Use of Human Epidermal Cells in the Study of Carcinogenesis

Because of the importance of human cells, particularly human epithelial cells, in cancer research, we have studied certain phases or events of carcinogenesis using human epidermal cells in primary culture. 1) We found that human epidermal cells are capable of metabolizing benzo[a]pyrene. Large inter-individual variations are found in the basal and induced arylhydrocarbon-hydroxylase activities. 2) UV-in-duced unscheduled DNA synthesis was demonstrated in human epidermal cells on autoradiographs. We also found that DNA repair is defective in epidermal cells isolated from xeroderma pigmentosum by a new explant-outgrowth culture. 3) Human epidermal cells are unique in that there is a large number of binding sites to phorbol esters compared with mouse epidermal cells, but there is no down-regulation. Further, human epidermal cells show essentially negative responses to tumor promoters, i.e., no stimulation of DNA synthesis, sugar uptake, and no induction of ornithine decarboxylase activity. 4) Human epidermal cells contain 1.5 × 105 binding sites per cell for epidermal growth factor (EGF), whereas squamous cell carcinomas of skin and oral cavity have larger amounts of EGF receptors in the order of 106 per cell. 5) Based on the above results, we attempted to transform human epidermal cells by the treatment with chemical carcinogens, but until now no transformation was obtained. J Invest Dermatol 92:271S–274S, 198

Protein Kinase C α Associates with Phospholipase D1 and Enhances Basal Phospholipase D Activity in a Protein Phosphorylation-Independent Manner in Human Melanoma Cells

It is well known that phospholipase D plays a crucial part in the signal transduction of many types of cells, and is activated by protein kinase C α when cells are stimulated. To elucidate the role of phospholipase D in melanoma, the expression of phospholipase D1 and protein kinase C α in primary and metastatic lesions of acral lentiginous melanoma and superficial spreading melanoma was investigated using immunohistologic techniques. In addition, the mechanism of regulation of phospholipase D1 by protein kinase C α was examined in a human melanoma cell line HM3KO using an adenovirus-mediated gene transfer technique. Both phospholipase D1 and protein kinase C α were strongly expressed in primary and metastatic lesions of superficial spreading melanoma. Conversely, in acral lentiginous melanoma lesions, the expression of these two proteins increased dramatically with tumor progression; the expression of both phospholipase D1 and protein kinase C α was almost negative in the radial growth phase of primary acral lentiginous melanoma lesions, and increased synchronously in a progression-related manner in advanced acral lentiginous melanoma lesions, including vertical growth phase and metastatic lesions. Immunoprecipitation study showed that phospholipase D1 and protein kinase C α are associated physiologically in resting melanoma cells. Further immunoprecipitation study using HM3KO cells after adenovirus-mediated simultaneous overexpression of phospholipase D1 and protein kinase C α, or phospholipase D1 and the kinase-negative mutant of protein kinase C α revealed that both protein kinase C α and the kinase-negative mutant of protein kinase C α are associated with phospholipase D1 in melanoma cells in the absence of an external signal. Overexpression of protein kinase C α or the kinase-negative mutant of protein kinase C α in melanoma cells by the adenovirus vectors resulted in the enhancement of basal phospholipase D activity in a viral concentration-dependent manner. Furthermore, enhanced basal phospholipase D activity increased the in vitro invasive potential of HM3KO cells. These results suggest that upregulation of phospholipase D1 and protein kinase C α plays a part in the progression of acral lentiginous melanoma from the radial growth phase to the vertical growth phase. The present results also suggest that protein kinase C α associates with phospholipase D1 and enhances basal phospholipase D activity in a protein phosphorylation-independent manner in melanoma cells, which contributes to the cell's high invasive potential

A novel type of binding specificity to phospholipids for rat mannose-binding proteins isolated from serum and liver

AbstractMannose-binding protein (MBP) belongs to the collectin subgroup of C-type lectins with specificity for mannose and N-acetylglucosamine sugars. We investigated whether rat MBPs isolated from serum (S-MBP) and liver (L-MBP) interact with phospholipids using antibody against each MBP. Both S- and L-MBPs bound to phosphatidylinositol coated onto microtiter wells in a concentration- and a Ca2+-dependent manner. L-MBP also bound to phosphatidylglycerol and weakly to phosphatidylserine. MBPs interacted with liposomes composed of these lipids. S- and L-MBPs bound to phosphatidylinositol 4-monophosphate. L-MBP also bound to cardiolipin. These results provide evidence for a novel type of ligand binding specificity for MBPs, and raise the possibility that phospholipids are ligands for collectins

PKCα mediates TGFβ-induced growth inhibition of human keratinocytes via phosphorylation of S100C/A11

Growth regulation of epithelial cells is of major concern because most human cancers arise from them. We demonstrated previously a novel signal pathway involving S100C/A11 for high Ca2+-induced growth inhibition of normal human keratinocytes (Sakaguchi, M., M. Miyazaki, M. Takaishi, Y. Sakaguchi, E. Makino, N. Kataoka, H. Yamada, M. Namba, and N.H. Huh. 2003. J. Cell Biol. 163:825–835). This paper addresses a question whether transforming growth factor β (TGFβ) shares the pathway with high Ca2+. On exposure of the cells to TGFβ1, S100C/A11 was phosphorylated, bound to nucleolin, and transferred to the nucleus, resulting in induction of p21WAF1/CIP1 and p15INK4B through activation of Sp1. Protein kinase C α (PKCα) was shown to phosphorylate 10Thr of S100C/A11, which is a critical event for the signal transduction. The TGFβ1-induced growth inhibition was almost completely mitigated when PKCα activity was blocked or when S100C/A11 was functionally sequestered. These results indicate that, in addition to the well-characterized Smad-mediated pathway, the PKCα–S100C/A11-mediated pathway is involved in and essential for the growth inhibition of normal human keratinocytes cells by TGFβ1

A Radiological Study of Rapidly Destructive Coxarthropathy

Rapidly destructive coxarthropathy (RDC) is a clinical concept propounded by Postel and Kerboull. RDC is characterized by joint destruction progression within a year, although the etiology of this disorder remains unknown. We evaluated 21 hips in 20 patients radiologically diagnosed with RDC. All patients underwent a total hip arthroplasty. The average age at surgery was 75 years. The affected side was more osteoporotic in all patients, and the pelvic angle, which indicates the spinopelvic alignment, was distributed below the normal range, i.e., the posterior tilt was more than the normal range. The affected side showed a higher center-edge (CE) angle and anterior-acetabular head index (AAHI) than the unaffected side, possibly due to severe head collapse. Our result supported that osteoporosis and/or mechanical factors influence the course of RDC. More investigations such as biochemical and immunopathological analyses would be necessary to clarify the etiology of RDC, which could be a terminal stage of some lesions

(N.N. Pcific Cruise) Northern North Pacific Ocean

航海番号: KH-74-2 ; 航海日程: April 30 - June 26, 197