双光子成像技术在大鼠放射性皮肤损伤修复评估中的应用

Application of two-photon imaging technology in the repair evaluation of radiation-induced skin injury in rats

目的采用双光子激发荧光(TPEF)成像技术,无创、活体评估X射线引起大鼠皮肤放射性损伤发展和修复过程。方法24只SD大鼠采用随机数表法分为4组,健康对照组、25 Gy组、35 Gy组和45 Gy组,每组6只。照射后不同时间评估皮肤损伤程度,通过TPEF成像技术在体检测表皮细胞尼克酰胺腺嘌呤二核苷酸(磷酸)[NAD(P)H]和真皮胶原纤维荧光信号的病理改变。结果第10天,各辐射组大鼠出现红斑和脱皮;第15~20天,随着辐射剂量地增加,辐射组出现递进性的渗出、水肿和溃疡;第25天,25 Gy组开始修复,其他组仍有渗出和溃疡。第10天,25、35和45 Gy组表皮细胞NAD(P)H荧光信号减少,乳头层和网状层荧光信号值减少,与健康对照组比较,差异有统计学意义(t=24.145、28.303、26.989、6.654、7.510、7.997,P<0.05);第30天,25 Gy组表皮细胞NAD(P)H和真皮胶原纤维开始修复,颗粒层、棘层和基底层细胞出现荧光信号,35、45 Gy组未出现NAD(P)H荧光信号;25 Gy组乳头层和网状层荧光信号均逐渐增高,但仍低于健康对照组(t=115.133、17.431,P<0.05),而45 Gy组未出现荧光信号。结论TPEF技术可以无创、活体检测X射线照射后表皮细胞和真皮胶原纤维信号损伤和修复的病理变化。

Objective To evaluate the skin development and repair process of X-ray radiation damage in rat with non-invasive two-photon excitation fluorescence(TPEF)imaging technology in vivo.Methods Totally 24 SD rats were randomly divided into four groups including X-ray irradiated group(25,35 and 45 Gy)and non-irradiation control group.At different times after irradiation,the degree of skin injury was evaluated,and the pathological changes of nicotinamide adenine dinucleotide(phosphate)[NAD(P)H]and collagen fiber fluorescence signals in epidermal cells were detected in vivo by TPEF imaging technology.Results At 10 d post-irradiation,the skin of irradiation groups showed erythema and desquamation.At 15-20 d post-irradiation,the skin of radiation groups developed progressive exudation,edema and ulcers with increasing radiation dose.On day 25,the skin began to repair in the 25 Gy group,however,the skin of other groups still had exudation and ulcers.On day 10,NAD(P)H fluorescence signal in epidermal cells of irradiation groups decreased and the fluorescence signal of collagen fibers in papillary layer and reticular layer of irradiation groups reduced,which were significantly lower than that of normal control group(t=24.145,28.303,26.989,6.654,7.510,7.997,P<0.05).On day 30,fluorescence signal of NAD(P)H and collagen fibers in epidermal cells and dermis began to repair,the cell from stratum granulosum,stratum spinosum,and stratum basale in the 25 Gy group showed fluorescence signal,the other groups did not show.The fluorescence signal of collagen fibers in the 25 Gy group were gradually increased in papillary layer and reticular layer,however,they were significantly lower than normal control group(t=115.133,17.431,P<0.05),the skin of 45 Gy group did not show fluorescence signal of collagen fibers.Conclusions The damage and repair process of epidermal cells and dermal collagen fiber can be detected noninvasively by TPEF imaging technology after X-ray irradiation in vivo.