Automated and online LC-MS/MS for multiplexed monitoring of hormone secretion from murine islets.

Abstract

Islets of Langerhans are the endocrine microorgans responsible for secretion of hormone peptides to regulate blood glucose. Five endocrine cell types make up islets, each releasing specific peptide hormone(s) in response to extra- and intra-islet signals. Secretory dysfunction is associated with several metabolic diseases; development of analytical technologies that can temporally resolve multiple secreted factors would therefore aid in understanding factors that govern healthy and disrupted secretion from these cells. To this end, an online liquid chromatography with tandem mass spectrometry (LC-MS/MS) approach that had been developed for analysis of hormone secretion from human islets of Langerhans was modified to be used with murine islets. Rodent islets provide easily accessible samples for many researchers and mimic the biology of human islets; however, there have been no published reports on the development of analytical approaches for measuring multi-hormone samples from these species. Online perfusion of ~ 30 murine islets was performed in a microfluidic system with the perfusate coupled to a 1.5 min LC separation via a six-port valve. The valve was automated to achieve a ~ 2 min sampling period from the perfusate, providing temporally resolved measurements of somatostatin, glucagon, insulin I, and insulin II. The triple quadrupole MS provided < 0.5 nM limits of quantitation for each peptide. Secretion from murine islets in response to elevated glucose was performed, as well as more complex sinusoidal glucose levels. The automation, quantitation, and ability to identify multiple hormone species simultaneously is expected to provide new insight into the factors that govern hormone secretion from islets.