Cellular Mechanism of Exocrine Pancreatic Insufficiency in Diabetes Mellitus

Patel, Rekha (2005) Cellular Mechanism of Exocrine Pancreatic Insufficiency in Diabetes Mellitus. Doctoral thesis, University of Central Lancashire.

[thumbnail of Thesis document]
PDF (Thesis document) - Accepted Version
Available under License Creative Commons Attribution Non-commercial Share Alike.



Diabetes mellitus (DM) is associated with the compromised digestion of carbohydrates.
This complication is described as exocrine pancreatic insufficiency. Whilst this causes
malnutrition in patients and contributes to diabetic morbidity, the physiology and molecular
biology leading to this state is not well defined. This disease-induced inability to digest
foodstuffs could have many levels of regulation. Obvious candidates are ligand proteins
involved in stimulating secretion, receptor defects, intracellular ion levels and post-receptor
signal transduction encompassing transcription and translation. To address these points, the
current studies aimed to characterise the effects of experimental type I DM upon both
physiological and molecular events.

The first study investigated the effects of cholecystokinin-octapeptide (CCK-8) and
exogenous insulin on exocrine pancreatic amylase secretion in streptozotocin (STZ)-
induced diabetic rats compared to healthy age-matched controls in vivo and in vitro. Seven
- eight weeks after the induction of diabetes, animals were either anaesthetised for the
study of in vivo exocrine secretion or humanely killed and pancreatic acinar cells isolated
for the measurement of intracellular free calcium and magnesium concentrations ([Ca 2+ ]1
and [Mg2+ ]1), total protein, and amylase output employing the Phadebas method. For rats in
both in vivo and in vitro studies, fasting blood glucose in control and diabetic rats was 73.3
± 3.4 mg dl-1 (n = 44) and 380.0 ± 25.9 mg dl-1 (n = 27), respectively. Basal pancreatic
juice flow rate in STZ-diabetic rats was significantly increased (P<0.001) whereas protein
and amylase outputs were significantly decreased (P<0.001) compared to control rats.
CCK-8 infusion (150 pmol kg -1 h-1 for 100 mm) resulted in marked elevations in flow rate
as well as in protein and amylase secretion in control animals (P<0.05 compared with the
corresponding basals). In contrast, in diabetic rats, CCK-8 evoked a small increase in flow
rate, which was not significant when compared to basal. In these animals, CCK-8
stimulated the secretion of amylase and protein output, but the secretory rates were
dramatically lower compared with those in control rats. Administration of insulin (1 U,
I.P.) in healthy rats significantly increased pancreatic flow rate, amylase secretion, protein
output and blood glucose levels in vivo compared to basal (P<0.05). Infusion of CCK-8
together with insulin (1 U) in control rats markedly potentiated pancreatic juice flow and
amylase secretion. Pretreatment with atropine (0.2 mg kg-1, I.P.) abolished the effects of
insulin on secretory parameters despite a similar reduction in glycaemia. In diabetic rats,
insulin (4 U, I.P.) did not modify exocrine pancreatic secretion either alone or in
combination with CCK-8. In vitro experiments revealed that either (ACh (10-8 – 10-4 M) or
CCK-8 (10" - 10-8 M)) can evoke total amylase release which was elevated in healthy
control pancreatic acinar cells compared to diabetic acinar cells. In contrast, 10-6 M insulin
produced a significant increase (Pc0.05) in the amount of total amylase output in control
acinar cells compared to diabetic acinar cells. Combining insulin (10-8 – 10-6 M) with
either ACh or CCK-8 had little or no effect on total amylase release in both control and
diabetic acinar cells. There were no significant differences among the groups in
unstimulated [Ca2+]j and [Mg2+]i. However, the peak [Ca 2+ ]i induced by 10-8 M CCK-8 was
depressed (P<0.05) in diabetic cells (275.3 ± 11.5 nM n = 8) compared to (359.7 ± 27.5
nM, n = 6) control cells. Similarly, CCK-8 significantly decreased (P<0.05) [Mg2 ']1 in
diabetic acinar cells compared to control.

On a molecular level, the gene encoding amylase was under transcriptional dysregulation.
Healthy control animals had a significantly lower (P>0.05) crossing point value (8.54 ±
0.131. n = 8) compared to STZ-induced diabetic animals (17.96 ± 0.272, n = 7),
respectively. On a protein level, those mediators controlling translation such as p70 S6K
and 4E-BPI were present at significantly lower (P>0.05) relative concentrations,
suggesting an impaired capacity for protein synthesis. Interestingly, the actual activity of
these proteins as measured by phosphorylation was slightly increased. It is suggested that
this is a cellular mechanism to counteract loss in transcription and/or translation of mRNA
encoding these proteins. Protein ubiquitination was also elevated suggesting increased
protein breakdown which could be responsible for pancreatic atrophy and net protein loss.
The NFkβ protein widely implicated in tissue atrophy was actually lower in STZ-induced
DM, and therefore probably does not contribute to pancreatic wasting.

To conclude, the results indicate that DM-induced exocrine pancreatic insufficiency is
associated with decreased levels of total protein output and amylase secretion and these
changes may be in part be associated with derangements in cellular Ca2+ and Mg2+
homeostasis. Furthermore, transcription of the α-amylase gene is reduced suggesting a
reduced protein level and thus capacity for stimulus-secretion coupling. Finally, there
appears impaired protein translation and elevated ATP-dependent protreasome mediated
protein breakdown in STZ-induced DM.

Repository Staff Only: item control page