Perlecan domain 1 recombinant proteoglycan augments BMP-2 activity and osteogenesis.
Perlecan domain 1 recombinant proteoglycan augments BMP-2 activity and osteogenesis.
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DOI：
10.1186/1472-6750-12-60
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发表时间：
2012-09-11
影响因子：
3.5
通讯作者：
Whitelock JM
中科院分区：
文献类型：
--
作者：
Decarlo AA;Belousova M;Ellis AL;Petersen D;Grenett H;Hardigan P;O'Grady R;Lord M;Whitelock JM
许多生长因子，如骨形态发生蛋白(BMP)-2，已被证明与被称为硫酸乙酰肝素(HS)糖胺聚糖(GAGS)的硫酸双链聚合物相互作用，这种聚合物存在于全身基质和细胞表面的蛋白多糖中。HS GAG和一些高度硫酸盐化形式的硫酸软骨素(CS)通过在GF与其受体相互作用中作为辅助因子或辅助受体来调节细胞功能，HS或CS GAG已被证明是诱导信号和GF活性所必需的，甚至在成骨细胞系中也是如此。然而，与重组蛋白不同的是，HS和CS GAG是非常异质性的，这在很大程度上是由于翻译后添加，然后去除，硫酸盐基团沿着GAG聚合物的不同位置。因此，我们研究了一种DNA前体药物是否可行，以在体内原位产生可溶的HS/CS蛋白多糖，从而增强包括BMP-2在内的生长因子的活性。利用从HEK 293细胞中表达的含有HS和CS GAG的重组人Perlecan结构域1(rh Pln.d1)，在体外证实了rh BMP-2活性的紧密结合和剂量增强。在体外，用硫酸盐化的HS和CS GAG对表达的重组人Pln.D1进行修饰。重组人骨形态发生蛋白-2的剂量增强由pln.d1表达载体作为冻干的单相一起在颗粒磷酸三钙支架上输送6周或更长时间，在大鼠上颌骨模型中产生的新生骨体积是没有pln.d1表达载体对照部位的9倍。使用明显更低的BMP-2剂量，这种组合提供了超过5倍的上颌骨隆起和更高的密度，使用胶原海绵(Inuse™)提供的重组BMP-2。在结合和基于细胞的分析中，重组HS/CS PG与BMP-2在功能上具有很强的相互作用，在体内，当以单相形式一起输送到支架上时，pln.247表达载体显著提高了BMP-2在体内成骨活性的剂量效应。HS/CS PGs的使用可能有助于加强GF的治疗，而基于质粒的方法在这里被证明是非常有效的。
Many growth factors, such as bone morphogenetic protein (BMP)-2, have been shown to interact with polymers of sulfated disacharrides known as heparan sulfate (HS) glycosaminoglycans (GAGs), which are found on matrix and cell-surface proteoglycans throughout the body. HS GAGs, and some more highly sulfated forms of chondroitin sulfate (CS), regulate cell function by serving as co-factors, or co-receptors, in GF interactions with their receptors, and HS or CS GAGs have been shown to be necessary for inducing signaling and GF activity, even in the osteogenic lineage. Unlike recombinant proteins, however, HS and CS GAGs are quite heterogenous due, in large part, to post-translational addition, then removal, of sulfate groups to various positions along the GAG polymer. We have, therefore, investigated whether it would be feasible to deliver a DNA pro-drug to generate a soluble HS/CS proteoglycan in situ that would augment the activity of growth-factors, including BMP-2, in vivo. Utilizing a purified recombinant human perlecan domain 1 (rhPln.D1) expressed from HEK 293 cells with HS and CS GAGs, tight binding and dose-enhancement of rhBMP-2 activity was demonstrated in vitro. In vitro, the expressed rhPln.D1 was characterized by modification with sulfated HS and CS GAGs. Dose-enhancement of rhBMP-2 by a pln.D1 expression plasmid delivered together as a lyophilized single-phase on a particulate tricalcium phosphate scaffold for 6 or more weeks generated up to 9 fold more bone volume de novo on the maxillary ridge in a rat model than in control sites without the pln.D1 plasmid. Using a significantly lower BMP-2 dose, this combination provided more than 5 times as much maxillary ridge augmentation and greater density than rhBMP-2 delivered on a collagen sponge (InFuse™). A recombinant HS/CS PG interacted strongly and functionally with BMP-2 in binding and cell-based assays, and, in vivo, the pln.247 expression plasmid significantly improved the dose-effectiveness of BMP-2 osteogenic activity for in vivo de novo bone generation when delivered together on a scaffold as a single-phase. The use of HS/CS PGs may be useful to augment GF therapeutics, and a plasmid-based approach has been shown here to be highly effective.
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影响因子：
4.8
作者：
Khan, Sanaullah;Rodriguez, Elizabeth;Perkins, Stephen J.
通讯作者：
Perkins, Stephen J.
影响因子：
4.3
作者：
Ito, Y;Hikino, M;Sugahara, K
通讯作者：
Sugahara, K
影响因子：
--
作者：
Muthusamy, Arivalagan;Cooper, Carlton R.;Gomes, Ronald R., Jr.
通讯作者：
Gomes, Ronald R., Jr.
影响因子：
4.8
作者：
Knox, S;Merry, C;Whitelock, J
通讯作者：
Whitelock, J
影响因子：
4.8
作者：
Robinson, CJ;Mulloy, B;Stringer, SE
通讯作者：
Stringer, SE