Extracellular matrix protein adsorption to phosphate-functionalized gels from serum promotes osteogenic differentiation of human mesenchymal stem cells

One of the primary goals for tissue engineering is to induce new tissue formation by stimulating specific cell function. Human mesenchymal stem cells (hMSCs) are a particularly important cell type that has been widely studied for differentiation down the osteogenic (bone) lineage, and we recently found that simple phosphate functional groups incorporated into poly(ethylene glycol) (PEG) hydrogels could induce osteogenesis without using differentiation medium by unknown mechanisms. Here, we aimed to determine whether direct or indirect cell/materials interactions were responsible for directing hMSCs down the osteogenic lineage on phosphate (PO₄)-functionalized PEG hydrogels. Our results indicated that serum components adsorbed onto PO₄-PEG hydrogels from medium in a presoaking step were sufficient for attachment and spreading of hMSCs, even when seeded in serum-free conditions. Blocking antibodies for collagen and fibronectin (targeted to the hydrogel), as well as β1 and β3 integrin blocking antibodies (targeted to the cells), each reduced attachment of hMSCs to PO₄-PEG hydrogels, suggesting that integrin-mediated interactions between cells and adsorbed matrix components facilitate attachment and spreading. Outside-in signaling, and not merely shape change, was found to be required for osteogenesis, as alkaline phosphatase activity and expression of CBFA1, osteopontin and collagen-1 were each significantly down regulated upon inhibition of focal adhesion kinase phosphorylation even though the focal adhesion structure or cell shape was unchanged. Our results demonstrate that complex function (i.e. osteogenic differentiation) can be controlled using simple functionalization strategies, such as incorporation of PO₄, but that the role of these materials may be due to more complex influences than has previously been appreciated.     

Synthetic hydrogel scaffold is an effective vehicle for delivery of INFUSE (rhBMP2) to critical‐sized calvaria bone defects in rats

Medtronic's INFUSE Bone Graft provides surgeons with a potent tool for stimulating bone formation. Current delivery vehicles that rely on Absorbable Collagen Sponges (ACS) require excessive quantities of the active ingredient in INFUSE, recombinant human Bone Morphogenic Protein‐2 (rhBMP2), to achieve physiologically relevant concentrations of the growth factor, driving up the cost of the product and increasing the likelihood of undesirable side effects in neighboring tissues. We demonstrate that a simple light‐mediated, thiol‐ene chemistry can be used to create an effective polymer delivery vehicle for rhBMP2, eliminating the use of xenographic materials and reducing the dose of rhBMP2 required to achieve therapeutic effects. Comprised entirely of synthetic components, this system entraps rhBMP2 within a biocompatible hydrogel scaffold that is degraded by naturally occurring remodeling enzymes, clearing the way for new tissue formation. When tested side‐by‐side with ACS in a critical‐sized bone defect model in rats, this polymeric delivery system significantly increased bone formation over ACS controls. © 2012 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 31: 401–406, 2013     

ADAMTS13 testing update: Focus on laboratory aspects of difficult thrombotic thrombocytopenic purpura diagnoses and effects of new therapies

TTP is a life-threatening disorder diagnosed using a combination of clinical information and laboratory results. ADAMTS13 activity and antibody testing represent a major advance in the field, but results can sometimes be difficult to interpret due to technical aspects of the tests and characteristics of the causative antibodies in acquired TTP. Genetic testing for ADAMTS13 mutations is also now available to assist with the diagnosis of inherited TTP. This review will focus on ADAMTS13 testing and will highlight patient and laboratory aspects that can lead to diagnostic difficulty. The effects of TTP therapies on test results will also be discussed.