General Description
Human PDGF (Platelet-Derived Growth Factor) is a general term for a small group of structurally-related and secreted growth factors. These growth factors are widely expressed, disulffde-linked, and represent the products of four distinct genes. There are currently five named PDGFs, all of which belong to the PDGF/VEGF family, cysteine-knot superfamily of proteins. Within the PDGF family, there are two subfamilies that are characterized by either the presence, or absence, of a CUB (C1r/Cls, Urchin EGF-like, and BMP1-1) domain (1-4). Two genes (PDGF-C and -D) contain the CUB domain, while the remaining two genes (PDGF-A and -B) do not. Although all four PDGF gene products form homodimers, a PDGF-AB covalent heterodimer is also formed. Heterodimer formation appears to be a random process (5). All PDGFs are synthesized as inactive proforms that undergo intracellular or extracellular proteolytic processing to become active (3), and all PDGFs contain at least one isoform that binds heparin (2).
Human PDGF-AB is a 28-30 kDa, disulffde-linked heterodimer containing one 14-16 kDa PDGF-A chain and one 12-14 kDa PDGF-B chain (6, 7, 8). Although it appears to be the first PDGF isoform recognized, its presence is suggested to be unusual in that the PDGF-A and -B chains are not commonly coexpressed (2). This may be particularly true in platelets where its appearance in α-granules may be a human phenomenon (2). When, however, the -A and -B chains are coexpressed, their association, albeit random, is preferred over the formation of homodimers (5, 9). Mature PDGF-AB is formed intracellularly by the initial dimerization of PDGF-A and -B proprecursors, followed by the subsequent cleavage of each molecule's proregions (5, 10). The PDGF-A proprecursor is 24 kDa in size, and 191 amino acids (aa) in length (6, 11). Although mature 15-16 kDa PDGF-A chain is generated following cleavage after Arg86, it appears that the 8 kDa cleaved proregion remains non-covalently associated with the mature chain during secretion, acting as a block against unwanted receptor activation (12). The 31 kDa PDGF-B proprecursor is 221 aa in length (7, 10). It has two proteolytic processing sites, one after Arg81, and a second after Thr190 that may or may not be utilized. This generates a 12-14 kDa 109 aa mature molecule (aa 82-190) that also likely continues a non-covalent association with its N-terminal proregion. Mature human PDGF-A and -B share 95% and 89% aa sequence identity with mouse PDGF-A and -B, respectively. Cells known to express PDGF-AB include platelets, endothelial cells, keratinocytes, macrophages, retinal pigment epithelium and neurons (5, 13, 14). PDGF-AB has documented chemotactic properties for multiple cell types, and is also known to be mitogenic for diverse cell types. Relative to the related homodimeric isoforms, PDGF-AB will induce mitosis but will not mediate IP3 (D-myo-inositol1,4,5-trisphosphate) production by vascular smooth muscle cells. PDGF-AA, by contrast, has no effect on either mitosis or IP3 production, while PDGF-BB induces both mitosis and IP3 production (14). Thus, it would appear that the three members (isoforms) of the non-CUB domain containing subfamily have some non-overlapping activities.
There are a number of molecules that either directly or indirectly participate in PDGF binding. The traditional receptor(s) for PDGF is considered to be either a homodimer or heterodimer created from two 170-180 kDa type I transmembrane RTKs termed PDGF Rα and PDGF Rβ (2, 5). In vitro studies have established that the αα homodimer will bind PDGF-AA, -AB, -BB, and -CC, the αβ heterodimer will bind -AB, -BB and -CC, and the ββ homodimer will bind -BB and -DD (4). In vivo studies have conffrmed PDGF-AA and -CC acting through αα, and -BB and -DD acting through ββ (2). Other molecules that participate in PDGF binding include LRP1 which forms a complex with PDGF Rβ (15, 16), FGF R1 that complexes with both PDGF Rβ and PDGF Rα (17, 18), and neuropilin-1, which complexes with PDGF Rα (19). PDGF-BB will also bind to SorLA/LR11 and to circulating SPARC (20, 21). PDGF-A and -B gene products also bind to circulating α2-macroglobulin (22).
The PDGF-AB Immunoassay is a 4.5 hour solid phase ELISA designed to measure human PDGF-AB in cell culture supernates, serum, and platelet-poor plasma. It contains E. coli-expressed recombinant human PDGF-AB and has been shown to accurately quantitate the recombinant factor. Results obtained using natural human PDGF-AB showed linear curves that were parallel to the standard curves obtained using the recombinant kit standards. These results indicate that this kit can be used to determine relative mass values for natural human PDGF-AB.
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