Elevated glucose alters global gene expression and tenascin-C alternative splicing in mesangial cells

2023

Front. Cell. Neurosci.

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The composition of the extracellular matrix (ECM) in nervous tissue plays an important role in controlling neuronal outgrowth and synapse development. Changes in both protein and glycosaminoglycan components of the ECM occur with tissue injury and may affect neuron growth. To investigate neuron responses to alterations in fibronectin (FN), a major component of the wound ECM, we grew cortical neurons on cell-derived decellularized matrices composed of wild type FN (FN+/+) or of a mutant form of FN (FNΔ/+) from which the III13 heparin-binding site had been deleted by CRISPR-Cas 9 gene editing. The most significant effect of the mutant FN was a reduction in dendrite outgrowth. Not only were dendrites shorter on mutant FNΔ/+-collagen (COL) matrix than on wild type (FN+/+-COL) matrix, but the number of dendrites and dendritic spines per neuron and the spine densities were also dramatically reduced on FNΔ/+-COL matrices. Mass spectrometry and immunostaining identified a reduction in tenascin-C (TN-C) levels in the mutant matrix. TN-C is an ECM protein that binds to the III13 site of FN and modulates cell-matrix interactions and has been linked to dendrite development. We propose that TN-C binding to FN in the wound matrix supports dendrite and spine development during repair of damaged neural tissue. Overall, these results show that changes in ECM composition can dramatically affect elaboration of neurites and support the idea that the ECM microenvironment controls neuron morphology and connectivity.

Section: Methodsmentioning

confidence: 99%

Extracellular matrix composition affects outgrowth of dendrites and dendritic spines on cortical neurons

Sharma

Hill

2023

Front. Cell. Neurosci.

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“…J o u r n a l P r e -p r o o f 13 RNA extraction and qPCR RNA from mock-treated and EXT1 knockdown cells was isolated 2-5 days post-transfection with TRIzol reagent (Invitrogen) followed by clean-up with a RNeasy column (Qiagen) (38). cDNA was prepared from 1 μg of total RNA using Protoscript II Reverse Transcriptase (NEB, Ipswich, MA).…”

Section: Ext1 Sirna Transfection Of Nih 3t3 Cellsmentioning

confidence: 99%

Heparan sulfate is necessary for the early formation of nascent fibronectin and collagen I fibrils at matrix assembly sites

Hill

Lovett

Journal of Biological Chemistry

2022

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“…In fact, staining of the fibrotic subendothelial matrix in a diabetic retina not only shows extensive deposition of FN but also high levels of type IV collagen and tenascin-C, 21 two ECM proteins that increase in high glucose conditions. 22,42,43 Changes in composition can affect the ultrastructure and properties of the matrix. The ECM deposited by retinal endothelial cells cultured in high glucose is stiffer by atomic force microscopy 44 and also more permeable to fluorescent 43 kDa dextran 45 than in normal -Binds to specific integrins and to other BM proteins.…”

Section: Ecm Alterations In Drmentioning

confidence: 99%

“…The small isoform binds FN in the matrix where it is positioned to regulate FN interactions with cell receptors whereas the large isoform is excluded from the matrix thus potentially allowing uncontrolled FN accumulation. 43…”

Section: Cell Regulation Of Ecm Assemblymentioning

confidence: 99%

“…High glucose conditions increase FN1 transcription, but the change is modest and insufficient to explain the dramatic increase in matrix FN. 23,43,46 Because plasma FN transits from the bloodstream into tissues, 47 plasma FN provides a source of protein for assembly without changes in gene expression. Changes in intracellular pathways have also been linked to FN matrix assembly in high glucose.…”

Section: Mechanisms Of Ecm Modulation In Elevated Glucosementioning

confidence: 99%

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Implications of fibrotic extracellular matrix in diabetic retinopathy

Resnikoff

Miller

2022

Exp Biol Med (Maywood)

Fibrosis is an accumulation of extracellular matrix (ECM) proteins and fibers in a disordered fashion, which compromises cell and tissue functions. High glucose-induced fibrosis, a major pathophysiological change of diabetic retinopathy (DR), severely affects vision by compromising the retinal vasculature and ultimately disrupting retinal tissue organization. The retina is a highly vascularized, stratified tissue with multiple cell types organized into distinct layers. Chronically high blood glucose stimulates certain retinal cells to increase production and assembly of ECM proteins resulting in excess ECM deposition primarily in the capillary walls on the basal side of the endothelium. This subendothelial fibrosis of the capillaries is the earliest histological change in the diabetic retina and has been linked to the vascular dysfunction that underlies DR. Proteins that are not normally abundant in the capillary basement membrane (BM) matrix, such as the ECM protein fibronectin, are assembled in significant quantities, disrupting the architecture of the BM and altering its properties. Cell culture models have identified multiple mechanisms through which elevated glucose can stimulate fibronectin matrix assembly, including intracellular signaling pathways, alternative splicing, and non-enzymatic glycation of the ECM. The fibrotic subendothelial matrix alters cell adhesion and supports further accumulation of other ECM proteins leading to disruption of endothelial cell–cell junctions. We review evidence supporting the notion that these molecular changes in the ECM contribute to the pathogenesis of DR, including vascular leakage, loss of endothelial cells and pericytes, changes in blood flow, and neovascularization. We propose that the accumulation of ECM, especially fibronectin matrix, first around the vasculature and later in extravascular locations, plays a critical role in DR and vision loss. Strategies for DR prevention and treatment should consider the ECM a potential therapeutic target.