The Role of MyoD in Myofibroblast Differentiation in Explants of Human Lens Tissue

Abstract

Purpose : Visual impairment in posterior capsule opacification (PCO) is primarily caused by myofibroblasts whose contractions deform the capsule. Myofibroblasts in explants of human lens tissue and rabbits after cataract surgery are derived from Myo/Nog cells that express the skeletal muscle specific transcription factor MyoD, bone morphogenetic protein inhibitor noggin and brain-specific angiogenesis inhibitor 1 (BAI1). In this study we tested whether MyoD was required for Myo/Nog cell differentiation into myofibroblasts.

Methods : Anterior lens tissue was obtained from patients undergoing cataract surgery. Tissue was incubated in serum free medium containing MYOD1 siRNA. Controls included no treatment, vehicle only and non-targeting siRNA. Cells were fixed on day 5. Double label immunofluorescence localization was performed with antibodies to MyoD, BAI1, Noggin, alpha smooth muscle actin (α-SMA), striated muscle myosin and Ki67, a marker of cell proliferation. Wrinkles in bare areas of the capsule were quantified by differential interference contrast microscopy.

Results : MyoD, α-SMA and striated muscle myosin were present in most BAI1 and Noggin-positive Myo/Nog cells but not in lens epithelial cells (LECs) in control cultures. MyoD protein was detected in less than 1% of Myo/Nog cells treated with MyoD siRNA. Knockdown of MyoD increased the number of Myo/Nog cells approximately 3-fold in parallel with an increase in Ki67 labeling. The effect of MyoD siRNA on α-SMA was variable. Few Myo/Nog cells contained detectable levels of striated muscle myosin with MyoD knockdown. In control cultures, areas denuded of LECs were surrounded by differentiated Myo/Nog cells and contained wrinkles in the capsule. Myo/Nog cells lacking striated muscle myosin surrounded holes in the epithelium in explants treated with MyoD siRNA; however, wrinkles in the capsule were significantly reduced. LECs appeared to be unaffected by MyoD siRNA.

Conclusions : MyoD appears to regulate the expression of striated muscle myosin in Myo/Nog cells as they differentiate into myofibroblasts in primary cultures of human lens tissue. α-SMA expression may be regulated independently of MyoD or synthesized prior to complete knockdown of MyoD with a slow rate of protein turnover. MyoD also is involved in withdrawal from the cell cycle. The presence of striated muscle myosin correlates with myofibroblast contractions that produce wrinkles in the capsule.

This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.