Matteini Paolo, Rossi Francesca, Menabuoni Luca, Pini Roberto
Consiglio Nazionale delle Ricerche, Istituto di Fisica Applicata Nello Carrara, Sesto Fiorentino, Italy.
Lasers Surg Med. 2007 Aug;39(7):597-604. doi: 10.1002/lsm.20532.
Laser welding of corneal tissue that employs diode lasers (810 nm) at low power densities (12-20 W/cm(2)) in association with Indocyanine Green staining of the wound is a technique proposed as an alternative to conventional suturing procedures. The aim of this study is to evaluate, by means of light (LM) and transmission electron microscopy (TEM) analyses, the structural modifications induced in laser-welded corneal stroma.
Experiments were carried out in 20 freshly enucleated pig eyes. A 3.5 mm in length full-thickness cut was produced in the cornea, and was then closed by laser welding. Birefringence modifications in samples stained with picrosirius red dye were analyzed by polarized LM to assess heat damage. TEM analysis was performed on ultra-thin slices, contrasted with uranyl acetate and lead citrate, in order to assess organization and size of type I collagen fibrils after laser welding.
LM evidenced bridges of collagen bundles between the wound edges, with a loss of regular lamellar organization at the welded site. Polarized LM indicated that birefringence properties were mostly preserved after laser treatment. TEM examinations revealed the presence of quasi-ordered groups of fibrils across the wound edges preserving their interfibrillar spacing. These fibrils appeared morphologically comparable to those in the control tissue, indicating that type I collagen was not denatured during the diode laser corneal welding.
The preservation of substantially intact, undenatured collagen fibrils in laser-welded corneal wounds supported the thermodynamic studies that we carried out recently, which indicated temperatures below 66 degrees C at the weld site under laser irradiation. This observation enabled us to hypothesize that the mechanism, proposed in the literature, of unwinding of collagen triple helixes followed by fibrils "interdigitation" is not likely to occur in the welding process that we set up for the corneal suturing.
利用二极管激光器（810纳米）在低功率密度（12 - 20瓦/平方厘米）下对角膜组织进行激光焊接，并结合伤口的吲哚菁绿染色，是一种被提议作为传统缝合手术替代方法的技术。本研究的目的是通过光学显微镜（LM）和透射电子显微镜（TEM）分析，评估激光焊接角膜基质中诱导的结构改变。
在20只新鲜摘除的猪眼中进行实验。在角膜上制作一个3.5毫米长的全层切口，然后通过激光焊接闭合。用天狼星红染料染色的样本中的双折射改变通过偏振光LM分析以评估热损伤。对超薄切片进行TEM分析，用醋酸铀和柠檬酸铅进行对比，以评估激光焊接后I型胶原纤维的组织结构和大小。
LM显示伤口边缘之间有胶原束桥接，焊接部位的规则板层组织丧失。偏振光LM表明激光处理后双折射特性大多得以保留。TEM检查显示伤口边缘有准有序的纤维组，其纤维间间距得以保留。这些纤维在形态上与对照组织中的纤维相似，表明在二极管激光角膜焊接过程中I型胶原未变性。
激光焊接的角膜伤口中基本完整、未变性的胶原纤维得以保留，这支持了我们最近进行的热力学研究，该研究表明激光照射下焊接部位温度低于66摄氏度。这一观察结果使我们能够假设，文献中提出的胶原三螺旋解旋然后纤维“交叉指状”的机制在我们为角膜缝合建立的焊接过程中不太可能发生。
