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A Method for Mouse Pancreatic Islet Isolation and Intracellular cAMP Determination
Published on: June 25, 2014
Biochemical and clinical implications of proinsulin conversion intermediates
The Journal of Clinical Investigation
|October 1, 1985
Summary
This study investigated proinsulin processing in humans, finding that cleavage at the Arg32-Glu33 bond is preferred. Subcutaneous administration of proinsulin leads to intermediate formation, unlike intravenous delivery.
Area of Science:
- Biochemistry
- Endocrinology
- Molecular Biology
Background:
- Proinsulin is the precursor to insulin, requiring specific cleavage steps for maturation.
- Understanding proinsulin conversion intermediates is crucial for mapping insulin-related peptides.
- Previous studies have identified various split proinsulin forms, but their in vivo dynamics are not fully elucidated.
Purpose of the Study:
- To investigate the formation of proinsulin conversion intermediates in vitro and in vivo.
- To determine the ratio of des-Arg31,Arg32-proinsulin to des-Lys64,Arg65-proinsulin in normal and pathological human tissues.
- To analyze the processing of human proinsulin after intravenous and subcutaneous administration in diabetic subjects.
Main Methods:
- High-performance liquid chromatography (HPLC) coupled with radioimmunoassay (RIA).
- Kinetic analysis of in vitro proinsulin processing using trypsin and carboxypeptidase B.
- Analysis of human pancreatic tissues (normal and insulinomas) and plasma samples.
Main Results:
- In vitro processing showed preferential cleavage at Arg32-Glu33, forming des-Arg31,Arg32-proinsulin and des-Lys64,Arg65-proinsulin at a 3.3:1 ratio.
- Normal pancreas tissue exhibited a similar ratio (approx. 3:1), while insulinomas predominantly contained des-Arg31,Arg32-proinsulin.
- Intravenous proinsulin infusion resulted in <1% processing, whereas subcutaneous infusion yielded 4-11% processing to intermediates.
Conclusions:
- Proinsulin to insulin conversion in vivo favors cleavage at the Arg32-Glu33 bond, similar to in vitro findings.
- Proinsulin processing is inefficient within the vascular compartment.
- Subcutaneous proinsulin administration can lead to the formation of biologically active conversion intermediates.
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