Corneal transplantation remains a critical treatment option for individuals with corneal disorders, but it faces challenges such as rejection, high associated medical costs, and donor scarcity. A promising alternative for corneal replacement involves fabricating artificial cornea from a patient’s own cells. Our study aimed to leverage bioprinting to develop a corneal model using human corneal stromal cells embedded in a collagen-based bioink. We generated both cellular and acellular collagen I (COL I) constructs. Cellular constructs were cultured for up to 4 weeks, and gene expression analysis was performed to assess extracellular matrix (ECM) remodeling and fibrotic markers. Our results demonstrated a significant decrease in the expression of COL I, collagen III (COL III), vimentin (VIM), and vinculin (VCL), indicating a dynamic remodeling process towards a more physiologically relevant corneal ECM. Overall, our study provides a foundational framework for developing customizable, corneal replacements using bioprinting technology. Further research is necessary to optimize the bioink composition and evaluate the functional and biomechanical properties of these bioengineered corneas.

Keratoconus (KC) is a corneal thinning dystrophy that leads to visual impairment. While the cause of KC remains poorly understood, changes in sex hormone levels have been correlated with KC development. This study investigated circulating gonadotropin-releasing hormone (GnRH) in control and KC subjects to determine if this master hormone regulator is linked to the KC pathology. Plasma and saliva were collected from KC subjects (n = 227 and n = 274, respectively) and non-KC controls (n = 58 and n = 101, respectively), in concert with patient demographics and clinical features. GnRH levels in both plasma and saliva were significantly lower in KC subjects compared to controls. This finding was retained in plasma when subjects were stratified based on age, sex, and KC severity. Control and KC corneal fibroblasts (HKCs) stimulated with recombinant GnRH protein in vitro revealed significantly increased luteinizing hormone receptor by HKCs and reduced expression of α-smooth muscle actin with treatment suggesting that GnRH may modulate hormonal and fibrotic responses in the KC corneal stroma. Further studies are needed to reveal the role of the hypothalamic–pituitary–gonadal axis in the onset and progression of KC and to explore this pathway as a novel therapeutic target.

Diabetic keratopathy is a corneal complication of diabetes mellitus (DM). Patients with diabetic keratopathy are prone to developing corneal haze, scarring, recurrent erosions, and significant wound healing defects/delays. The purpose of this study was to determine the contractility profiles in the diabetic human corneal stromal cells and characterize their molecular signatures. Primary human corneal fibroblasts from healthy, Type 1 DM (T1DM), and Type 2 DM (T2DM) donors were cultured using an established 3D collagen gel model. We tracked, measured, and quantified the contractile footprint over 9 days and quantified the modulation of specific corneal/diabetes markers in the conditional media and cell lysates using western blot analysis. Human corneal fibroblasts (HCFs) exhibited delayed and decreased contractility compared to that from T1DMs and T2DMs. Compared to HCFs, T2DMs demonstrated an initial downregulation of collagen I (day 3), followed by a significant upregulation by day 9. Collagen V was significantly upregulated in both T1DMs and T2DMs based on basal secretion, when compared to HCFs. Cell lysates were upregulated in the myofibroblast-associated marker, α-smooth muscle actin, in T2DMs on day 9, corresponding to the significant increase in contractility rate observed at the same time point. Furthermore, our data demonstrated a significant upregulation in IGF-1 expression in T2DMs, when compared to HCFs and T1DMs, at day 9. T1DMs demonstrated significant downregulation of IGF-1 expression, when compared to HCFs. Overall, both T1DMs and T2DMs exhibited increased contractility associated with fibrotic phenotypes. These findings, and future studies, may contribute to better understanding of the pathobiology of diabetic keratopathy and ultimately the development of new therapeutic approaches.

This study investigated the interplay between transforming growth factor beta (TGF-β1/T1 and TGF-β3/T3), and sex hormone receptors using our 3D in vitro cornea stroma model. Primary human corneal fibroblasts (HCFs) from healthy donors were plated in transwells at 10⁶ cells/well and cultured for four weeks. HCFs were supplemented with stable vitamin C (VitC) and stimulated with T1 or T3. 3D construct proteins were analyzed for the androgen receptor (AR), progesterone receptor (PR), estrogen receptor alpha (ERα) and beta (ERβ), luteinizing hormone receptor (LHR), follicle-stimulating hormone receptor (FSHR), gonadotropin-releasing hormone receptor (GnRHR), KiSS1-derived peptide receptor (KiSS1R/GPR54), and follicle-stimulating hormone subunit beta (FSH-B). In female constructs, T1 significantly upregulated AR, PR, ERα, FSHR, GnRHR, and KiSS1R. In male constructs, T1 significantly downregulated FSHR and FSH-B and significantly upregulated ERα, ERβ, and GnRHR. T3 caused significant upregulation in expressions PR, ERα, ERβ, LHR, FSHR, and GNRHR in female constructs, and significant downregulation of AR, ERα, and FSHR in male constructs. Semi-quantitative Western blot findings present the interplay between sex hormone receptors and TGF-β isoforms in the corneal stroma, which is influenced by sex as a biological variable (SABV). Additional studies are warranted to fully delineate their interactions and signaling mechanisms.