Objective To explore the mechanism of recombinant thymosin β4 (Tβ4) accelerating skin wound heal ing in rats by regulating laminin 5 expression. Methods Two full thickness 8 mm punch wounds were made at the costovertebralangle on dorsal surface of each adult male rats weighing 200-250 g. Sixty rats were randomized into the control group (n=15) and the experimental group (n=45), which was subdivided into low, medium and high dose groups (n=15). Tβ4 was appl ied topically at 2, 6, 18 μg in 50 μL PBS for every 12 hours after model making in the experimental group. The identical amounts of phosphate buffered sal ine was appl ied in the control group. Wound heal ing was observed after model making and immunohistochemical observation was conducted 2, 4 and 7 days after operation. Results Seven days after operation, wound contracted obviously and most of the wounds connected well with the margin. In the control group, low dose group, medium dose group and high dose group, the wound heal ing rate were 7.67% ± 5.46%, 29.01% ± 7.43%, 26.54% ± 11.49% and 10.39% ± 3.96% respectively 2 days after operation; 28.16% ± 13.76%, 37.99% ± 13.05%, 42.00% ± 9.56% and 39.58% ± 12.74% respectively 4 days after operation; 59.08% ± 19.02%, 64.15% ± 17.92%, 77.39% ± 8.45% and 69.78% ± 8.45% respectively 7 days after operation. At 2 days after operation, significant differences were notified in heal ing rats between 3 sub-experimental groups and the control group (P lt; 0.05). Immunohistochemistry staining showed that there was a l ittle more positive expression of laminin 5 2 days after operation that beneficial to promote the prol iferation and differentiation of cell in every group, including positive cells andECM. But in medium group there was fewer expression, only at the borderl ine and bottom of the wound, while the expression significantly increased 4 days after operation (P lt; 0.05) and there was a relative high expression 7 days after operation (P lt; 0.01). Conclusion Tβ4 can inhibit the expression of laminin 5 early, and then up-regulate laminin 5 expression to moderate the reformation of ECM, promote the migration of epidemic cell and accelerate skin wound heal ing.
目的 观察局灶性脑缺血梗死区周围层粘连蛋白（Laminin）及基质金属蛋白酶（MMP）-9表达，探讨其在脑缺血再灌注损伤发病机制中的作用。 方法 将45只体重250～300 g、4个月龄的Sprague Dawley（SD）雄性大鼠随机分为假手术（Sham）组与局灶性缺血再灌注（MCAO）组。MCAO组又分6、24、72 h及7 d组，每组各9只。光学显微镜下观察脑组织病理改变，免疫组织化学染色检测各组Laminin及MMP-9的表达情况。 结果 再灌注后6 h，即有MMP-9表达明显增高，表达高峰出现在再灌注后24 h，3 d时有所下降，至7 d时仍明显高于基础水平（P＜0.01）。Laminin的表达于再灌注6 h开始降低，24 h降至最低，3 d后开始缓慢回升。 结论 脑缺血再灌注后，病变区MMP-9与Laminin表达变化可反映脑血管壁受损状况，并与脑缺血损伤及修复过程有关。
ObjectiveBased on the cell-extracellular matrix adhesion theory in selective cell retention (SCR) technology, demineralized bone matrix (DBM) modified by simplified polypeptide surface was designed to promote both bone regeneration and angiogenesis.MethodsFunctional peptide of α4 chains of laminin protein (LNα4), cyclic RGDfK (cRGD), and collagen-binding domain (CBD) peptides were selected. CBD-LNα4-cRGD peptide was synthesized in solid phase and modified on DBM to construct DBM/CBD-LNα4-cRGD scaffold (DBM/LN). Firstly, scanning electron microscope and laser scanning confocal microscope were used to examine the characteristics and stability of the modified scaffold. Then, the adhesion, proliferation, and tube formation properties of CBD-LNα4-cRGD peptide on endothelial progenitor cells (EPCs) were detected, respectively. Western blot method was used to verify the molecular mechanism affecting EPCs. Finally, 24 10-week-old male C57 mice were used to establish a 2-mm-length defect of femoral bone model. DBM/LN and DBM scaffolds after SCR treatment were used to repair bone defects in DBM/LN group (n=12) and DBM group (n=12), respectively. At 8 weeks after operation, the angiogenesis and bone regeneration ability of DBM/LN scaffolds were evaluated by X-ray film, Micro-CT, angiography, histology, and immunofluorescence staining [CD31, endomucin (Emcn), Ki67].ResultsMaterial related tests showed that the surface of DBM/LN scaffold was rougher than DBM scaffold, but the pore diameter did not change significantly (t=0.218, P=0.835). After SCR treatment, DBM/LN scaffold was still stable and effective. Compared with DBM scaffold, DBM/LN scaffold could adhere to more EPCs after the surface modification of CBD-LNα4-cRGD (P<0.05), and the proliferation rate and tube formation ability increased. Western blot analysis showed that the relative expressions of VEGF, phosphorylated FAK (p-FAK), and phosphorylated ERK1/2 (p-ERK1/2) proteins were higher in DBM/LN than in DBM (P<0.05). In the femoral bone defect model of mice, it was found that mice implanted with DBM/LN scaffold had stronger angiogenesis and bone regeneration capacity (P<0.05), and the number of CD31hiEmcnhi cells increased significantly (P<0.05).ConclusionDBM/LN scaffold can promote the adhesion of EPCs. Importantly, it can significantly promote the generation of H-type vessels and realize the effective coupling between angiogenesis and bone regeneration in bone defect repair.