αVβ8 integrin targeting to prevent posterior capsular opacification
Mahbubul H. Shihan, … , Adam P. Faranda, Melinda K. Duncan
Mahbubul H. Shihan, … , Adam P. Faranda, Melinda K. Duncan
Published September 23, 2021
Citation Information: JCI Insight. 2021;6(21):e145715. https://doi.org/10.1172/jci.insight.145715.
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Research Article Cell biology Ophthalmology

αVβ8 integrin targeting to prevent posterior capsular opacification

Abstract

Fibrotic posterior capsular opacification (PCO), a major complication of cataract surgery, is driven by transforming growth factor–β (TGF-β). Previously, αV integrins were found to be critical for the onset of TGF-β–mediated PCO in vivo; however, the functional heterodimer was unknown. Here, β8 integrin–conditional knockout (β8ITG-cKO) lens epithelial cells (LCs) attenuated their fibrotic responses, while both β5 and β6 integrin–null LCs underwent fibrotic changes similar to WT at 5 days post cataract surgery (PCS). RNA-Seq revealed that β8ITG-cKO LCs attenuated their upregulation of integrins and their ligands, as well as known targets of TGF-β–induced signaling, at 24 hours PCS. Treatment of β8ITG-cKO eyes with active TGF-β1 at the time of surgery rescued the fibrotic response. Treatment of WT mice with an anti-αVβ8 integrin function blocking antibody at the time of surgery ameliorated both canonical TGF-β signaling and LC fibrotic response PCS, and treatment at 5 days PCS, after surgically induced fibrotic responses were established, largely reversed this fibrotic response. These data suggest that αVβ8 integrin is a major regulator of TGF-β activation by LCs PCS and that therapeutics targeting αVβ8 integrin could be effective for fibrotic PCO prevention and treatment.

Authors

Mahbubul H. Shihan, Samuel G. Novo, Yan Wang, Dean Sheppard, Amha Atakilit, Thomas D. Arnold, Nicole M. Rossi, Adam P. Faranda, Melinda K. Duncan

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Figure 5

LCs lacking the β8 integrin gene fail to activate TGF-β signaling PCS, but this can be rescued by treatment with active TGF-β1.

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LCs lacking the β8 integrin gene fail to activate TGF-β signaling PCS, b...
(A) Heatmap of genes known to participate in TGF-β pathways expressed at lower levels in β8ITG-cKO LCs at 24 hours PCS compared with WT. Expression levels are reported as fragments per kilobase million (FPKM). (B and C) SMAD3 phosphorylation (p-SMAD3) is detected in WT LCs by 48 hours PCS (P = 0.013), and this upregulates further by 5 days PCS (P ≤ 0.001; P ≤ 0.002). In contrast, β8ITG-cKO LCs exhibit attenuated SMAD3 phosphorylation 48 hours (P = 0.042) and 5 days PCS (P ≤ 0.001). (D–I) Treatment of β8ITG-cKO capsular bags with active TGF-β1 at the time of lens fiber cell removal rescued both p-SMAD3 levels (D and E; P ≤ 0.001), and the robust expression of α-SMA (D and F; P = 0.011), tenascin C (D and G; P = 0.007), fibronectin (D and H; P = 0.012) and collagen I (D and I; P = 0.003) 5 days PCS. Scale bar: 35 μm. p-SMAD3, tenascin C, fibronectin, and collagen I (red); α-SMA (green); DNA detected by Draq5 (blue). All experiments had n = 3. Values are expressed as mean ± SEM presented for 1 representative experiment of 2 independent experiments, with similar results; asterisks indicate statistically significant MFI between WT and/or β8ITG-cKO and/or β8ITG-cKO (TGF-β) (*P ≤ 0.05; **P ≤ 0.01; ***P ≤ 0.001); both Student’s 2-tailed t test (corrected for multiple comparisons using the Holm-Šídák method) and 1-way ANOVA with Tukey’s post hoc test (C) or 1-way ANOVA with Tukey’s post hoc test (E–I). Graph colors: (C) red (different PCS time points of WT), green (different PCS time points of β8ITG-cKO); (E–I) red (WT), green (β8ITG-cKO), blue (β8ITG-cKO [TGF-β]. C, lens capsule; LC, remnant lens cells; PCS, post cataract surgery; a, anterior; p, posterior.