[PubMed] [Google Scholar] 51. without hindering the neurite-promoting potential of laminin. Likewise, neuritic growth by DRGn cultured on normal nerve sections was increased markedly by first treating the nerve sections with MMP-2. The proteolytic deinhibition by MMP-2 was equivalent to and nonadditive with that achieved by chondroitinase, suggesting that both enzymes inactivated inhibitory CSPG. Additionally, the increases in neuritic growth resulting from treating nerve sections with MMP-2 or chondroitinase were blocked by anti-laminin antibodies. From these results we Ioversol conclude that MMP-2 provides a mechanism for the deinhibition of laminin in the endoneurial basal lamina and may play an important role in the regeneration of peripheral nerve. (Muir et al., 1989a; Snow et al., 1991). CSPGs permeate boundary structures confronted by developing and regenerating axons (Oakley and Tosney, 1991; Brittis et al., 1992;Pindzola et al., 1993). Normal adult peripheral nerve contains neurite-inhibiting activity associated with CSPG, which is increased further in the distal nerve after injury (Zuo et al., 1998). CSPG colocalizes with laminin within the Schwann cell basal lamina and appears to have a wider distribution within the endoneurial compartment in degenerating nerve (Kuecherer-Ehret et al., 1990; Tona et al., 1993). Because of its considerable regenerative capacity, it may seem counter-intuitive that peripheral nerve contains neurite inhibitors. One possibility is that growth inhibitors help to stabilize axons in normal nerve. By blocking the neurite-promoting potential of laminin in the endoneurial basal lamina, inhibitory CSPG may prevent axons from sprouting collaterals, especially at nodes of Ranvier. This implies that nerve sheaths actually might suppress axonal growth under normal conditions. However, this suppression somehow must be reversed in nerve regeneration. The growth cone, the leading structure of growing axons, senses guidance cues from the surrounding environment and implements directed outgrowth. Axonal regeneration requires extensive growth cone motility and infiltration within damaged and degenerating nervous tissue. Substantial evidence now indicates that neurons secrete matrix-degrading enzymes and actively remodel surrounding ECM substrata (Monard, 1988; Pittman and Buettner, 1989; Fambrough et al., 1996). Proteinases, including plasminogen activators and matrix metalloproteinases (MMPs), are expressed by peripheral neurons and released by growth cones, implicating the growing tip of axons Ioversol in the proteolysis of matrix components (Pittman and Williams, 1988; McGuire and Seeds, 1990; Muir, 1994; Nordstrom et al., 1995; Hayden and Seeds, 1996). MMPs are believed to be the physiologically relevant mediators of ECM degradation and matrix remodeling (Matrisian, 1992). MMPs also participate in the proteolytic processing of both ECM and cell surface components, which can result in alterations of cell properties such as attachment and migration (Chantry et al., 1992; DiStefano et al., 1993;Ray and Stetler-Stevenson, 1995; Giannelli et al., 1997). Thus, the function of MMPs, at first thought to be limited to the catabolism and LASS2 antibody cellular infiltration of ECM, also includes an important regulatory mechanism for the proteolytic activation of cryptic molecular domains. In the present report, axonal growth by regenerating embryonic dorsal root ganglionic neurons (DRGn) cultured on sections of normal adult nerve and on a synthetic substratum composed of laminin and inhibitory CSPG was found to be dependent on metalloproteinase activity. DRGn expressed MMP-2, which inactivated inhibitory CSPG and unmasked the neurite-promoting activity of associated laminin. From these studies we conclude that MMP-2 provides a mechanism for the deinhibition of laminin in the endoneurial basal lamina. MATERIALS AND METHODS (Muir, 1994). In the present study a defined culture medium (DMEM/N2+NGF) was formulated without cysteine to minimize metalloproteinase inhibition (see Materials and Methods). All DRGn bioassays included NGF in the culture medium. In a standard 4 hr bioassay, neuritic growth on a NIFClaminin substratum was Ioversol virtually the same with or without cysteine in the medium. However, in the absence of cysteine, gradual neuritic growth was observed on NIFClaminin that was not seen previously by neurons grown in the presence of cysteine. To quantitate this effect, we established a NIFClaminin substratum, using a submaximal concentration of NIF (2 NIU/ml) and a maximal concentration of laminin (1 g/ml). On.