Silencing SMOC2 ameliorates kidney fibrosis by inhibiting fibroblast to myofibroblast transformation
Casimiro Gerarduzzi, Ramya K. Kumar, Priyanka Trivedi, Amrendra K. Ajay, Ashwin Iyer, Sarah Boswell, John N. Hutchinson, Sushrut S. Waikar, Vishal S. Vaidya
Casimiro Gerarduzzi, Ramya K. Kumar, Priyanka Trivedi, Amrendra K. Ajay, Ashwin Iyer, Sarah Boswell, John N. Hutchinson, Sushrut S. Waikar, Vishal S. Vaidya
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Research Article Nephrology

Silencing SMOC2 ameliorates kidney fibrosis by inhibiting fibroblast to myofibroblast transformation

Abstract

Secreted modular calcium-binding protein 2 (SMOC2) belongs to the secreted protein acidic and rich in cysteine (SPARC) family of matricellular proteins whose members are known to modulate cell-matrix interactions. We report that SMOC2 is upregulated in the kidney tubular epithelial cells of mice and humans following fibrosis. Using genetically manipulated mice with SMOC2 overexpression or knockdown, we show that SMOC2 is critically involved in the progression of kidney fibrosis. Mechanistically, we found that SMOC2 activates a fibroblast-to-myofibroblast transition (FMT) to stimulate stress fiber formation, proliferation, migration, and extracellular matrix production. Furthermore, we demonstrate that targeting SMOC2 by siRNA results in attenuation of TGFβ1-mediated FMT in vitro and an amelioration of kidney fibrosis in mice. These findings implicate that SMOC2 is a key signaling molecule in the pathological secretome of a damaged kidney and targeting SMOC2 offers a therapeutic strategy for inhibiting FMT-mediated kidney fibrosis — an unmet medical need.

Authors

Casimiro Gerarduzzi, Ramya K. Kumar, Priyanka Trivedi, Amrendra K. Ajay, Ashwin Iyer, Sarah Boswell, John N. Hutchinson, Sushrut S. Waikar, Vishal S. Vaidya

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Figure 2

SMOC2-overexpressing mice are more susceptible to kidney fibrosis than WT mice.

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SMOC2-overexpressing mice are more susceptible to kidney fibrosis than W...
(A) Confirmation of SMOC2 overexpression in SMOC2 transgenic (SMOC2 Tg) mice by PCR (above, primers specific to recognize Tg insert) and Western blotting (below) (Supplemental Figure 3A). (B) Representative Western blot (n = 5/condition; Supplemental Figure 3B and Supplemental Figure 5B) of αSMA, collagen 1α1, fibronectin, and SMOC2 expression using kidney samples obtained from SMOC2 Tg and WT mice subjected to 7 and 14 days of unilateral ureteral obstruction (UUO). (C) Representative images of immunofluorescent staining for αSMA in CoK and fibrotic kidneys from WT and SMOC2 Tg mice at day 7 following UUO (n = 5/condition, 5 visual fields/tissue). (D) Representative Western blot (n = 5/condition; Supplemental Figure 3C and Supplemental Figure 6B) of αSMA, collagen 1α1, fibronectin, and SMOC2 expression using kidney samples obtained from SMOC2 Tg and WT mice subjected to 7 and 14 days of folic acid (FA). (E) Representative images of immunofluorescent staining for αSMA of normal and fibrotic kidneys from WT and SMOC2 Tg mice at day 7 following FA (n = 5/condition, 10 visual fields/tissue). (F) Representative images of picrosirius red (n = 5/condition, 10 visual fields/tissue) and Masson’s trichrome (n = 5/condition, 5 visual fields/tissue) staining of CoK versus 7 and 14 day UUO–treated kidneys. (G) Representative images of picrosirius red and Masson’s trichrome staining of normal versus 7 and 14 day FA–treated kidneys (n = 5/condition, 5 visual fields/tissue). Confocal and light microscopy images are 20× magnification. Scale bars: 50 μM. Relative quantifications of images are represented as box plots, which describe the median (line within box), upper and lower quartiles (bounds of box), and minimum and maximum values (bars). *P < 0.05 (CoK [UUO] or Normal [FA]) and #P < 0.05 (WT at respective time point) determined by one-way ANOVA with Tukey post-hoc analysis.