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

While the lens is an avascular tissue with an immune-privileged status, studies have now revealed that there are immune responses specifically linked to the lens. The response to lens injury, such as following cataract surgery, has been shown to involve the activation of the resident immune cell population of the lens and the induction of immunomodulatory factors by the wounded epithelium. However, there has been limited investigation into the immediate response of the lens to wounding, particularly those induced factors that are intrinsic to the lens and its associated resident immune cells. Using an established chick embryo ex vivo cataract surgery model has made it possible to determine the early immune responses of this tissue to injury, including its resident immune cells, through a transcriptome analysis. RNA-seq studies were performed to determine the gene expression profile at 1 h post wounding compared to time 0. The results provided evidence that, as occurs in other tissues, the resident immune cells of the lens rapidly acquired a molecular signature consistent with their activation. These studies also identified the expression of many inflammatory factors by the injured lens that are associated with both the induction and regulation of the immune response.

Fibrosis-the tale of H3K27 histone methyltransferases and demethylases.

Fibrosis, or excessive scarring, is characterized by the emergence of alpha-smooth muscle actin (αSMA)-expressing myofibroblasts and the excessive accumulation of fibrotic extracellular matrix (ECM). Currently, there is a lack of effective treatment options for fibrosis, highlighting an unmet need to identify new therapeutic targets. The acquisition of a fibrotic phenotype is associated with changes in chromatin structure, a key determinant of gene transcription activation and repression. The major repressive histone mark, H3K27me3, has been linked to dynamic changes in gene expression in fibrosis through alterations in chromatin structure. H3K27-specific homologous histone methylase (HMT) enzymes, Enhancer of zeste 1 and 2 (EZH1, EZH2), which are the alternative subunits of the Polycomb Repressive Complex 2 (PRC2) and demethylase (KDM) enzymes, Ubiquitously transcribed tetratricopeptide repeat, X chromosome (UTX), and Lysine demethylase 6B (KDM6B), are responsible for regulating methylation status of H3K27me3. In this review, we explore how these key enzymes regulate chromatin structure to alter gene expression in fibrosis, highlighting them as attractive targets for the treatment of fibrosis.

Changes in nascent chromatin structure regulate activation of the pro-fibrotic transcriptome and myofibroblast emergence in organ fibrosis.

Cell reprogramming to a myofibroblast responsible for the pathological accumulation of extracellular matrix is fundamental to the onset of fibrosis. Here, we explored how condensed chromatin structure marked by H3K72me3 becomes modified to allow for activation of repressed genes to drive emergence of myofibroblasts. In the early stages of myofibroblast precursor cell differentiation, we discovered that H3K27me3 demethylase enzymes UTX/KDM6B creates a delay in the accumulation of H3K27me3 on nascent DNA revealing a period of decondensed chromatin structure. This period of decondensed nascent chromatin structure allows for binding of pro-fibrotic transcription factor, Myocardin-related transcription factor A (MRTF-A) to nascent DNA. Inhibition of UTX/KDM6B enzymatic activity condenses chromatin structure, prevents MRTF-A binding, blocks activation of the pro-fibrotic transcriptome, and results in an inhibition of fibrosis in lens and lung fibrosis models. Our work reveals UTX/KDM6B as central coordinators of fibrosis, highlighting the potential to target its demethylase activity to prevent organ fibrosis.

PI3K Isoform-Specific Regulation of Leader and Follower Cell Function for Collective Migration and Proliferation in Response to Injury.

To ensure proper wound healing it is important to elucidate the signaling cues that coordinate leader and follower cell behavior to promote collective migration and proliferation for wound healing in response to injury. Using an ex vivo post-cataract surgery wound healing model we investigated the role of class I phosphatidylinositol-3-kinase (PI3K) isoforms in this process. Our findings revealed a specific role for p110α signaling independent of Akt for promoting the collective migration and proliferation of the epithelium for wound closure. In addition, we found an important role for p110α signaling in orchestrating proper polarized cytoskeletal organization within both leader and wounded epithelial follower cells to coordinate their function for wound healing. p110α was necessary to signal the formation and persistence of vimentin rich-lamellipodia extensions by leader cells and the reorganization of actomyosin into stress fibers along the basal domains of the wounded lens epithelial follower cells for movement. Together, our study reveals a critical role for p110α in the collective migration of an epithelium in response to wounding.

The local wound environment is a key determinant of the outcome of TGFß signaling on the fibrotic response of CD44+ leader cells in an ex vivo post-cataract-surgery model.

The cytokine transforming growth factor beta (TGFß) has a role in regulating the normal and pathological response to wound healing, yet how it shifts from a pro-repair to a pro-fibrotic function within the wound environment is still unclear. Using a clinically relevant ex vivo post-cataract surgery model that mimics the lens fibrotic disease posterior capsule opacification (PCO), we investigated the influence of two distinct wound environments on shaping the TGFß-mediated injury response of CD44+ vimentin-rich leader cells. The substantial fibrotic response of this cell population occurred within a rigid wound environment under the control of endogenous TGFß. However, TGFß was dispensable for the role of leader cells in wound healing on the endogenous basement membrane wound environment, where repair occurs in the absence of a major fibrotic outcome. A difference between leader cell function in these distinct environments was their cell surface expression of the latent TGFß activator, αvß3 integrin. This receptor is exclusively found on this CD44+ cell population when they localize to the leading edge of the rigid wound environment. Providing exogenous TGFß to bypass any differences in the ability of the leader cells to sustain activation of TGFß in different environments revealed their inherent ability to induce pro-fibrotic reactions on the basement membrane wound environment. Furthermore, exposure of the leader cells in the rigid wound environment to TGFß led to an accelerated fibrotic response including the earlier appearance of pro-collagen + cells, alpha smooth muscle actin (αSMA)+ myofibroblasts, and increased fibrotic matrix production. Collectively, these findings show the influence of the local wound environment on the extent and severity of TGFß-induced fibrotic responses. These findings have important implications for understanding the development of the lens fibrotic disease PCO in response to cataract surgery wounding.