Transforming growth factor-beta 1 produced by vascular smooth muscle cells predicts fibrosis in the gastrocnemius of patients with peripheral artery disease

Duy M. Ha, Lauren C. Carpenter, Panagiotis Koutakis, Stanley A. Swanson, Zhen Zhu, Mina Hanna, Holly K. DeSpiegelaere, Iraklis I Pipinos, George P Casale

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Abstract

Background: Lower leg ischemia, myopathy, and limb dysfunction are distinguishing features of peripheral artery disease (PAD). The myopathy of PAD is characterized by myofiber degeneration in association with extracellular matrix expansion, and increased expression of transforming growth factor-beta 1 (TGF-β1; a pro-fibrotic cytokine). In this study, we evaluated cellular expression of TGF-β1 in gastrocnemius of control (CTRL) and PAD patients and its relationship to deposited collagen, fibroblast accumulation and limb hemodynamics. Methods: Gastrocnemius biopsies were collected from PAD patients with claudication (PAD-II; N=25) and tissue loss (PAD-IV; N=20) and from CTRL patients (N=20). TGF-β1 in slide-mounted specimens was labeled with fluorescent antibodies and analyzed by quantitative wide-field, fluorescence microscopy. We evaluated co-localization of TGF-β1 with vascular smooth muscle cells (SMC) (high molecular weight caldesmon), fibroblasts (TE-7 antigen), macrophages (CD163), T cells (CD3) and endothelial cells (CD31). Collagen was stained with Masson Trichrome and collagen density was determined by quantitative bright-field microscopy with multi-spectral imaging. Results: Collagen density increased from CTRL to PAD-II to PAD-IV specimens (all differences p<0.05) and was prominent around microvessels. TGF-β1 expression increased with advancing disease (all differences p<0.05), correlated with collagen density across all specimens (r=0.864; p<0.001), associated with fibroblast accumulation, and was observed exclusively in SMC. TGF-β1 expression inversely correlated with ankle-brachial index across PAD patients (r=-0.698; p<0.001). Conclusions: Our findings support a progressive fibrosis in the gastrocnemius of PAD patients that is caused by elevated TGF-β1 production in the SMC of microvessels in response to tissue hypoxia.

Original languageEnglish (US)
Article number39
JournalJournal of Translational Medicine
Volume14
Issue number1
DOIs
StatePublished - Feb 4 2016

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Peripheral Arterial Disease
Vascular Smooth Muscle
Transforming Growth Factor beta
Smooth Muscle Myocytes
Muscle
Fibrosis
Cells
Collagen
Fibroblasts
Muscular Diseases
Microvessels
Extremities
Calmodulin-Binding Proteins
Ankle Brachial Index
Tissue
T-cells
Biopsy
Macrophages
Fluorescence microscopy
Endothelial cells

Keywords

  • Fibrosis
  • Microvasculature
  • Peripheral artery disease
  • Skeletal muscle
  • Transforming growth factor-beta 1
  • Vascular smooth muscle cells

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)

Cite this

Transforming growth factor-beta 1 produced by vascular smooth muscle cells predicts fibrosis in the gastrocnemius of patients with peripheral artery disease. / Ha, Duy M.; Carpenter, Lauren C.; Koutakis, Panagiotis; Swanson, Stanley A.; Zhu, Zhen; Hanna, Mina; DeSpiegelaere, Holly K.; Pipinos, Iraklis I; Casale, George P.

In: Journal of Translational Medicine, Vol. 14, No. 1, 39, 04.02.2016.

Research output: Contribution to journalArticle

Ha, Duy M. ; Carpenter, Lauren C. ; Koutakis, Panagiotis ; Swanson, Stanley A. ; Zhu, Zhen ; Hanna, Mina ; DeSpiegelaere, Holly K. ; Pipinos, Iraklis I ; Casale, George P. / Transforming growth factor-beta 1 produced by vascular smooth muscle cells predicts fibrosis in the gastrocnemius of patients with peripheral artery disease. In: Journal of Translational Medicine. 2016 ; Vol. 14, No. 1.
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abstract = "Background: Lower leg ischemia, myopathy, and limb dysfunction are distinguishing features of peripheral artery disease (PAD). The myopathy of PAD is characterized by myofiber degeneration in association with extracellular matrix expansion, and increased expression of transforming growth factor-beta 1 (TGF-β1; a pro-fibrotic cytokine). In this study, we evaluated cellular expression of TGF-β1 in gastrocnemius of control (CTRL) and PAD patients and its relationship to deposited collagen, fibroblast accumulation and limb hemodynamics. Methods: Gastrocnemius biopsies were collected from PAD patients with claudication (PAD-II; N=25) and tissue loss (PAD-IV; N=20) and from CTRL patients (N=20). TGF-β1 in slide-mounted specimens was labeled with fluorescent antibodies and analyzed by quantitative wide-field, fluorescence microscopy. We evaluated co-localization of TGF-β1 with vascular smooth muscle cells (SMC) (high molecular weight caldesmon), fibroblasts (TE-7 antigen), macrophages (CD163), T cells (CD3) and endothelial cells (CD31). Collagen was stained with Masson Trichrome and collagen density was determined by quantitative bright-field microscopy with multi-spectral imaging. Results: Collagen density increased from CTRL to PAD-II to PAD-IV specimens (all differences p<0.05) and was prominent around microvessels. TGF-β1 expression increased with advancing disease (all differences p<0.05), correlated with collagen density across all specimens (r=0.864; p<0.001), associated with fibroblast accumulation, and was observed exclusively in SMC. TGF-β1 expression inversely correlated with ankle-brachial index across PAD patients (r=-0.698; p<0.001). Conclusions: Our findings support a progressive fibrosis in the gastrocnemius of PAD patients that is caused by elevated TGF-β1 production in the SMC of microvessels in response to tissue hypoxia.",
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AU - Ha, Duy M.

