• 1.

    Singh, N, DG Armstrong, and BA Lipsky. :Preventing foot ulcers in patients with diabetes. .JAMA 293::217. ,2005. .

  • 2.

    Jeffcoate, WJ . :Theories concerning the pathogenesis of the acute Charcot foot suggest future therapy. .Curr Diab Rep 5::430. ,2005. .

  • 3.

    Hampton, S . :Caring for the diabetic patient with a foot ulcer. .Br J Nurs 15::S22. ,2006. .

  • 4.

    Lobmann, R, A Ambrosch, G Schultz, et al. :Expression of matrix-metalloproteinases and their inhibitors in the wounds of diabetic and non-diabetic patients. .Diabetologia 45::1011. ,2002. .

    • Crossref
    • PubMed
    • Search Google Scholar
    • Export Citation
  • 5.

    Varani, J, RL Warner, M Gharaee-Kermani, et al. :Vitamin A antagonizes decreased cell growth and elevated collagen-degrading matrix metalloproteinase and stimulates collagen accumulation in naturally aged human skin. .J Invest Dermatol 114::480. ,2000. .

    • Crossref
    • PubMed
    • Search Google Scholar
    • Export Citation
  • 6.

    Visse, R and H Nagase. :Matrix metalloproteinases and tissue inhibitors of metalloproteinases: structure, function, and biochemistry. .Circ Res 92::827. ,2003. .

    • Crossref
    • PubMed
    • Search Google Scholar
    • Export Citation
  • 7.

    Iyer, V, K Pumiglia, and CM DiPersio. :α3β1 integrin regulates MMP-9 mRNA stability in immortalized keratinocytes: a novel mechanism of integrin-mediated MMP gene expression. .J Cell Sci 118 ( pt 6:):1185. ,2005. .

    • Crossref
    • PubMed
    • Search Google Scholar
    • Export Citation
  • 8.

    Das, S, M Mandal, T Chakraborti, et al. :Structure and evolutionary aspects of matrix metallo-proteinases: a brief overview. .Mol Cell Biochem 253::31. ,2003. .

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 9.

    Andreea, SI, C Marietta, and D Anca. :AGEs and glucose levels modulate type I and III procollagen mRNA synthesis in dermal fibroblasts cells culture. .Exp Diabetes Res. [Published online ahead of print April 3, 2008; 10.1155/2008/473603].

    • PubMed
    • Search Google Scholar
    • Export Citation
  • 10.

    Lobmann, R, T Pap, A Ambrosch, et al. :Differential effects of PDGF-BB on matrix metalloproteases and cytokine release in fibroblasts of Type 2 diabetic patients and normal controls in vitro. .J Diabetes Complications 20::105. ,2006. .

    • Crossref
    • PubMed
    • Search Google Scholar
    • Export Citation
  • 11.

    Lai, CF, V Seshadri, K Huang, et al. :An osteopontin–NADPH oxidase signaling cascade promotes pro–matrix metalloproteinase 9 activation in aortic mesenchymal cells. .Circ Res 98::1479. ,2006. .

    • Crossref
    • PubMed
    • Search Google Scholar
    • Export Citation
  • 12.

    Lee, CW, CC Lin, SF Luo, et al. :TNF-α induced MMP-9 expression via activation of Src/EGFR, PDGFR/PI3K/Akt cascade and promotion of NF-κB/p300 binding in human tracheal smooth muscle cells. .Am J Physiol Lung Cell Mol Physiol 292::L799. ,2007. .

    • Crossref
    • PubMed
    • Search Google Scholar
    • Export Citation
  • 13.

    Solini, A, E Santini, M Nannipieri, et al. :High glucose and homocysteine synergistically affect the metalloproteinases-tissue inhibitors of metalloproteinases pattern, but not TGFB expression, in human fibroblasts. .Diabetologia 49::2499. ,2006. .

    • Crossref
    • Search Google Scholar
    • Export Citation
  • 14.

    Fire, A, S Xu, MK Montgomery, et al. :Potent and specific genetic interference by double-stranded RNA in Caenorhabditis elegans. .Nature 391::806. ,1998. .

  • 15.

    Elbashir, SM, J Harborth, W Lendeckel, et al. :Duplexes of 21-nucleotide RNAs mediate RNA interference in cultured mammalian cells. .Nature 411::494. ,2001. .

  • 16.

    Ren, JL, JS Pan, T Cheng, et al. :RNA interference inhibits hepatitis B virus gene expression and replication in HepG2-N10 cells. .Chin J Dig Dis 7::230. ,2006. .

    • Crossref
    • PubMed
    • Search Google Scholar
    • Export Citation
  • 17.

    Armstrong, DG and EB Jude. :The role of matrix metalloproteinases in wound healing. .JAPMA 92::12. ,2002. .

  • 18.

    Reiss, MJ, YP Han, E Garcia, et al. :Matrix metalloproteinase-9 delays wound healing in a murine wound model. .Surgery 147::295. ,2010. .

