Viral Vector Effects on Exoenzyme C3 Transferase-Mediated Actin Disruption and on Outflow Facility

Investigative Ophthalmology & Visual Science
Sarah R SlausonC R Brandt

Abstract

Purified Clostridium botulinum exoenzyme C3 transferase (C3) effects on the actin cytoskeleton in human trabecular meshwork cells (HTM) and on the outflow facility response in monkey organ-cultured anterior segments (MOCAS) were determined in the presence or absence of viral vectors. Human adenovirus type 5 (AdV) and feline immunodeficiency virus (FIV) vectors were produced using kits. Cell soluble purified C3 (C3cs) was purchased commercially. Recombinant C3 (C3rec) cDNA was overexpressed in Escherichia coli and purified. The HTM cells were incubated with up to 10 μg/mL C3cs or with 5 μg of C3rec and/or viral vector (multiplicity of infection [MOI] = 25). Cells then were fixed and stained for actin. Outflow facility in MOCAS was measured at baseline, 4 hours, 24 hours, and 3 to 4 days following bolus injection of AdV (1.6 × 107 transducing units) and/or 2.5 μg C3rec. The HTM cells treated for 4 hours with C3cs (all doses) or for 24 hours with C3rec developed a rounded morphology and lost stress fibers. Cells transduced with vectors alone showed no changes at any time point. Cells exposed to C3rec and cotransduced with either viral vector showed significant disruption of the actin cytoskeleton within 4 hours after exposure, whi...Continue Reading

Citations

Jun 21, 2016·Infection, Genetics and Evolution : Journal of Molecular Epidemiology and Evolutionary Genetics in Infectious Diseases·A PrisillaP Chellapandi
Oct 17, 2018·Investigative Ophthalmology & Visual Science·Junkai TanXuyang Liu
Jan 14, 2021·Experimental Eye Research·Monica M Sauter, Curtis R Brandt
Jul 14, 2021·Translational Vision Science & Technology·Coralia LunaPedro Gonzalez

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