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Background: Cucurbitacin (Cuc) and triterpene-derived natural products exhibit anti-cancer potential in addition to their conspicuous anti-bacterial and anti-inflammatory activity. Recently, inhibition of Janus kinase 2 (JAK2)/signal transducer and activator of transcription 3 (STAT3) signaling was shown to underlie the effects of Cuc family on inducing cell death in cancer.Method: We purified Cuc IIa, the active component from the medicinal plant Hemsleya amalils Diels, which shows different structural modifications from other Cuc derivatives. We investigated the mechanisms of its inhibitory effects on cancer cells in vitro and tumour growth in vivo.Results: Cuc IIa induced the irreversible clustering of filamentous actin and arrested cell cycle by the increases in G2/M populations. Cuc IIa resulted in the reduced phospho-Histone H3 and markedly increased cleavage of poly-(ADP-ribose) polymerase or PARP, immediate upstream of DNA breakdown as the result of caspase activation, consistent with mitotic blockage-induced cell death. However, unlike other Cuc members, Cuc IIa did not suppress JAK2/STAT3 phosphorylation or alter phosphorylation of mitogen-activated protein kinases. Instead, the expression of the cell cycle-regulated Inhibitor of Apoptosis Protein (IAP) survivin was reduced. Introducing oncoprotein δ-catenin, which increased survivin expression and suppressed small GTPase RhoA, reduced efficacy of Cuc IIa to induce cell death. Supporting the effects of Cuc IIa on actin cytoskeletal signaling, RhoA phosphorylation was reduced suggesting its increased activity.Conclusion: Cuc IIa is a novel class of anti-cancer drug in suppression of cancer cell expansion by disrupting the actin cytoskeleton and directing the cell to undergo PARP-mediated apoptosis through the inhibition of survivin downstream of JAK2/STAT3. [ABSTRACT FROM AUTHOR]

The fibroblast-like cells in the marrow stromal system were separated from endothelial cells and macrophages by negative selection of magnetic beads. Immunocytochemistry confirmed that these fibroblast-like cells expressed fibronectin and collagen Type III, but not Factor VIII and epithelial membrane antigen (endothelial cell markers) or Mac I (macrophage marker). The fibroblast-like stromal cells (FSC) synthesized the insulin-like growth factors (IGF)-I and -II in amounts equivalent to that produced by unfractionated marrow stromal cells (UMSC); in both, the concentration of IGF-II was 10 times higher than that of IGF-I. Northern analysis revealed that FSC and UMSC expressed identical patterns of mRNAs for IGF-I and transforming growth factor (TGF) -beta 2, for osteopontin, and for procollagen Types I and III (Type I > Type III). Type II procollagen mRNA was not expressed in both cell populations. The TGF-beta 2 gene mRNA was expressed at a lower level by the FSC than UMSC. The pattern of gene expression in these cells is consistent with an osteoprogenitor phenotype. Both FSCs and UMSCs express parathyroid hormone (PTH) and estrogen receptor genes (rtPCR technique). The study provides additional evidence that fibroblast-like marrow stromal cells have an osteoblast signature, and that they are largely responsible for the osteogenic performance of cells in unfractionated marrow. [ABSTRACT FROM AUTHOR]

Fibroblast-like rat marrow stromal cell (CFU-F) cultures have been characterized in terms of their responsiveness to calciotropic hormones, metal ions, the nonsteroidal antiinflammatory drug, and by their putative paracrine role in the maintenance of active populations of osteoblasts at the marrow-bone interface. These studies indicate that CFU-Fs lack a complete osteoblast signature. Subconfluent CFU-Fs grown in the presence or absence of 10(-7) M dexamethasone lack receptors for PTH and calcitonin, and fail to show enhanced cAMP or cGMP responses to 10(-7) M 1-34 PTH (rat), or any evidence of osteocalcin production [+/- 10(-9) M 1,25-(OH)2D3]. Low concentrations of fluoride [10(-12) and 10(-9) M] stimulated CFU-F grown in vitro in serum-free media, though higher levels (10(-7) and 10(-6) M), inhibited growth in vivo and in vitro. Aluminum (10(-12)-10(-7) M) and ibuprofen (10(-7) M) did not alter normal growth patterns, indicating an action on bone cells more differentiated than CFU-Fs. Serum-free conditioned medium (CM) from control and ovariectomized (OVX)/OVX+ dihydrotachysterol-Rx rat CFU-F cultures was mitogenic for neonatal rat calvarial osteoblasts in vitro, but not for ROS 17/2.8 cells. The studies affirm the mesenchymal-like character of CFU-Fs and project their significant role in sustaining functional endosteal osteogenic cell populations. [ABSTRACT FROM AUTHOR]