Immunohistochemical Detection of CTGF in the Human Eye

<p><i>Purpose/Aim of the study</i>: Connective tissue growth factor (CTGF) is a key player in the control of extracellular matrix remodeling, fibrosis, and angiogenesis. It is also involved in the modification of the trabecular meshwork, thus potentially modulating outflow facility and intraocular pressure (IOP). As a consequence, CTGF might be relevant for the development of elevated IOP, a major risk factor in glaucoma-pathogenesis. While comprehensive information on the origins of CTGF in the human eye is not available, the goal of this study is to identify ocular sources of CTGF using morphological methods.</p> <p><i>Materials and Methods</i>: Human donor eyes were prepared for immunohistochemical analysis of CTGF, α-smooth muscle-actin (ASMA), and CD31. Confocal laser scanning microscopy was used for documentation.</p> <p><i>Results</i>: In the cornea, CTGF-immunoreactivity (CTGF-IR) was detected in the epithelium, mainly in basal layers, stromal keratinocytes, and endothelial cells. Adjacent conjunctiva showed also CTGF-IR in epithelial cells. In the iris, both, the sphincter and dilator muscles displayed CGTF-IR, as did iris and ciliary body vessels, deriving at this location from the vascular endothelium, as detected with CD31, but not from vascular smooth muscle cells, as detected with ASMA. In the ciliary body, CTGF-IR was detected in smooth-muscle cells of the ciliary muscle and further in the non-pigmented epithelium. In the retina, CTGF-IR was detected in the NFL and weakly in the IPL/OPL. In the choroid, the choriocapillaris and blood vessels displayed CTGF-IR. Further, few cells in the optic nerve head and the lamina cribrosa were CTGF-positive.</p> <p><i>Conclusion</i>: CTGF was detected in various structures of the human eye. Since CTGF has been also described in aqueous humor, the identified structures might be the sources of CTGF in the aqueous humor. By means of aqueous flow, CTGF is transported into the trabecular meshwork, where it could change outflow facility and therefore affecting IOP homeostasis.</p

Table1_Pericyte-derived cells participate in optic nerve scar formation.DOCX

Introduction: Pericytes (PCs) are specialized cells located abluminal of endothelial cells on capillaries, fulfilling numerous important functions. Their potential involvement in wound healing and scar formation is achieving increasing attention since years. Thus, many studies investigated the participation of PCs following brain and spinal cord (SC) injury, however, lacking in-depth analysis of lesioned optic nerve (ON) tissue. Further, due to the lack of a unique PC marker and uniform definition of PCs, contradicting results are published.Methods: In the present study the inducible PDGFRβ-P2A-CreERT2-tdTomato lineage tracing reporter mouse was used to investigate the participation and trans-differentiation of endogenous PC-derived cells in an ON crush (ONC) injury model, analyzing five different post lesion time points up to 8 weeks post lesion.Results: PC-specific labeling of the reporter was evaluated and confirmed in the unlesioned ON of the reporter mouse. After ONC, we detected PC-derived tdTomato+ cells in the lesion, whereof the majority is not associated with vascular structures. The number of PC-derived tdTomato+ cells within the lesion increased over time, accounting for 60–90% of all PDGFRβ+ cells in the lesion. The presence of PDGFRβ+tdTomato- cells in the ON scar suggests the existence of fibrotic cell subpopulations of different origins.Discussion: Our results clearly demonstrate the presence of non-vascular associated tdTomato+ cells in the lesion core, indicating the participation of PC-derived cells in fibrotic scar formation following ONC. Thus, these PC-derived cells represent promising target cells for therapeutic treatment strategies to modulate fibrotic scar formation to improve axonal regeneration.</p

Image1_Pericyte-derived cells participate in optic nerve scar formation.JPEG

Introduction: Pericytes (PCs) are specialized cells located abluminal of endothelial cells on capillaries, fulfilling numerous important functions. Their potential involvement in wound healing and scar formation is achieving increasing attention since years. Thus, many studies investigated the participation of PCs following brain and spinal cord (SC) injury, however, lacking in-depth analysis of lesioned optic nerve (ON) tissue. Further, due to the lack of a unique PC marker and uniform definition of PCs, contradicting results are published.Methods: In the present study the inducible PDGFRβ-P2A-CreERT2-tdTomato lineage tracing reporter mouse was used to investigate the participation and trans-differentiation of endogenous PC-derived cells in an ON crush (ONC) injury model, analyzing five different post lesion time points up to 8 weeks post lesion.Results: PC-specific labeling of the reporter was evaluated and confirmed in the unlesioned ON of the reporter mouse. After ONC, we detected PC-derived tdTomato+ cells in the lesion, whereof the majority is not associated with vascular structures. The number of PC-derived tdTomato+ cells within the lesion increased over time, accounting for 60–90% of all PDGFRβ+ cells in the lesion. The presence of PDGFRβ+tdTomato- cells in the ON scar suggests the existence of fibrotic cell subpopulations of different origins.Discussion: Our results clearly demonstrate the presence of non-vascular associated tdTomato+ cells in the lesion core, indicating the participation of PC-derived cells in fibrotic scar formation following ONC. Thus, these PC-derived cells represent promising target cells for therapeutic treatment strategies to modulate fibrotic scar formation to improve axonal regeneration.</p