Abstract：
StrataGraft® (allogeneic cultured keratinocytes and dermal fibroblasts in murine collagen-dsat) is an FDA-approved viable bioengineered allogeneic cellularized construct for adult patients with deep partial-thickness burns requiring surgery. We characterized the structural and functional properties of StrataGraft to improve product understanding by evaluating extracellular matrix (ECM) molecule distribution and secreted protein factor expression in vitro.
ECM protein expression was determined using indirect immunofluorescence on construct cross sections using commercial antibodies against collagen III, IV, VI, laminin-332, and decorin. Human collagen I expression was verified by enzyme-linked immunosorbent assay (ELISA) for collagen I C-terminal propeptide. Soluble protein factor secretion was quantified by multiplex biomarker assays and singleplex ELISA in conditioned media from meshed constructs.
StrataGraft cellular components produced collagen I, collagen III, collagen VI, and decorin in patterns indicating an organized ECM. Distributions of collagen IV and laminin-332 indicated formation of basement membranes and dermal-epidermal junctions. Soluble protein factors were observed in the pg/cm2/h range from 1 h to the experiment end at 168 h.
The organization of the ECM proteins was like human skin and the viable cellular components provided sustained secretion of soluble wound healing factors, making StrataGraft an attractive option for treating severe burns.
ECM protein expression was determined using indirect immunofluorescence on construct cross sections using commercial antibodies against collagen III, IV, VI, laminin-332, and decorin. Human collagen I expression was verified by enzyme-linked immunosorbent assay (ELISA) for collagen I C-terminal propeptide. Soluble protein factor secretion was quantified by multiplex biomarker assays and singleplex ELISA in conditioned media from meshed constructs.
StrataGraft cellular components produced collagen I, collagen III, collagen VI, and decorin in patterns indicating an organized ECM. Distributions of collagen IV and laminin-332 indicated formation of basement membranes and dermal-epidermal junctions. Soluble protein factors were observed in the pg/cm2/h range from 1 h to the experiment end at 168 h.
The organization of the ECM proteins was like human skin and the viable cellular components provided sustained secretion of soluble wound healing factors, making StrataGraft an attractive option for treating severe burns.
摘要：
背景：StrataGraft®（同种异体培养的角质形成细胞和小鼠胶原dsat中的真皮成纤维细胞）是FDA批准的可行的生物工程同种异体细胞化构建体，适用于需要手术的部分深度烧伤成年患者。我们表征了Strata移植物的结构和功能特性，以通过评估细胞外基质（ECM）分子分布和分泌的蛋白质因子表达来提高产品的理解。
方法：ECM蛋白的表达使用抗胶原蛋白III的商业抗体在构建体横截面上使用间接免疫荧光进行测定，IV,VI,层粘连蛋白-332和decorin.通过酶联免疫吸附测定(ELISA)证实人I型胶原C端前肽的表达。通过多重生物标志物测定和单重ELISA在来自网状构建体的条件培养基中定量可溶性蛋白质因子分泌。
结果：StrataGraft细胞成分产生胶原蛋白I，胶原蛋白III，胶原蛋白VI，和表明有组织的ECM的模式中的核心蛋白。胶原IV和层粘连蛋白332的分布表明基底膜和真皮-表皮连接的形成。从1小时到168小时的实验结束，在pg/cm2/h范围内观察到可溶性蛋白质因子。
结论：ECM蛋白的组织类似于人类皮肤，活的细胞成分提供可溶性伤口愈合因子的持续分泌，使StrataGraft成为治疗严重烧伤的有吸引力的选择。
方法：ECM蛋白的表达使用抗胶原蛋白III的商业抗体在构建体横截面上使用间接免疫荧光进行测定，IV,VI,层粘连蛋白-332和decorin.通过酶联免疫吸附测定(ELISA)证实人I型胶原C端前肽的表达。通过多重生物标志物测定和单重ELISA在来自网状构建体的条件培养基中定量可溶性蛋白质因子分泌。
结果：StrataGraft细胞成分产生胶原蛋白I，胶原蛋白III，胶原蛋白VI，和表明有组织的ECM的模式中的核心蛋白。胶原IV和层粘连蛋白332的分布表明基底膜和真皮-表皮连接的形成。从1小时到168小时的实验结束，在pg/cm2/h范围内观察到可溶性蛋白质因子。
结论：ECM蛋白的组织类似于人类皮肤，活的细胞成分提供可溶性伤口愈合因子的持续分泌，使StrataGraft成为治疗严重烧伤的有吸引力的选择。
