Multiphoton Images of fibrosis in vitreoretinal diseases using a SHG microscope.

多光子显微镜（MPM）可以利用内源性二次谐波（SHG）对组织和细胞化胶原凝胶进行成像，提供胶原含量和微观结构的独特信息。我们使用SHG显微镜观察视网膜前纤维血管膜、患者的内限制膜和视网膜色素上皮（RPE）培养的胶原凝胶。从增生性玻璃体视网膜病变（PVR）、增生性糖尿病视网膜病变（PDR）和特发性视网膜前膜（iERM）患者的视网膜前组织中检测SHG信号。我们用面积分数和平均强度来评估SHG信号。眼底照片显示PDR患者纤维血管膜平均SHG信号强度随纤维化进展有增加的趋势。在3D培养的胶原凝胶模型中，TGF β 2刺激较非刺激组显著增加SHG面积分数和平均SHG信号强度。由于我们无需任何固定或染色即可评估SHG信号变化，因此这种细胞化胶原凝胶模型可能有助于实时评估纤维化。

Multiphoton microscopy(MPM) can image tissue and cellularized collagen gels using endogenous second-harmonic generation(SHG) to provide unique information about collagen content and microstructure. We observed preretinal fibrovascular membranes, inner limiting membrane obtained from patients and retinal pigment epithelium(RPE) cultured collagen gels using a SHG microscope. SHG signals were detected from preretinal tissue obtained from patients with proliferative vitreoretinopathy(PVR), proliferative diabetic retinopathy(PDR) and idiopathic epiretinal membrane(iERM). We evaluated SHG signals as the area fraction and mean intensity. Mean SHG signal intensity of fibrovasucular membranes obtained from PDR patients had a trend to increase in the progress of fibrosis based on the fundus photographs. In 3D cultured collagen gel model, TGF beta2 stimulation significantly increased both SHG area fraction and mean SHG signal intensity compared to non-stimulant control. Since we can evaluate SHG signal changes without any fixation or staining, this cellularized collagen gel model may be useful to evaluate fibrosis in real time.