Abstract
The glycoprotein YKL-40 (CHI3L1) is a secreted chitinase family protein that induces angiogenesis, cell survival, and cell proliferation, and plays roles in tissue remodeling and immune regulation. It is expressed primarily in cells of mesenchymal origin, is overexpressed in numerous aggressive carcinomas and sarcomas, but is rarely expressed in normal ectodermal tissues. Bone marrow-derived mesenchymal stem cells (MSCs) can be induced to differentiate into various mesenchymal tissues and trans-differentiate into some non-mesenchymal cell types. Since YKL-40 has been used as a mesenchymal marker, we followed YKL-40 expression as undifferentiated MSCs were induced to differentiate into bone, cartilage, and neural phenotypes. Undifferentiated MSCs contain significant levels of YKL-40 mRNA but do not synthesize detectable levels of YKL-40 protein. MSCs induced to differentiate into chondrocytes and osteocytes soon began to express and secrete YKL-40 protein, as do ex vivo cultured chondrocytes and primary osteocytes. In contrast, MSCs induced to trans-differentiate into neurons did not synthesize YKL-40 protein, consistent with the general absence of YKL-40 protein in normal CNS parenchyma. However, these trans-differentiated neurons retained significant levels of YKL-40 mRNA, suggesting the mechanisms which prevented YKL-40 translation in undifferentiated MSCs remained in place, and that these trans-differentiated neurons differ in at least this way from neurons derived from neuronal stem cells. Utilization of a differentiation protocol containing β-mercaptoethanol resulted in cells that expressed significant amounts of intracellular YKL-40 protein that was not secreted, which is not seen in normal cells. Thus the synthesis of YKL-40 protein is a marker for MSC differentiation into mature mesenchymal phenotypes, and the presence of untranslated YKL-40 mRNA in non-mesenchymal cells derived from MSCs reflects differences between differentiated and trans-differentiated phenotypes.
Highlights
The human chitinase family glycoprotein YKL-40 (CHI3L1) has been implicated in tissue remodeling, angiogenesis, and cell survival in both normal and pathological conditions [1,2,3,4]
Undifferentiated MSCs have been reported to express YKL40 mRNA, so initial experiments were performed to establish the levels of YKL-40 mRNA and the corresponding levels of YKL-40 protein synthesized by undifferentiated mesenchymal stem cells [35,36]
To insure that YKL-40 was not being translated and rapidly degraded by the proteasome, undifferentiated MSCs were incubated with the proteosome inhibitor MG132 for up to 6 hours, western blot analysis demonstrated that YKL-40 protein was not present in either the cell lysate or media supernatant at any time within this 6 hour period (Figure 2B)
Summary
The human chitinase family glycoprotein YKL-40 (CHI3L1) has been implicated in tissue remodeling, angiogenesis, and cell survival in both normal and pathological conditions [1,2,3,4]. YKL-40 protein is present in normal human serum at concentrations in the low nanomolar range [10]. It is elevated in the serum of patients and in the affected cells of a number of noncancerous pathological conditions including rheumatoid arthritis [11], asthma [12], and hepatic fibrosis/cirrhosis [13,14,15]. YKL-40 serum protein levels were directly correlated with morbidity and or mortality in patients suffering from cancers of the breast [16,17,18], colon [19,20], ovaries [21], and brain [22]. YKL-40 is expressed in a limited number of brain cancers, and is the most highly upregulated gene in high grade glioblastoma multiformae. YKL40 expression in these high grade glioblastoma multiformae appears linked to the degree of tumor vascularization [23]
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