Naked plasmid-mediated gene transfer to skeletal muscle ameliorates diabetes mellitus

Abstract

Background

The ability of tissues to take up naked plasmid DNA in vivo suggests an approach for reconstituting systemic metabolic deficiencies without the disadvantages of viral vectors and lipid-DNA complexes. Plasmid-mediated gene transfer into skeletal muscle was investigated as a means of providing a therapeutic source of insulin.

Methods

Four plasmid constructs, each bearing a mouse furin cDNA transgene and rat proinsulin cDNA (modified for processing by furin) driven by four different promoters were injected into the calf muscles of male Balb/c mice. Insulin and C-peptide concentrations were measured by radioimmunoassays having minimal crossreactivity for proinsulin and partially processed proinsulin.

Results

Intramuscular insulin concentrations increased by up to 3.6-fold over controls seven days after single injections of CMV, β-actin, hsp70 and myoglobin promoter constructs. The optimal dose for most constructs was 100 µg plasmid DNA. Intramuscular plasmid injection into streptozotocin-induced diabetic Balb/c mice raised plasma insulin and C-peptide concentrations, and reduced hyperglycaemia. Two injections (100 µg plasmid DNA each) caused higher plasma insulin concentrations and significantly reduced hyperglycemia in diabetic mice than a single injection. Best results were obtained when plasmid injections preceded induction of diabetes by 14 days.

Conclusions

Skeletal muscle is a potentially useful platform for ectopic secretion of insulin using naked plasmid as a gene transfer vector. Injection at two sites 14 days before the onset of severe hyperglycemia is optimal. This approach could protect Type I diabetics from fatal ketoacidosis and enhance the action of agents that sensitize tissues to insulin in type II diabetes. Copyright © 1999 John Wiley & Sons, Ltd.