干预Periostin-TGF-beta环路逆转大鼠移植肾小管间质纤维化的进展

Chronic renal allograft damage remains a main cause of late graft loss. It is characterized by progressive tubulointerstitial fibrosis, which finally develops into irreversible graft dysfunction.In our previous studies on a rat model of F344-Lewis kidney transplantation, we found that upregulation of TGF-beta in the graft is closely associated with the progression of nephropathy. Thus far studies have focused on profibrotic mechanisms of TGF-beta. However,it is unclear how the expression and activity of TGF-beta is maintained at high level in tissues undergoing fibrosis, which might drive the progression of renal allograft dyfunction. Recent studies indicated that periostin may serve as a marker of the reversibility of chronic renal disease. Periostin is a secreted matricellular protein and acts as a signal molecule as well. It positively cross-regulated with TGF-beta in the profibrotic process. We hypothesized that the high level of periostin induced in graft is involved in sustained upregulation of TGF-beta signaling in profibrotic processes and leads to progression of chronic renal allograft tubulointersitial injuries. This project is to investigate the formation and modulation of periostin-TGF-beta circuit in rat renal allogtafts and its implication in the non-return tubulointerstitial injuries，and to explore the potential therapeutics to rescue renal allografts from late loss.

以肾小管间质进行性损伤和纤维化为特征的移植物肾病进展至不可逆转是移植肾失功的重要原因。我们在F344-Lewis大鼠肾移植模型的研究发现TGF-beta高表达与病变的进展密切相关。大量研究证实TGF-beta在组织纤维化过程中起重要作用，TGF-beta持续高活性可能是移植肾小管间质纤维化进展的推动力，然而维持TGF-beta高活性的机制尚未阐明。最新的研究提示periostin上调与肾脏慢性病变不可逆转有关。periostin是一种分泌性间质信号蛋白，由TGF-beta诱导生成，与TGF-beta共同促进组织纤维化并互为正性调控，参与TGF-beta高活性的维持。本项目研究大鼠移植肾Periostin-TGF-beta环路的形成及其对小管间质损伤逆转性的调控机制，探讨通过干预Periostin-TGF-beta环路挽救移植肾失功的新途径。

近年来的研究表明，periostin是慢性肾脏病肾脏纤维化进展的重要标志物，其肾脏组织表达水平增高预示病变进展的不可逆性。然而，periostin在肾脏纤维化进展中的作用机制尚未阐明。本项目拟探索periostin在肾脏纤维化过程中的变化以及作为干预靶点对治疗移植物肾小管间质纤维化不可逆进展的潜在意义。本项目在前期研究发现大鼠同种异体移植肾脏内TGF-beta1高水平表达的基础上，通过体外实验证明大鼠肾小管上皮细胞在TGF-beta1作用下产生periostin，同时在TGF-beta1作用下的肾小管上皮细胞自身能够上调TGF-beta1的表达水平和分泌，从而放大TGF-beta1-periostin效应，构成推动纤维化病变进展的分子基础。通过沉默periostin，我们发现periostin基因敲低可以使肾小管上皮细胞Vimentin, PAI-1蛋白表达减少，抑制TGF-beta1诱导的alpha-SMA和Col1A的mRNA上调，有助于减轻肾脏纤维化；然而，我们也发现，periostin基因敲低也导致Fibronectin，FSP-1的mRNA水平增高，对纤维化可能产生促进作用。periostin基因敲低的肾小管上皮细胞活力与生长受到明显抑制。此外，我们还发现TGF-beta1对肾小管上皮细胞单核细胞趋化因子MCP-1具有很强的诱导作用，伴有BAMBI表达水平下调。MCP-1也是促进肾脏纤维化进展的关键分子，而BAMBI是假性I型TGF-beta受体，在细胞中拮抗TGF-beta信号通路的过度激活。我们发现TGF-beta1可能通过激活MEK/ERK1/2抑制BAMBI的表达，而BAMBI的下调则通过增强TGF-beta1-MEK/ERK1/2信号转导，进一步促进TGF-beta1对MCP-1的诱导作用，形成一条新的促纤维化正性环路。我们的研究结果提示，肾脏组织 periostin的过度产生可能作为肾脏纤维化进展的标志物，但它们正常水平的表达也有其功能学意义。肾脏纤维化的不可逆进展可能与多种促纤维化分子环路的存在有关，单纯阻断一个靶分子可能不足以逆转纤维化的进程，未来的研究方向应探讨多环节联合干预。