scholarly journals Development of 3D Printed Bruch’s Membrane-Mimetic Substance for the Maturation of Retinal Pigment Epithelial Cells

2021 ◽  
Vol 22 (3) ◽  
pp. 1095
Author(s):  
Jongmin Kim ◽  
Ju Young Park ◽  
Jeong Sik Kong ◽  
Hyungseok Lee ◽  
Jae Yon Won ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
3D Printing ◽  
Retinal Pigment ◽  
Bruch's Membrane ◽  
Bruch’S Membrane ◽  
Printing Technology ◽  
Rpe Cells ◽  
3D Printing Technology ◽  
Membrane Mimetic ◽  
Rpe Transplantation

Retinal pigment epithelium (RPE) is a monolayer of the pigmented cells that lies on the thin extracellular matrix called Bruch’s membrane. This monolayer is the main component of the outer blood–retinal barrier (BRB), which plays a multifunctional role. Due to their crucial roles, the damage of this epithelium causes a wide range of diseases related to retinal degeneration including age-related macular degeneration, retinitis pigmentosa, and Stargardt disease. Unfortunately, there is presently no cure for these diseases. Clinically implantable RPE for humans is under development, and there is no practical examination platform for drug development. Here, we developed porcine Bruch’s membrane-derived bioink (BM-ECM). Compared to conventional laminin, the RPE cells on BM-ECM showed enhanced functionality of RPE. Furthermore, we developed the Bruch’s membrane-mimetic substrate (BMS) via the integration of BM-ECM and 3D printing technology, which revealed structure and extracellular matrix components similar to those of natural Bruch’s membrane. The developed BMS facilitated the appropriate functions of RPE, including barrier and clearance functions, the secretion of anti-angiogenic growth factors, and enzyme formation for phototransduction. Moreover, it could be used as a basement frame for RPE transplantation. We established BMS using 3D printing technology to grow RPE cells with functions that could be used for an in vitro model and RPE transplantation.

scholarly journals FHL-1 interacts with human RPE cells through the α5β1 integrin and confers protection against oxidative stress

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Rawshan Choudhury ◽  
Nadhim Bayatti ◽  
Richard Scharff ◽  
Ewa Szula ◽  
Viranga Tilakaratna ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Cell Fate ◽  
Retinal Pigment ◽  
Factor H ◽  
Age Related Macular Degeneration ◽  
Bruch's Membrane ◽  
Bruch’S Membrane ◽  
Rpe Cells ◽  
Age Related

AbstractRetinal pigment epithelial (RPE) cells that underlie the neurosensory retina are essential for the maintenance of photoreceptor cells and hence vision. Interactions between the RPE and their basement membrane, i.e. the inner layer of Bruch’s membrane, are essential for RPE cell health and function, but the signals induced by Bruch’s membrane engagement, and their contributions to RPE cell fate determination remain poorly defined. Here, we studied the functional role of the soluble complement regulator and component of Bruch’s membrane, Factor H-like protein 1 (FHL-1). Human primary RPE cells adhered to FHL-1 in a manner that was eliminated by either mutagenesis of the integrin-binding RGD motif in FHL-1 or by using competing antibodies directed against the α5 and β1 integrin subunits. These short-term experiments reveal an immediate protein-integrin interaction that were obtained from primary RPE cells and replicated using the hTERT-RPE1 cell line. Separate, longer term experiments utilising RNAseq analysis of hTERT-RPE1 cells bound to FHL-1, showed an increased expression of the heat-shock protein genes HSPA6, CRYAB, HSPA1A and HSPA1B when compared to cells bound to fibronectin (FN) or laminin (LA). Pathway analysis implicated changes in EIF2 signalling, the unfolded protein response, and mineralocorticoid receptor signalling as putative pathways. Subsequent cell survival assays using H2O2 to induce oxidative stress-induced cell death suggest hTERT-RPE1 cells had significantly greater protection when bound to FHL-1 or LA compared to plastic or FN. These data show a non-canonical role of FHL-1 in protecting RPE cells against oxidative stress and identifies a novel interaction that has implications for ocular diseases such as age-related macular degeneration.

Bruch’s membrane pathology: A mechanistic perspective

2020 ◽  
Vol 30 (6) ◽  
pp. 1195-1206 ◽  
Author(s):  
Aishwarya Murali ◽  
Subramanian Krishnakumar ◽  
Anuradha Subramanian ◽  
Sowmya Parameswaran
Keyword(s):  
Extracellular Matrix ◽  
Retinal Pigment Epithelium ◽  
Pigment Epithelium ◽  
Retinal Pigment ◽  
Vital Role ◽  
Age Related Macular Degeneration ◽  
Ocular Diseases ◽  
Bruch's Membrane ◽  
Bruch’S Membrane ◽  
Age Related

