A Limited Role for Retinoic Acid and Retinoic Acid Receptors RARα and RARβ in Regulating Keratin 19 Expression and Keratinization in Oral and Epidermal Keratinocytes

Different types of stratified squamous epithelia—for example, the “orthokeratinized” epidermis, the “parakeratinized” gingiva, and the “nonkeratinized” oral lining mucosal epithelia—are formed by intrinsically distinct keratinocyte subtypes. These subtypes exhibit characteristic patterns of keratin protein expression in vivo and in culture. Keratin 19 is an informative subtype-specific marker because the basal cells of only nonkeratinizing epithelia express K19 in vivo and in culture. Epidermal keratinocytes normally do not express K19, but can be induced to do so in culture by retinoic acid (RA). Keratinocyte subtypes express the retinoic acid receptor (RAR)β at levels roughly correlated with their level of K19 expression in culture and their potential for forming a nonkeratinized epithelium in vivo. We tested the hypothesis that the level of RARβ expressed by a keratinocyte determines its K19 expression and its form of suprabasal differentiation. Normal human epidermal and gingival keratinocytes stably overexpressing either RARβ or RARα were generated by defective retroviral transduction. Overexpression of either receptor enhanced the RA inducibility of K19 in conventional culture, in that the proportion of the transductants becoming K19+ in response to RA was markedly increased compared with controls. The pattern of differentiation of the epithelium formed in organotypic culture, assessed by basal K19 and suprabasal K1, K4, and filaggrin expression, however, was unaltered by PAR overexpression. Thus, the susceptibility of keratinocytes to regulation of K19 expression by retinoids is conditional, and levels of neither RARβ nor RARα are limiting to the intrinsic mechanism that specifies alternate differentiation pathways for stratified squamous epithelia

NF-κB Inhibition through Proteasome Inhibition or IKKβ Blockade Increases the Susceptibility of Melanoma Cells to Cytostatic Treatment through Distinct Pathways

Metastasized melanoma is almost universally resistant to chemotherapy. Given that constitutive or drug-induced upregulation of NF-κB activity is associated with this chemoresistance, NF-κB inhibition may increase the susceptibility to antitumoral therapy. On the cellular level, two principles of NF-κB inhibition, proteasome inhibition by bortezomib and IκB kinase-β (IKKβ) inhibition by the kinase inhibitor of NF-κB-1 (KINK-1), significantly increased the antitumoral efficacy of camptothecin. When combined with camptothecin, either of the two NF-κB-inhibiting principles synergistically influenced progression-related in vitro functions, including cell growth, apoptosis, and invasion through an artificial basement membrane. In addition, when C57BL/6 mice were intravenously injected with B16F10 melanoma cells, the combination of cytostatic treatment with either of the NF-κB-inhibiting compounds revealed significantly reduced pulmonary metastasis compared to either treatment alone. However, on the molecular level, nuclear translocation of p65, cell cycle analysis, and expression of NF-κB-dependent gene products disclosed distinctly different molecular mechanisms, resulting in the same functional effect. That proteasome inhibition and IKKβ inhibition affect distinct molecular pathways downstream of NF-κB, both leading to increased chemosensitivity, is previously unreported. Thus, it is conceivable that switching the two principles of NF-κB inhibition, once resistance to one of the agents occurs, will improve future treatment regimens

Inositoylated Platelet-Activating Factor (Ino-C2-PAF) Modulates Dynamic Lymphocyte–Endothelial Cell Interactions and Alleviates Psoriasis-Like Skin Inflammation in Two Complementary Mouse Models

Psoriasis, a tumor necrosis factor alpha (TNFα)-governed inflammatory disorder with prominent dysregulation of cutaneous vascular functions, has evolved into a model disorder for studying anti-inflammatory therapies. We present experimental in vitro and in vivo data on 1-O-octadecyl-2-O-(2-(myo-inositolyl)-ethyl)-sn-glycero-3-(R/S)-phosphatidyl-choline (Ino-C2-PAF), the lead compound of a class of synthetic glycosylated phospholipids, in anti-inflammatory therapy. Ino-C2-PAF strongly induced apoptosis only in TNFα-stimulated, but not in untreated human vascular endothelial cells. Moreover, TNFα-induced endothelial adhesion molecules that mediated the rolling and firm adhesion of leukocytes (vascular cell adhesion protein-1 (VCAM-1), E-selectin, and ICAM-1) were selectively downregulated by Ino-C2-PAF. Similarly, expression of L-selectin, VCAM-1 receptor α4β1 integrin, and lymphocyte function-associated antigen-1 on human peripheral blood mononuclear cells was reduced without induction of apoptosis. Functionally, these changes were accompanied by significant impairment of rolling and adhesion of human peripheral blood lymphocytes on TNFα-activated endothelial cells in a dynamic flow chamber system. When the therapeutic potential of Ino-C2-PAF was assessed in two complementary mouse models of psoriasis, K5.hTGFβ1 transgenic and JunB/c-Jun-deficient mice, Ino-C2-PAF led to significant alleviation of the clinical symptoms and normalized the pathological cutaneous changes including vascularization. There were no overt adverse effects. These findings suggested that Ino-C2-PAF is a potential candidate in the therapy of inflammatory skin diseases that include abnormal vascular functions

INTS6/DICE1 inhibits growth of human androgen-independent prostate cancer cells by altering the cell cycle profile and Wnt signaling

<p>Abstract</p> <p>Background</p> <p>The gene encoding integrator complex subunit 6 (<it>INTS6</it>), previously known as deleted in cancer cells 1 (<it>DICE1</it>, OMIM 604331) was found to be frequently affected by allelic deletion and promoter hypermethylation in prostate cancer specimens and cell lines. A missense mutation has been detected in prostate cancer cell line LNCaP. Together, these results suggest <it>INTS6/DICE1 </it>as a putative tumor suppressor gene in prostate cancer. In this study, we examined the growth inhibitory effects of <it>INTS6/DICE1 </it>on prostate cancer cells.</p> <p>Results</p> <p>Markedly decreased <it>INTS6/DICE1 </it>mRNA levels were detected in prostate cancer cell lines LNCaP, DU145 and PC3 as well as CPTX1532 as compared to a cell line derived from normal prostate tissue, NPTX1532. Exogenous re-expression of <it>INTS6/DICE1 </it>cDNA in androgen-independent PC3 and DU145 cell lines substantially suppressed their ability to form colonies <it>in vitro</it>. This growth inhibition was not due to immediate induction of apoptosis. Rather, prostate cancer cells arrested in G1 phase of the cell cycle. Expression profiling of members of the Wnt signaling pathway revealed up-regulation of several genes including disheveled inhibitor CXXC finger 4 (<it>CXXC4</it>), frizzled homologue 7 (<it>FZD7</it>), transcription factor 7-like 1 (<it>TCF7L1</it>), and down-regulation of cyclin D1.</p> <p>Conclusion</p> <p>These results show for the first time a link between <it>INTS6/DICE1 </it>function, cell cycle regulation and cell-cell communication involving members of the Wnt signaling pathway.</p