Developmentally Arrested Basket/Stellate Cells in Postnatal Human Brain as Potential Tumor Cells of Origin for Cerebellar Hemangioblastoma in von Hippel-Lindau Patients.

von Hippel-Lindau (VHL) disease is an autosomal dominant hereditary cancer disorder caused by a germline mutation in the VHL tumor suppressor gene. Loss of the wild-type allele results in VHL deficiency and the potential formation of cerebellar hemangioblastomas, which resemble embryonic hemangioblast proliferation and differentiation processes. Multiple, microscopic, VHL-deficient precursors, termed developmentally arrested structural elements (DASEs), consistently involve the cerebellar molecular layer in VHL patients, indicating the tumor site of origin. Unlike hemangioblastomas, however, cerebellar DASEs do not express brachyury, a mesodermal marker for hemangioblasts. In this study, neuronal progenitors occupying the molecular layer were investigated as tumor cells of origin. By immunohistochemistry, cerebellar DASEs and hemangioblastomas lacked immunoreactivity with antibody ZIC1 (Zic family member 1), a granule cell progenitor marker with concordance from oligonucleotide RNA expression array analyses. Rather, cerebellar DASEs and hemangioblastomas were immunoreactive with antibody PAX2 (paired box 2), a marker of basket/stellate cell progenitors. VHL cerebellar cortices also revealed PAX2-positive cells in Purkinje and molecular layers, resembling the histological and molecular development of basket/stellate cells in postnatal non-VHL mouse and human cerebella. These data suggest that VHL deficiency can result in the developmental arrest of basket/stellate cells in the human cerebellum and that these PAX2-positive, initiated cells await another insult or signal to form DASEs and eventually, tumors.

An oxidative stress mechanism mediates chelerythrine-induced heparin-binding EGF-like growth factor ectodomain shedding.

Regulated shedding of cell surface proteins is a mechanism for rapid activation of autocrine and paracrine signaling. Here we report that chelerythrine, a protein kinase C (PKC) inhibitor that possesses a variety of biological functions, is a potent inducer of heparin-binding epidermal growth factor-like growth factor (HB-EGF) shedding from the cell surface. Chelerythrine induced a time- and dose-dependent shedding of an HB-EGF-alkaline phosphatase (HB-EGF-AP) fusion protein expressed in MC2 rat prostate epithelial cells. The soluble form of HB-EGF-AP bound to heparin and exhibited potent biological activity as measured by DNA synthesis assay. Chelerythrine-induced HB-EGF shedding was metalloproteinase-(MMP-) mediated because specific MMP antagonists inhibited shedding by > or =60%. Chelerythrine stimulated production of reactive oxygen species, and antioxidants prevented chelerythrine-induced HB-EGF shedding, suggesting that the production of intracellular peroxides is necessary for this event. Consistent with this possibility, antioxidant- and MMP-inhibitable shedding was also demonstrated when hydrogen peroxide was used as an inducer. Although JNK/SAPK and p38 MAPK pathways were activated by chelerythine, these signaling mechanisms were not required to mediate the shedding event. However, JNK signaling was involved in chelerythrine-stimulated apoptosis. Our results suggest that HB-EGF shedding induced by chelerythrine is mediated predominantly via the production of reactive oxygen species.