Bruch’s membrane, an extracellular matrix located between the retinal pigment epithelium and the choroid, plays a vital role as structural and functional support to the retinal pigment epithelium. Dysfunction of Bruch’s membrane in both age-related macular degeneration and other ocular diseases is caused mostly by extracellular matrix degeneration, deposit formation, and angiogenesis. Although these factors are dealt in greater detail with respect to the cells that are degenerated such as the retinal pigment epithelium and the endothelial cells, the pathology involving the Bruch’s membrane is often underrated. Since in most of the macular degenerations early degenerative changes are also observed in the Bruch’s membrane, addressing only the cellular component without the underlying membrane will not yield an ideal clinical benefit. This review aims to discuss the factors and the mechanisms affecting the integrity of the Bruch’s membrane, which would aid in developing an effective therapy for these pathologies.

scholarly journals FHL-1 interacts with human RPE cells through the α5β1 integrin and confers protection against oxidative stress

2020 ◽  
Author(s):  
Rawshan Choudhury ◽  
Nadhim Bayatti ◽  
Richard Scharff ◽  
Ewa Szula ◽  
Viranga Tilakaratna ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Cell Fate ◽  
Retinal Pigment ◽  
Factor H ◽  
Age Related Macular Degeneration ◽  
Bruch's Membrane ◽  
Bruch’S Membrane ◽  
Rpe Cells ◽  
Age Related

AbstractRetinal pigment epithelial (RPE) cells that underlie the neurosensory retina are essential for the maintenance of photoreceptor cells and hence vision. Interactions between the RPE and their basement membrane, i.e. the inner layer of Bruch’s membrane, are essential for RPE cell health and function, but the signals induced by Bruch’s membrane engagement, and their contributions to RPE cell fate determination remain poorly defined. Here, we studied the functional role of the soluble complement regulator and component of Bruch’s membrane, Factor H-like protein 1 (FHL-1). Human primary RPE cells adhered to FHL-1 in a manner that was eliminated by either mutagenesis of the integrin-binding RGD motif in FHL-1 or by using competing antibodies directed against the α5 and β1 integrin subunits. The results obtained from primary RPE cells were replicated using the hTERT-RPE cell line. RNAseq expression analysis of hTERT-RPE cells bound to FHL-1 showed an increased expression of the heat-shock protein genes HSPA6, CRYAB, HSPA1A and HSPA1B when compared to cells bound to fibronectin (FN) or laminin (LA). Pathway analysis implicated changes in EIF2 signalling, the unfolded protein response, and mineralocorticoid receptor signalling as putative pathways. Subsequent cell survival assays using H2O2 to induce oxidative stress-induced cell death showed hTERT-RPE cells had significantly greater protection when bound to FHL-1 or LA compared to plastic or FN. These data show a non-canonical role of FHL-1 in protecting RPE cells against oxidative stress and identifies a novel interaction that has implications for ocular diseases such as age-related macular degeneration.

scholarly journals Genetic LAMP2 deficiency accelerates the age-associated formation of basal laminar deposits in the retina

2019 ◽  
Vol 116 (47) ◽  
pp. 23724-23734 ◽  
Author(s):  
Shoji Notomi ◽  
Kenji Ishihara ◽  
Nikolaos E. Efstathiou ◽  
Jong-Jer Lee ◽  
Toshio Hisatomi ◽  
...  
Keyword(s):  
Basal Lamina ◽  
Pigment Epithelium ◽  
Retinal Pigment ◽  
Critical Role ◽  
Age Related Macular Degeneration ◽  
Bruch's Membrane ◽  
Bruch’S Membrane ◽  
Rpe Cells ◽  
Age Related ◽  
Deficient Mice

The early stages of age-related macular degeneration (AMD) are characterized by the accumulation of basal laminar deposits (BLamDs). The mechanism for BLamDs accumulating between the retinal pigment epithelium (RPE) and its basal lamina remains elusive. Here we examined the role in AMD of lysosome-associated membrane protein-2 (LAMP2), a glycoprotein that plays a critical role in lysosomal biogenesis and maturation of autophagosomes/phagosomes. LAMP2 was preferentially expressed by RPE cells, and its expression declined with age. Deletion of the Lamp2 gene in mice resulted in age-dependent autofluorescence abnormalities of the fundus, thickening of Bruch’s membrane, and the formation of BLamDs, resembling histopathological changes occurring in AMD. Moreover, LAMP2-deficient mice developed molecular signatures similar to those found in human AMD—namely, the accumulation of APOE, APOA1, clusterin, and vitronectin—adjacent to BLamDs. In contrast, collagen 4, laminin, and fibronectin, which are extracellular matrix proteins constituting RPE basal lamina and Bruch’s membrane were reduced in Lamp2 knockout (KO) mice. Mechanistically, retarded phagocytic degradation of photoreceptor outer segments compromised lysosomal degradation and increased exocytosis in LAMP2-deficient RPE cells. The accumulation of BLamDs observed in LAMP2-deficient mice was eventually followed by loss of the RPE and photoreceptors. Finally, we observed loss of LAMP2 expression along with ultramicroscopic features of abnormal phagocytosis and exocytosis in eyes from AMD patients but not from control individuals. Taken together, these results indicate an important role for LAMP2 in RPE function in health and disease, suggesting that LAMP2 reduction may contribute to the formation of BLamDs in AMD.

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