In Depth Quantification of Extracellular Matrix Proteins from Human Pancreas

Extracellular matrix (ECM) is an
important component of the pancreatic
microenvironment which regulates β cell proliferation, differentiation,
and insulin secretion. Protocols have recently been developed for
the decellularization of the human pancreas to generate functional
scaffolds and hydrogels. In this work, we characterized human pancreatic
ECM composition before and after decellularization using isobaric
dimethylated leucine (DiLeu) labeling for relative quantification
of ECM proteins. A novel correction factor was employed in the study
to eliminate the bias introduced during sample preparation. In comparison
to the commonly employed sample preparation methods (urea and FASP)
for proteomic analysis, a recently developed surfactant and chaotropic
agent assisted sequential extraction/on pellet digestion (SCAD) protocol
has provided an improved strategy for ECM protein extraction of human
pancreatic ECM matrix. The quantitative proteomic results revealed
the preservation of matrisome proteins while most of the cellular
proteins were removed. This method was compared with a well-established
label-free quantification (LFQ) approach which rendered similar expressions
of different categories of proteins (collagens, ECM glycoproteins,
proteoglycans, etc.). The distinct expression of ECM proteins was
quantified comparing adult and fetal pancreas ECM, shedding light
on the correlation between matrix composition and postnatal β
cell maturation. Despite the distinct profiles of different subcategories
in the native pancreas, the distribution of matrisome proteins exhibited
similar trends after the decellularization process. Our method generated
a large data set of matrisome proteins from a single tissue type.
These results provide valuable insight into the possibilities of constructing
a bioengineered pancreas. It may also facilitate better understanding
of the potential roles that matrisome proteins play in postnatal β
cell maturation.