Cholinergic properties of the SN56.B5.G4 cell line derived from the fusion of neurons of the mouse postnatal day 21 septum and the murine neuroblastoma cell line N18TG2 were investigated and correlated with morphological differentiation. In basal serum-containing growth medium, few cells developed neurites. Neurite extension occurred in cells grown for 2 d with forskolin or dibutyryl cAMP (dbcAMP) but not with butyrate. In cells treated with these compounds, the activity of ChAT and ACh content were two- to threefold higher relative to controls. The cells synthesized ACh from choline taken up by the sodium-dependent high-affinity transport. Forskolin-, dbcAMP-, and butyrate-treated cells (but not the controls) were capable of spontaneous and depolarization-evoked ACh release. The results indicate that the morphological and the neurochemical aspects of SN56.B5.G4 cell differentiation are independently regulated.

Laminin-332 (LN-332), which is essential for epithelial cell adhesion and migration, is up-regulated in most invasive carcinomas. Association between LN-332 and carcinoma cell integrins and stroma collagen is thought to be important for tumor growth and metastasis. Here, we show that function blocking LN-332 antibodies interfering with cellular adhesion and migration in vitro evoke apoptotic pathways. The antibodies also target epithelial tumors in vivo. Antibodies against the cell binding domain of the a3 chain of LN-332 inhibited tumor growth by up to 68%, and antibodies against the matrix binding domains of the b3 and c2 chains significantly decreased lung metastases. The LN-332 antibodies appear to induce tumor cell anoikis and subsequent programmed cell death and reduce migration by interfering with tumor cell matrix interactions. ' UICCKey words: laminin; basement membranes; metastasis; immunotherapy Laminins are trimeric basement membrane (BM) glycoproteins with roles in cell adhesion, proliferation, migration and differentiation. In mammals, 5 genetically distinct a, 3b and 3g chains can form at least 14 different combinations of these chains.1,2 Laminin-332 (LN-332, previously termed laminin-5) that has chain composition a3:b3:g2 ( Supplementary Fig. 1) is essential for anchorage of epithelial cells and specifically found in epithelial BMs.3-5 LN-332 defects lead to detachment of epithelia and the fatal skin blistering disease junctional epidermolysis bullosa.6-8 LN-332 also has a role in proliferation and locomotion of epithelial cells, such as in keratinocytes of healing wounds.9,10 The globular domain (Gdomain) of the a3 chain binds to the cell surface through integrin receptors a6b4 and a3b1 11 and evokes anti-apoptotic signals through focal adhesion kinase, 12,13 while the short arms of the b3 and g2 chains bind to Type VII collagen in the stroma. 14 LN-332 is up-regulated in various epithelial cancers, including colon, gastric, mammary duct and squamous cell carcinomas, as well as melanomas, [15][16][17][18] but not in mesenchymal cancers. 15,16 High expression of the g2 chain of LN-332 has been found to correlate with poor prognosis of cervical squamous cell carcinomas.19 LN-332 is also a major scattering factor stimulating invasive and metastatic capacity of several tumor cell lines in vitro. 20,21 In the cancer tissue, the protein is primarily expressed at the invasive front, as well as in micro-metastases. Down-regulation of LN-332 has been reported in epithelial prostate cancer 22 and also in breast cancers. 23LN-332 expression has been associated with tumorigenesis. Thus, when HT1080 tumor cells constitutively expressing laminin b3 and g2 chains but not a3 were transfected with laminin a3 cDNA, the cells grew significantly larger tumors in nude mice than untransformed cells. 24 Moreover, LN-332 negative (as well as a4 integrin negative) keratinocytes did not become tumorigenic upon transfection with Ras-IjBa in contrast to normal keratinocytes. 25 Since most cancers are of epithelial origi...

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