非线性光学显微成像技术诊断移植肾纤维化的临床研究

Diagnosis of interstitial fibrosis of renal allograft by non-linear optical imaging technology

目的探索使用非线性光学显微成像技术对移植肾间质纤维化进行成像及定量分析的效果。方法分析2005年1月至2013年3月间40例肾移植受者的移植。肾组织样本，其中轻度纤维化22例，中重度纤维化18例，分别行双光子激发荧光（TPEF）和二次谐波（SHG）成像以及Masson染色成像，通过肉眼观察和Image-Pro—Plus软件初步分析比较两种方法对同一区域纤维化的显像情况。结果对于轻度纤维化组肾组织样本，SHG和TPEF网像可以清晰地显示在肾小球、肾小血管、肾小管周围及肾间质内有少量散在绿色信号的胶原纤维沉积，而Masson染色则显示基本正常，未能发现早期轻度的纤维化表现。对于中重度纤维化组肾组织样本，SHG和TPEF成像则能清晰显示正常胶原的轮廓和结构，并通过计算机程序对其进行较准确的定量分析。40例SHG和TPEF图像的纤维化指数为（27．4±15．8）％，Masson染色图像纤维化指数为（26．8±16．0）％，二者间的差异无统计学意义（z=-0．72，P=0．94）。结论与Masson染色相比较，SHG和TPEF技术对胶原纤维成像更敏感、清晰，能够更直观、早期的发现移植肾问质纤维化，是一种有研究潜力的新型诊断方法。

Objective To figure out the image quality of renal allograft interstitial fibrosis by non-linear optical imaging technology and compare with the traditional pathological staining method to confirm its advantages. Method Forty cases of paraffin-embedded renal allograft biopsy specimens who were diagnosed as mild （n = 22） to moderate-severe （n = 18） renal interstitial fibrosis （RIF） were obtained from Jan. 2005 to Mar. 2013, and subjected to second harmonic generation and two photon fluorescence （SHG/TPEF） imaging and Masson staining. The findings of the fibrosis at the same region by the above methods were compared and analyzed by the naked eyes and Image-Pro-Plus software. Result SHG/TPEF images could clearly display ,the construction of renal tissue as traditional Masson staining. In mild renal allograft interstitial fibrosis specimens, SHG/TPEF images could reveal a few＇interspersed green collagen deposit around glomerulus, renal tubule, renal vessel, and in interstitium, but the pathologic results of Masson staining were nearly normal by the naked eyes. For moderate-severe renal allograft interstitial fibrosis specimens, SHG/TPEF images could more easily show the distribution range and deposit position of green collagen than the Masson staining. There was no significant difference in the firosis index between the SHG/TPEF imaging and Masson staining （Z = - 0. 72, P = 0. 94）. Conclusion Non-linear optical technology can clearly and early display renal allograft interstitial fibrosis and tissue structure. SHG/TPEF is a better method than traditional staining method for accessing renal allograft interstitial fibrosis.