AU - Carpenter, Lauren C.

AU - Koutakis, Panagiotis

AU - Swanson, Stanley A.

AU - Zhu, Zhen

AU - Hanna, Mina

AU - DeSpiegelaere, Holly K.

AU - Pipinos, Iraklis I

AU - Casale, George P

PY - 2016/2/4

Y1 - 2016/2/4

N2 - Background: Lower leg ischemia, myopathy, and limb dysfunction are distinguishing features of peripheral artery disease (PAD). The myopathy of PAD is characterized by myofiber degeneration in association with extracellular matrix expansion, and increased expression of transforming growth factor-beta 1 (TGF-β1; a pro-fibrotic cytokine). In this study, we evaluated cellular expression of TGF-β1 in gastrocnemius of control (CTRL) and PAD patients and its relationship to deposited collagen, fibroblast accumulation and limb hemodynamics. Methods: Gastrocnemius biopsies were collected from PAD patients with claudication (PAD-II; N=25) and tissue loss (PAD-IV; N=20) and from CTRL patients (N=20). TGF-β1 in slide-mounted specimens was labeled with fluorescent antibodies and analyzed by quantitative wide-field, fluorescence microscopy. We evaluated co-localization of TGF-β1 with vascular smooth muscle cells (SMC) (high molecular weight caldesmon), fibroblasts (TE-7 antigen), macrophages (CD163), T cells (CD3) and endothelial cells (CD31). Collagen was stained with Masson Trichrome and collagen density was determined by quantitative bright-field microscopy with multi-spectral imaging. Results: Collagen density increased from CTRL to PAD-II to PAD-IV specimens (all differences p<0.05) and was prominent around microvessels. TGF-β1 expression increased with advancing disease (all differences p<0.05), correlated with collagen density across all specimens (r=0.864; p<0.001), associated with fibroblast accumulation, and was observed exclusively in SMC. TGF-β1 expression inversely correlated with ankle-brachial index across PAD patients (r=-0.698; p<0.001). Conclusions: Our findings support a progressive fibrosis in the gastrocnemius of PAD patients that is caused by elevated TGF-β1 production in the SMC of microvessels in response to tissue hypoxia.

AB - Background: Lower leg ischemia, myopathy, and limb dysfunction are distinguishing features of peripheral artery disease (PAD). The myopathy of PAD is characterized by myofiber degeneration in association with extracellular matrix expansion, and increased expression of transforming growth factor-beta 1 (TGF-β1; a pro-fibrotic cytokine). In this study, we evaluated cellular expression of TGF-β1 in gastrocnemius of control (CTRL) and PAD patients and its relationship to deposited collagen, fibroblast accumulation and limb hemodynamics. Methods: Gastrocnemius biopsies were collected from PAD patients with claudication (PAD-II; N=25) and tissue loss (PAD-IV; N=20) and from CTRL patients (N=20). TGF-β1 in slide-mounted specimens was labeled with fluorescent antibodies and analyzed by quantitative wide-field, fluorescence microscopy. We evaluated co-localization of TGF-β1 with vascular smooth muscle cells (SMC) (high molecular weight caldesmon), fibroblasts (TE-7 antigen), macrophages (CD163), T cells (CD3) and endothelial cells (CD31). Collagen was stained with Masson Trichrome and collagen density was determined by quantitative bright-field microscopy with multi-spectral imaging. Results: Collagen density increased from CTRL to PAD-II to PAD-IV specimens (all differences p<0.05) and was prominent around microvessels. TGF-β1 expression increased with advancing disease (all differences p<0.05), correlated with collagen density across all specimens (r=0.864; p<0.001), associated with fibroblast accumulation, and was observed exclusively in SMC. TGF-β1 expression inversely correlated with ankle-brachial index across PAD patients (r=-0.698; p<0.001). Conclusions: Our findings support a progressive fibrosis in the gastrocnemius of PAD patients that is caused by elevated TGF-β1 production in the SMC of microvessels in response to tissue hypoxia.

KW - Fibrosis

KW - Microvasculature

KW - Peripheral artery disease

KW - Skeletal muscle

KW - Transforming growth factor-beta 1

KW - Vascular smooth muscle cells

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