head, body and tail of the pancreas was 1.13, 0.47 and 0.47%, respectively. There was statistically significant (approaching) difference between the means of the head versus the tail and body

13 VMAT2 Staining Fractional VMAT2 Staining Area Head ( n=2) Body (n=2) Tail (n=2) The average fractional area for VMAT2 staining in the head, body and tail of the pancreas was 1.19, 1.57 and 1.63, respectively. There was no statistically significant difference between the means

14 Data Collection

15 Discussion It is believed that the great majority of VMAT2 cells are insulin-producing beta cells. We therefore were able to evaluate pancreatic tissue after autopsy from humans without diabetes mellitus. We showed that insulin-producing beta cells are abundant throughout the pancreas. There appears to be no significant difference in insulin staining between head, body and tail. Consistent with older literature, pancreatic polypeptide appears to be more abundant in the head of the pancreas

16 Limitations The limitations of this study are mostly related to evaluations of tissue staining. The tissue staining at times was less than adequate and therefore the antibodies did not show well VMAT2 Staining Immunofluorescence staining to evaluate the contribution of insulin and pancreatic polypeptide to VMAT2 distribution.

17 Bibliography 1. Cowie, C.C., et al., Full accounting of diabetes and pre-diabetes in the U.S. population in and Diabetes Care, (2): p Freeby, M., et al., VMAT2 quantitation by PET as a biomarker for beta-cell mass in health and disease. Diabetes Obes Metab, Suppl 4: p Watkins, P.J., Cardiovascular disease, hypertension, and lipids. BMJ, (7394): p Raffo, A., et al., Role of vesicular monoamine transporter type 2 in rodent insulin secretion and glucose metabolism revealed by its specific antagonist tetrabenazine. J Endocrinol, (1): p Souza, F., et al., Longitudinal noninvasive PET-based beta cell mass estimates in a spontaneous diabetes rat model. J Clin Invest, (6): p Anlauf, M., et al., Expression of the two isoforms of the vesicular monoamine transporter (VMAT1 and VMAT2) in the endocrine pancreas and pancreatic endocrine tumors. J Histochem Cytochem, (8): p Goland, R., et al., 11C-dihydrotetrabenazine PET of the pancreas in subjects with long-standing type 1 diabetes and in healthy controls. J Nucl Med, (3): p Gepts, W., Pathologic anatomy of the pancreas in juvenile diabetes mellitus. Diabetes, (10): p Saisho, Y., et al., Relationship between pancreatic vesicular monoamine transporter 2 (VMAT2) and insulin expression in human pancreas. J Mol Histol, (5): p Gersell, D.J., R.L. Gingerich, and M.H. Greider, Regional distribution and concentration of pancreatic polypeptide in the human and canine pancreas. Diabetes, (1): p Miyazaki, K. and A. Funakoshi, Distribution of pancreatic polypeptide-like immunoreactivity in rat tissues. Regul Pept, (1-2): p

18 Conclusion Findings confirm previous reports that there is a significant enrichment of PPY cells in the head of the pancreas relative to the body and tail VMAT2 staining paralleled that of insulin staining Future studies will establish the frequencies of cells that stain double positive for insulin and VMAT2 and the frequency of cells that stain positive for both PPY and VMAT2