Dp44mT targets the AKT, TGF-ß and ERK pathways via the metastasis suppressor NDRG1 in normal prostate epithelial cells and prostate cancer cells.

BACKGROUND: Effective treatment of prostate cancer should be based on targeting interactions between tumour cell signalling pathways and key converging downstream effectors. Here, we determined how the tumourigenic phosphoinositide 3-kinase/protein kinase B (PI3K/AKT), tumour-suppressive phosphatase and tensin homologue deleted on chromosome 10 (PTEN) and transforming growth factor-ß (TGF-ß) pathways are integrated via the metastasis suppressor, N-myc downstream-regulated gene-1 (NDRG1). Moreover, we assessed how the novel anti-tumour agent, Dp44mT, may target these integrated pathways by increasing NDRG1 expression. METHODS: Protein expression in Dp44mT-treated normal human prostate epithelial cells and prostate cancer cells (PC-3, DU145) was assessed by western blotting. The role of NDRG1 was examined by transfection using an NDRG1 overexpression vector or shRNA. RESULTS: Dp44mT increased levels of tumour-suppressive PTEN, and decreased phosphorylation of ERK1/2 and SMAD2L, which are regulated by oncogenic Ras/MAPK signalling. Importantly, the effects of Dp44mT on NDRG1 and p-SMAD2L expression were more marked in prostate cancer cells than normal prostate epithelial cells. This may partly explain the anti-tumour selectivity of these agents. Silencing NDRG1 expression increased phosphorylation of tumourigenic AKT, ERK1/2 and SMAD2L and decreased PTEN levels, whereas NDRG1 overexpression induced the opposite effect. Furthermore, NDRG1 silencing significantly reduced the ability of Dp44mT to suppress p-SMAD2L and p-ERK1/2 levels. CONCLUSION: NDRG1 has an important role in mediating the tumour-suppressive effects of Dp44mT in prostate cancer via selective targeting of the PI3K/AKT, TGF-ß and ERK pathways.

Photoprotection by 1alpha,25-dihydroxyvitamin D and analogs: further studies on mechanisms and implications for UV-damage.

Ultraviolet (UV) irradiation causes DNA damage in skin cells, immunosuppression and photocarcinogenesis. 1alpha,25-dihydroxyvitamin D3 (1,25D) reduces UV-induced DNA damage in the form of cyclobutane pyrimidine dimers (CPD) in human keratinocytes in culture and in mouse and human skin. UV-induced immunosuppression is also reduced in mice by 1,25D, in part due to the reduction in CPD and a reduction in interleukin (IL-6. The cis-locked analog, 1alpha,25-dihydroxylumisterol3 (JN), which has almost no transactivating activity, reduces UV-induced DNA damage, apoptosis and immunosuppression with similar potency to 1,25D, consistent with a non-genomic signalling mechanism. The mechanism of the reduction in DNA damage in the form of CPD is unclear. 1,25D doubles nuclear expression of p53 compared to UV alone, which suggests that 1,25D facilitates DNA repair. Yet expression of a key DNA repair gene, XPG is not affected by 1,25D. Chemical production of CPD has been described. Incubation of keratinocytes with a nitric oxide donor, SNP, induces CPD in the dark. We previously reported that 1,25D reduced UV-induced nitrite in keratinocytes, similar to aminoguanidine, an inhibitor of nitric oxide synthase. A reduction in reactive nitrogen species has been shown to facilitate DNA repair, but in view of these findings may also reduce CPD formation via a novel mechanism.

Detection of urine and blood clenbuterol following short-term oral administration in the horse.

BACKGROUND AND AIM: The pharmacokinetics of clenbuterol in equine urine and blood was investigated. MATERIAL AND METHODS: Urine and blood samples were collected following 3-day multiple oral administrations. The samples were examined using enzyme-linked immunosorbent assay and further confirmed by solid phase extraction and capillary electrophoresis. RESULTS: Urinary clenbuterol was detectable until day 14 after the last dose. The urinary excretion of clenbuterol was characterized by a biphasic pattern. The half-lives of the bi-exponential elimination (t(1/2alpha) and t(1/2beta)) for urinary clenbuterol were about 12.1 and 48 hours. After a single oral administration (4 microg/kg) of clenbuterol, the half-life of serum clenbuterol was approximately 11.4 hours.

In vivo relevance for photoprotection by the vitamin D rapid response pathway.

