We have recently reported that CBFb is critical for Vifinduced A3G polyubiquitination and degradation

es with the observation that YKL-40 protein is not usually found in normal human CNS neurons, nor generally in normal brain tissues with the exception of low level expression in a subpopulation of microglia and reactive astrocytes. The ATRA/bFGF/B27 differentiation protocol was previously dem- 5 YKL-40 Expression in MSCs 6 YKL-40 Expression in MSCs onstrated to differentiate MSCs into a phenotype that expressed glial and neuronal progenitor markers, synthesized, packaged and released neurotransmitters and produced spontaneous post-synaptic currents, thus further corroborating the absence of YKL-40 protein production with the neuronal phenotype. While very low levels of YKL-40 mRNA have been reported in homogenates of normal brain tissues, the continued presence of high levels of YKL40 mRNA following ATRA/bFGF/B27 neuronal differentiation suggests that despite the many similarities to mature neurons, some differences exist between normal neurons differentiated from neural stem cells and neurons trans-differentiated from MSCs using this protocol. This also indicates that the mechanisms suppressing YKL-40 translation in undifferentiated MSCs remain active in ATRA/bFGF/B27 differentiated MSCs. The presence of YKL-40 protein 17526600 in MSCs following treatment with -mercaptoethanol as a trans-differentiating agent suggests that MSCs differentiated using this protocol represent even less of a true neuronal phenotype than those transdifferentiated using the 7623957 ATRA/bFGF/B27 protocol. While MSCs differentiated using small molecule chemical inducers such as mercaptoethanol and DMSO show increases in the expression of some neuronal markers, they also show increased apoptosis and do not show the electrophysiological properties of neurons in patch clamp experiments. It should be noted that any in vitro transdifferentiation protocol may push MSCs to assume a phenotype that is beyond their normal differentiation limits in vivo. Transdifferentiation may result in cells with significant epigenetic differences as compared to the normal phenotype, and care must be taken both in interpreting results of studies that push cells well beyond the natural limits of differentiation, and in utilizing transdifferentiated cells for therapeutic applications. The mechanism of suppression of YKL-40 mRNA translation in undifferentiated MSCs remains unknown. A common mechanism of translational suppression is miRNA binding to mRNA 39 sequences, and reports have documented significant changes in the levels of specific miRNAs in MSC differentiation, including miR27a, miR-148b and miR-489 in MSC differention into osteoblasts and miR-145 in MSC differentiation into chondrocytes. None of these miRNAs were identified by the major miRNA target search algorithms as being able to target YKL-40 mRNA, and the miRNAs that were listed as being able to target YKL40 mRNA sequences were not among those identified as being significantly up or downregulated during MSC differentiation. Thus if YKL-40 mRNA is being silenced in undifferentiated MSCs by miRNAs, it is being silenced either by miRNA species that bind in a PR619 manner that is not recognized by the current algorithms, or by miRNA species whose total levels do not change appreciably during differentiation but whose available levels change due to significant changes in the levels of other binding partners during osteogenic and chondrogenic differentation. These results further support the utility of YKL-40 protein expression as a marker for MSC di