Delivery of sh RNA via lentivirus in human pseudoislets provides a model to test dynamic regulation of insulin secretion and gene function in human islets

M Harata, etc
Physiological Reports, 2018

Rodent islets are widely used to study the pathophysiology of beta cells and
islet function, however, structural and functional differences exist between
human and rodent islets, highlighting the need for human islet studies.
Human islets are highly variable, deteriorate during culture, and are difficult
to genetically modify, making mechanistic studies difficult to conduct and
reproduce. To overcome these limitations, we tested whether pseudoislets, created
by dissociation and reaggregation of islet cell suspensions, allow for
assessment of dynamic islet function after genetic modulation. Characterization
of pseudoislets cultured for 1 week revealed better preservation of
first-phase glucose-stimulated insulin secretion (GSIS) compared with cultured-intact
islets and insulin secretion profiles similar to fresh islets when
challenged by glibenclamide and KCl. qPCR indicated that pseudoislets are
similar to the original islets for the expression of markers for cell types, beta
cell function, and cellular stress with the exception of reduced proinflammatory
cytokine genes (IL1B, CCL2, CXCL8). The expression of extracellular
matrix markers (ASPN, COL1A1, COL4A1) was also altered in pseudoislets
compared with intact islets. Compared with intact islets transduced by adenovirus,
pseudoislets transduced by lentivirus showed uniform transduction
and better first-phase GSIS. Lastly, the lentiviral-mediated delivery of short
hairpin RNA targeting glucokinase (GCK) achieved significant reduction of
GCK expression in pseudoislets as well as marked reduction of both first and
second phase GSIS without affecting the insulin secretion in response to KCl.
Thus, pseudoislets are a tool that enables efficient genetic modulation of
human islet cells while preserving insulin secretion.

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Physiological Reports
doi: 10.14814/phy2.13907
University of Iowa,