The Immediate Early Response of Lens Epithelial Cells to Lens Injury

Abstract

Cataracts are treated by lens fiber cell removal followed by intraocular lens (IOL) implantation into the lens capsule. While effective, this procedure leaves behind numerous lens epithelial cells (LECs) which undergo a wound healing response that frequently leads to posterior capsular opacification (PCO). In order to elucidate the acute response of LECs to lens fiber cell removal which models cataract surgery (post cataract surgery, PCS), RNA-seq was conducted on LECs derived from wild type mice at 0 and 6 h PCS. This analysis found that LECs upregulate the expression of numerous proinflammatory cytokines and profibrotic regulators by 6 h PCS suggesting rapid priming of pathways leading to inflammation and fibrosis PCS. LECs also highly upregulate the expression of numerous immediate early transcription factors (IETFs) by 6 h PCS and immunolocalization found elevated levels of these proteins by 3 h PCS, and this was preceded by the phosphorylation of ERK1/2 in injured LECs. Egr1 and FosB were among the highest expressed of these factors and qRT-PCR revealed that they also upregulate in explanted mouse lens epithelia suggesting potential roles in the LEC injury response. Analysis of lenses lacking either Egr1 or FosB revealed that both genes may regulate a portion of the acute LEC injury response, although neither gene was essential for expression of either proinflammatory or fibrotic markers at later times PCS suggesting that IETFs may work in concert to mediate the LEC injury response following cataract surgery.

Immune Responses Induced at One Hour Post Cataract Surgery Wounding of the Chick Lens

“…This approach made it possible to focus on the response of the injured epithelium and its associated resident immune cells in the absence of any immune contribution from outside of the lens. In this reductionist lens wounding model, we found that the immediate early transcription factors were induced within 1 h of injury, including the transcription factors Egr1 and FosB (Supplemental File S1), which had been previously reported for the mouse model at 6 h post cataract surgery [27]. Also, consistent with studies performed at later times post lens wounding, we found that pro-inflammatory cytokine genes are induced within 1 h post lens wounding, which at this early time included the chemokine CCL4, also known as macrophage inflammatory protein.…”

“…The cytokines induced are consistent with a role in the activation of an innate immune response, with cytokine expression preceding the detection of neutrophils with the post-cataract surgery lens tissue [9]. The upregulation of pro-inflammatory cytokines was also detected in studies performed at 6 h post mouse cataract surgery [27].…”

“…However, they did not address the specific response of the lens-associated resident immune cells, including their role in orchestrating the recruitment of immune cells to the lens and modulating the functional outcome of the lens to insult. Previous RNA-seq analyses showed the induction of immediate early transcription factors, including Egr1 and FosB, in addition to pro-inflammatory cytokines, at 6 h after ex vivo mouse cataract surgery, and that Egr1 and FosB were also induced within 6 h of lens epithelial wounding performed ex vivo [27]. In the mouse cataract surgery model, it was also found that, by 24 h post surgery, genes associated with the innate immune response are induced [9].…”

ATF4 May Be Essential for Adaption of the Ocular Lens to Its Avascular Environment

“…It is currently unknown how Atf4 transcript levels are regulated in the developing lens, the precise time point in lens development when ATF4 function becomes necessary, the key mechanisms that fine-tune its function to drive appropriate lens amino acid levels, or whether/how it interfaces with HIF1a during adaption of the lens to chronic tissue hypoxia. Notably, Atf4 mRNA levels also upregulate sharply in lens epithelial cells by 6 h after lens injury and remain elevated for several days, while Hif1a mRNA levels decrease in injured LECs over this same time frame [22,61,108,109]. As ATF4 is best known as a mediator of cellular responses to acute stress in other tissues [26], the lens may be an ideal system to investigate how the diverse functions of Atf4 are differentially regulated during both normal tissue development and as a consequence of injury responses.…”