We present a method to investigate the functional beta cell mass in the pancreas,
defined as the number of beta cells and their functional state. It consists in
determining total beta cell numbers in the organ and their distribution over
functionally and/or topographically different subpopulations. The procedure was
developed in the adult rodent pancreas and then applied to models of beta cell growth
and regeneration. Prior quantifications of the beta cell population were mostly indirect,
involving an assessment of the weight or volume of the beta cell mass in pancreatic
sections; they were also often conducted on samples with unknown representativity.
Consequently, differences in measured values do not necessarily reflect
(patho)physiological differences in beta cell number. With the present method, beta
cell numbers are directly obtained through an automatic nuclear count in a
representative pancreas sample, keeping the measurement error under 10%. In
addition, the distribution of the beta cell population was determined over
subpopulations that differed in their degree of aggregation and/or of expression of a
major functional marker: the size of the cellular insulin store. In 10-week old rats, the
pancreas was found to contain 3 million beta cells, distributed over 40 000 insulinpositive
units with different size ranging from single cells to aggregates with a
diameter of 500?m. Units smaller than 50µm were most numerous (>90 percent) but
contained only 10% of the total beta cell number; those larger than 200?m
represented a minority (10 percent) while containing 50% of the cells. Beta cells in
small units presented a higher insulin staining intensity than those in larger
aggregates; moreover, they were significantly less responsive to a glibenclamideinduced
degranulation. Our in situ observations indicate that studies on isolated islets
can be seen as representative for a large beta cell subpopulation but not for the smaller
fraction of beta cells that is present in the majority of beta cell units, i.e. those smaller
than 50?m. The number of units after birth (from 7000 to 27000) and then remains constant. In addition, a minor
number of these small units grow to a larger size, a process which continuously
proceeds throughout the 10 week study period. This combination of neogenesis and
growth of beta cell units is responsible for postnatal formation of 90 percent of the
beta cells counted at week 10. The majority of neoformed beta cells is present in
aggregates of larger size; they appear the result, at least in part, of replication of a beta
2 cell subpopulation which has grown in size during the first 4 weeks and then remained
constant; it is at present unclear to which extent this growth is also the result of fusion
of small beta cell units. Our methods and observations on postnatal growth of the beta
cell population in rats were taken as basis to investigate the occurrence of beta cell
regeneration in duct-ligated mouse pancreases, which have been reported to exhibit an
increased beta cell weight two weeks after injury. We confirmed the increase in beta
cell weight following pancreatic duct ligation (PDL) but this was not associated with
an increase in the number of beta cells; the discrepancy was attributed to interference
of the severely altered PDL-pancreatic tissue with measurements of beta cell mass.
PDL did induce a neoformation of small beta cell units in the adult mouse pancreas,
but this was not accompanied by growth of more particles to a larger size. Further
work is needed to identify factors that induce neogenesis and those that stimulate
growth to larger aggregates. Our method provides a useful tool for these studies,
allowing also a quantification of the associated increases in beta cell number and an
assessment of the functionally different beta cell subpopulations.
Datum prijs7 sep 2012
Originele taalEnglish
Prijsuitreikende instantie
  • Vrije Universiteit Brussel
BegeleiderDaniel Pipeleers (Promotor), Miriam Marichal (Co-promotor), Zhidong Ling (Co-promotor), Christiaan Van Schravendijk (Jury), Tony Lahoutte (Jury), Luc Bouwens (Jury), A. Lernmark (Jury) & Frank Roels (Jury)

Citeer dit