Xenbase Image ID: 149308

Figure 5. Mammalian Schwann cells promote synapse formation in vitro. A , The expression of TGF-β1 mRNA in mammalian tissues was examined using RT-PCR. Cultured rat Schwann cells, as well as sciatic nerves from neonatal rats, expressed TGF-β1 mRNA. B , The presence of TGF-β1 protein was investigated by Western blot. TGF-β1 protein was detected in mammalian SC-CM. Human recombinant TGF-β1 was used as a positive control and culture medium (DMEM) was used as a negative control for Western blot. C , D , Xenopus nerve–muscle cocultures were treated with trophic stimulation only. The formation of AChR clusters was reduced along the nerve–muscle contacts (arrowheads). E , F , Nerve–muscle cocultures with trophic stimulation were treated with TGF-β1. The formation of AChR clusters was restored at nerve–muscle contacts (arrowheads), as demonstrated by the positive staining of α-BTX. G , H , Nerve–muscle cocultures were treated with mammalian SC-CM in the presence of trophic stimulation. The addition of mammalian SC-CM increased the formation of AChR clusters at nerve–muscle contacts (arrowheads). I , J , Nerve–muscle cocultures were treated with mammalian SC-CM that was immunodepleted of TGF-β1 protein. In this example, note that no α-BTX staining was observed at nerve–muscle contacts (arrowheads). Scale bar in J applies to C–I . K , Quantification of the formation of AChR clusters at nerve–muscle contacts. Data from five independent experiments were combined. In control nerve–muscle cocultures with trophic stimulation alone, only 18.3 ± 2.5% of nerve–muscle contacts (n = 242) were associated with AChR clusters. The addition of mammalian SC-CM increased the percentage of nerve–muscle contacts associated with AChR clusters to 66.8 ± 4.5% (mSC-CM, 247 contacts), which is similar to the effect produced by treatment with TGF-β1 (TGF-β1, 68.1 ± 2.5%, 279 contacts). When TGF-β1 protein was immunodepleted from SC-CM, SC-CM was no longer effective in promoting the formation of AChR clusters at nerve–muscle contacts (mSC-CM after immunoprecipitation with mAb TGF-β1, 18.6 ± 2.2, n = 311 contacts]. Because mammalian Schwann cells were cultured in DMEM, DMEM was used as a control. DMEM alone had no effect on the formation of AChR clusters at nerve–muscle contacts (DMEM, 17.4 ± 1.3%, n = 238 contacts). Data are mean ± SEM, *p < 0.05; two-tailed, unequal variance Student's t test. Image published in: Feng Z and Ko CP (2008) Copyright © 2008. OA ARTICLE, images redisplayed under a Creative Commons license. Gene Synonyms Species Stage(s) Tissue syn1.L syn1-a, syn1-b, synapsin, synapsin 1, synapsin I X. laevis Sometime during NF stage 1 to NF stage 66 neuron Image source: Published Larger Image Printer Friendly View

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