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In the current study, to elucidate the pathological characteristics of myopic scleral stroma, three-dimensional (3D) cultures of human scleral stroma fibroblasts (HSSFs) with several axial lengths (AL, 22.80–30.63 mm) that were obtained from patients (n = 7) were examined. Among the three groups of ALs,  30 mm (n = 3), the physical properties of the 3D HSSFs spheroids with respect to size and stiffness, the expressions of extracellular matrix (ECM) molecules, including collagen (COL) 1, 4, and 6 and fibronectin (FN) by qPCR and immunohistochemistry (IHC), and the mRNA expression of ECM metabolism modulators including hypoxia-inducible factor 1A (HIF 1A), HIF 2A, lysyl oxidase (LOX), tissue inhibitor of metalloproteinase (TIMP) 1–4, and matrix metalloproteinase (MMP) 2, 9, and 14 as well as several endoplasmic reticulum (ER) stress-related factors were compared. In the largest AL group (&gt;30 mm), the 3D HSSFs spheroids were (1) significantly down-sized and less stiff compared to the other groups, and (2) significant changes were detected in the expression of some ECMs (qPCR; the up-regulation of COL1 and COL4, and the down-regulation of FN, IHC; the up-regulation of COL1 and FN, and down-regulation of COL4). The mRNA expressions of ECM modulators and ER stress-related genes were also altered with increasing AL length (up-regulation of HIF2A, MMP2, XBP1, and MMP14, down-regulation of LOX, TIMP 2 and 3, GRP78, GRP94, IRE1, and ATF6). In addition, a substantial down-regulation of some ER stress-related genes (ATF4, sXPB1 and CHOP) was observed in the 25–30 mm AL group. The findings presented herein suggest that small and stiffer 3D HSSFs spheroids in the largest AL group may accurately replicate the pathological significance of scleral thinning and weakening in myopic eyes. In addition, the modulation of several related factors among the different AL groups may also provide significant insights into our understanding of the molecular mechanisms responsible for causing myopic changes in the sclera.

current issues in molecular biology

Modulation of the Physical Properties of 3D Spheroids Derived from Human Scleral Stroma Fibroblasts (HSSFs) with Different Axial Lengths Obtained from Surgical Patients