  • 19.

    Burrow, JW, JA Koch, H Chuang, et al. :Nitric oxide donors selectively reduce the expression of matrix metalloproteinases-8 and -9 by human diabetic skin fibroblasts. .J Surg Res 140::90. ,2007. .

    • Crossref
    • PubMed
    • Search Google Scholar
    • Export Citation
  • 20.

    Harborth, J, SM Elbashir, K Bechert, et al. :Identification of essential genes in cultured mammalian cells using small interfering RNAs. .J Cell Sci 114::4557. ,2001. .

    • Crossref
    • PubMed
    • Search Google Scholar
    • Export Citation
  • 21.

    Sui, G, C Soohoo, EB Affar, et al. :A DNA vector-based RNAi technology to suppress gene expression in mammalian cells. .Proc Natl Acad Sci U S A 99::5515. ,2002. .

  • 22.

    Buysschaert, M, AS Dramais, PE Wallemacq, et al. :Hyperhomocysteinemia in type 2 diabetes: relation to macroangiopathy, nephropathy and insulin resistance. .Diabetes Care 23.:1846. ,2000. .

    • Search Google Scholar
    • Export Citation
  • 23.

    Fonseca, VA, A Stone, M Munshi, et al. :Oxidative stress in diabetic macrovascular disease: does homocysteine play a role. .South Med J 90::903. ,1997. .

  • 24.

    Boykin Jr, JV and C Baylis. :Homocysteine: a stealth mediator of impaired wound healing: a preliminary study. .Wounds 18::26. ,2006. .

  • 25.

    Death, AK, EJ Fisher, KC McGrath, et al. :High glucose alters matrix metalloproteinase expression in two key vascular cells: potential impact on atherosclerosis in diabetes. .Atherosclerosis 168::263. ,2003. .

    • Crossref
    • PubMed
    • Search Google Scholar
    • Export Citation
  • 26.

    Branski, LK, CT Pereira, DN Herndon, et al. :Gene therapy in wound healing: present status and future directions. .Gene Therapy 14::1. ,2007. .

Inhibition of Matrix Metalloproteinase 9 Expression in Rat Dermal Fibroblasts Using Small Interfering RNA

Xiao-Ying Xie Department of Endocrinology and Metabolism, The Second Affiliated Hospital of Sun Yat-sen University, Guangzhou, China.

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Chuan Yang Department of Endocrinology and Metabolism, The Second Affiliated Hospital of Sun Yat-sen University, Guangzhou, China.

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Meng Ren Department of Endocrinology and Metabolism, The Second Affiliated Hospital of Sun Yat-sen University, Guangzhou, China.

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Shao-Yun Hao Department of Endocrinology and Metabolism, The Second Affiliated Hospital of Sun Yat-sen University, Guangzhou, China.

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Ping Zhu Department of Endocrinology and Metabolism, The Second Affiliated Hospital of Sun Yat-sen University, Guangzhou, China.

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Li Yan Department of Endocrinology and Metabolism, The Second Affiliated Hospital of Sun Yat-sen University, Guangzhou, China.

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Background:

Matrix metalloproteinases (MMPs) degrade extracellular matrix components. Increased MMP-9 content in diabetic skin contributes to skin vulnerability and refractory foot ulcers. To identify ways to decrease MMP-9 levels in skin, inhibition of MMP-9 expression in dermal fibroblasts using small interfering RNA was investigated in vitro.

Methods:

A full-thickness wound was created on the midback of streptozotocin-induced diabetic rats; skin biopsies were performed 3 days later. Skin MMP-9 expression was observed by immunohistochemical analysis. Dermal fibroblasts from 1-day-old normal Sprague Dawley rats cultured with high glucose and homocysteine concentrations were transfected with small interfering RNA complexes. Cells were collected 30, 48, and 72 hours after transfection, and reverse transcription–polymerase chain reaction, Western blot analysis, and gelatin zymography for MMP-9 were performed.

Results:

Expression of MMP-9 was increased in diabetic rat skin, especially around wounds. After 30-, 48-, and 72-hour transfection with each MMP-9–specific small interfering RNA, reverse transcription–polymerase chain reaction showed markedly decreased MMP-9 messenger RNA expression, protein abundance, and activity. Of four MMP-9 small interfering RNAs, one sequence had a stable high inhibition rate (>70% at 30 and 48 hours after transfection).

Conclusions:

Expression of MMP-9 was increased in diabetic rat skin, especially around wounds, and was markedly inhibited after MMP-9 small interfering RNA transfection in vitro (P < .05). These findings may provide new treatments for diabetic skin wounds. (J Am Podiatr Med Assoc 102(4): 299–308, 2012)

Corresponding author: Li Yan, MM, Department of Endocrinology and Metabolism, The Second Affiliated Hospital of Sun Yat-sen University, 107 YanJiangXi Rd, Guangzhou 510120, China. (E-mail: ellen1130@163.com)