Ultrastructural and immunohistochemical analysis of the 8-20 week human fetal pancreas

Abstract

Abbreviations: SEM, scanning electron microscopy; TEM, transmission electron microscopy; CK19, Cytokeratin 19; ECM, extracellular matrix; PDX-1, pancreatic and duodenal homeobox 1.Development of the human pancreas is well-known to involve tightly controlled differentiation of pancreatic precursors to mature cells that express endocrine-or exocrine-specific protein products. However, details of human pancreatic development at the ultrastructural level are limited. The present study analyzed 8-20 week fetal age human pancreata using scanning and transmission electron microscopy (TEM), TEM immunogold and double or triple immunofluorescence staining. Primary organization of islets and acini occurred during the developmental period examined. Differentiating endocrine and exocrine cells developed from the ductal tubules and subsequently formed isolated small clusters. Extracellular matrix fibers and proteins accumulated around newly differentiated cells during their migration and cluster formation. Glycogen expression was robust in ductal cells of the pancreas from 8-15 weeks of fetal age; however, this became markedly reduced at 20 weeks, with a concomitant increase in acinar cell glycogen content. Insulin secretory granules transformed from being dense and round at 8 weeks to distinct geometric (multilobular, crystalline) structures by 14-20 weeks. Initially many of the differentiating endocrine cells were multihormonal and contained polyhormonal granules; by 20 weeks, monohormonal cells were in the majority. Interestingly, certain secretory granules in the early human fetal pancreatic cells showed positivity for both exocrine (amylase) and endocrine proteins. This combined ultrastructural and immunohistochemical study showed that, during early developmental stages, the human pancreas contains differentiating epithelial cells that associate closely with the extracellular matrix, have dynamic glycogen expression patterns and contain polyhormonal as well as mixed endocrine/ exocrine granules.

SOX9+/PTF1A+ Cells Define the Tip Progenitor Cells of the Human Fetal Pancreas of the Second Trimester

“…D), suggesting that patterning of the tip cells occurred in the hFP between 13.5 and 17WGA. However, contrary to a prior report , we did not observe expression of amylase at the protein level in any specimen (Fig. C).…”

Pancreatic β cell identity requires continual repression of non–β cell programs

“…The development of polyhormonal cells during T1D and T2D has been postulated to be indicative of a more primitive endocrine cell phenotype that is known to exist during human islet cell development (38). Interestingly, during the early stages of human pancreas formation, the endocrine cells are initially polyhormonal and only gradually resolve into monohormonal cell identities (39).…”

Human pancreas development

“…More recently, the generation of bihormonal cells in vitro has been associated with the use of BMP inhibitors during pancreatic specification (Russ et al, 2015). However, it is perhaps an over-simplification to regard these insulin + /glucagon + dual-stained cells as an in vitro artifact; they are a normal aspect of human fetal development (Polak et al, 2000;Piper et al, 2004;Sarkar et al, 2008;Jeon et al, 2009;Riedel et al, 2012;Riopel et al, 2014). The frequency of their detection has been strikingly variable, from ∼5% to 92% of endocrine cells, perhaps owing to differing sensitivity of immunohistochemistry protocols across different groups or variations in the age of material studied.…”