Gene expression analysis of the pleiotropic effects of TGF-β1 in an in vitro model of flexor tendon healing.
Gene expression analysis of the pleiotropic effects of TGF-β1 in an in vitro model of flexor tendon healing.
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DOI：
10.1371/journal.pone.0051411
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发表时间：
2012
期刊：
影响因子：
3.7
通讯作者：
Awad HA
中科院分区：
文献类型：
--
作者：
Farhat YM;Al-Maliki AA;Chen T;Juneja SC;Schwarz EM;O'Keefe RJ;Awad HA
屈肌腱损伤是手外科医生和组织工程师面临的最具挑战性的问题之一。屈肌腱损伤不仅愈合时机械强度差，而且还可能形成使人衰弱的粘连，从而可能永久性地损害手的功能。虽然TGF-β1是恢复肌腱强度的必要因素，但它与屈肌腱和其他组织中的瘢痕和粘连形成以及纤维化疾病有关。TGF-β1对肌腱细胞和组织的多效性作用尚未详细描述。本研究的目的是确定TGF-β1对肌腱愈合的影响可以通过哪些靶点来改变。为了实现这一点，我们用1、10或100 ng/mL的TGF-β1处理在固定的胶原凝胶中培养的屈肌腱腱细胞，并在处理后48小时内使用RT-PCR测量凝胶收缩和基因表达。具体而言，我们研究了TGF-β1对胶原、纤连蛋白、蛋白多糖、MMP、MMP抑制剂以及新腱转录因子Scleraxis和Mohawk表达的影响。凝胶的面积收缩与测试的TGF-β1浓度无剂量依赖性。我们观察到MMP-16（MT3-MMP）和核心蛋白聚糖的剂量依赖性下调，以及双糖链蛋白聚糖、胶原V、胶原XII、派-1、巩膜轴蛋白和Mohawk的TGF-β1上调。还进行了基因间分析以进一步表征在肌腱细胞接种的胶原凝胶中ECM和MMP基因的表达。这些分析表明，TGF-β1使基因表达的平衡倾向于ECM合成，而不是基质重塑MMPs，这可能是TGF-β1促进粘附形成的一种方式。
Flexor tendon injuries are among the most challenging problems for hand surgeons and tissue engineers alike. Not only do flexor tendon injuries heal with poor mechanical strength, they can also form debilitating adhesions that may permanently impair hand function. While TGF-β1 is a necessary factor for regaining tendon strength, it is associated with scar and adhesion formation in the flexor tendons and other tissues as well as fibrotic diseases. The pleiotropic effects of TGF-β1 on tendon cells and tissue have not been characterized in detail. The goal of the present study was to identify the targets through which the effects of TGF-β1 on tendon healing could be altered. To accomplish this, we treated flexor tendon tenocytes cultured in pinned collagen gels with 1, 10 or 100 ng/mL of TGF-β1 and measured gel contraction and gene expression using RT-PCR up to 48 hours after treatment. Specifically, we studied the effects of TGF-β1 on the expression of collagens, fibronectin, proteoglycans, MMPs, MMP inhibitors, and the neotendon transcription factors, Scleraxis and Mohawk. Area contraction of the gels was not dose-dependent with the TGF-β1 concentrations tested. We observed dose-dependent downregulation of MMP-16 (MT3-MMP) and decorin, and upregulation of biglycan, collagen V, collagen XII, PAI-1, Scleraxis, and Mohawk by TGF-β1. Inter-gene analyses were also performed to further characterize the expression of ECM and MMP genes in the tenocyte-seeded collagen gels. These analyses illustrate that TGF-β1 tilts the balance of gene expression in favor of ECM synthesis rather than the matrix-remodeling MMPs, a possible means by which TGF-β1 promotes adhesion formation.
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