Vitamin D is produced by exposure of 7-dehydrocholesterol in the skin to UV irradiation (UVR) and further converted in the skin to the biologically active metabolite, 1,25-dihydroxyvitamin D(3) (1,25(OH)(2)D(3)) and other compounds. UVR also results in DNA damage producing cyclobutane pyrimidine dimers (CPD). We previously reported that 1,25(OH)(2)D(3) at picomolar concentrations, protects human skin cells from UVR-induced apoptosis, and decreases CPD in surviving cells. 1,25(OH)(2)D(3) has been shown to generate biological responses via two pathways-the classical steroid receptor/genomic pathway or a rapid, non-genomic pathway mediated by a putative membrane receptor. Whether the rapid response pathway is physiologically relevant is unclear. A cis-locked, rapid-acting agonist 1,25(OH)(2)lumisterol(3) (JN), entirely mimicked the actions of 1,25(OH)(2)D(3) to reduce fibroblast and keratinocyte loss and CPD damage after UVR. The effects of 1,25(OH)(2)D(3) were abolished by a rapid-acting antagonist, but not by a genomic antagonist. Skh:hr1 mice exposed to three times the minimal erythemal dose of solar-simulated UVR and treated topically with 1,25(OH)(2)D(3) or JN immediately after UVR showed reduction in UVR-induced UVR-induced sunburn cells (p<0.01 and <0.05, respectively), CPD (p<0.01 for both) and immunosuppression (p<0.001 for both) compared with vehicle-treated mice. These results show for the first time an in vivo biological response mediated by a rapid-acting analog of the vitamin D system. The data support the hypothesis that 1,25(OH)(2)D(3) exerts its photoprotective effects via the rapid pathway and raise the possibility that other D compounds produced in skin may contribute to the photoprotective effects.

Skin cancer prevention: a possible role of 1,25dihydroxyvitamin D3 and its analogs.

We previously reported that the natural hormone 1,25dihydroxyvitamin D3 (1,25(OH)(2)D(3)) protects human skin cells from ultraviolet radiation (UVR)-induced apoptosis. UVR-induced pre-mutagenic cyclobutane pyrimidine dimers are diminished in number from 0.5h after cessation of UVR in all skin cell types, by treatment with three different Vitamin D compounds: by 1,25(OH)(2)D(3), by the rapid acting, low calcemic analog, 1alpha,25(OH)(2)lumisterol(3) (JN) and by the low calcemic but transcriptionally active hybrid analog 1alpha-hydroxymethyl-16-ene-24,24-difluoro-25-hydroxy-26,27-bis-homovitamin D3 QW-1624F2-2 (QW), which may explain the enhanced cell survival. The rapid response antagonist analog 1beta,25(OH)(2)D(3) (HL) abolished the photoprotective effects of 1,25(OH)(2)D(3) whilst a genomic antagonist, (23S)-25-dehydro-1alpha-hydroxyvitamin D(3)-26,23-lactone (TEI-9647), had no effect. UVR increased p53 expression in human skin cells, whilst concurrent treatment with 1,25(OH)(2)D(3) further enhanced this effect several fold, at 3 and 6h after UVR. Combined with previously reported lower nitrite levels with 1,25(OH)(2)D(3), this increased p53 expression may favor DNA repair over apoptosis. We now report that topical application of 1,25(OH)(2)D(3) or QW also suppressed solar simulated UV (SSUVR-induced pyrimidine dimers in the epidermis of irradiated hairless Skh:HR1 mice, measured 24h after irradiation. Furthermore, UVR-induced immunosuppression in the mice was markedly reduced by topical application of either 1,25(OH)(2)D(3) or QW. These preliminary results show, for the first time, a protective effect of Vitamin D compounds against DNA photodamage in vivo.

1,25-Dihydroxyvitamin D and three low-calcemic analogs decrease UV-induced DNA damage via the rapid response pathway.

1,25-Dihydroxyvitamin D(3) [1,25(OH)(2)D(3)] is anti-apoptotic in human keratinocytes, melanocytes and fibroblasts after ultraviolet (UV)-exposure. To date, there is no published data on the effects of 1,25(OH)(2)D(3) or its analogs on DNA damage in irradiated skin cells. In these skin cells, 24h pre-treatment with 1,25(OH)(2)D(3) dose-dependently (10(-12) to 10(-8)M) decreased CPD damage by up to 60%. This photoprotective effect was also seen if the 1,25(OH)(2)D(3) was added immediately after irradiation and was mimicked by QW-1624F2-2 (QW), a low-calcemic 1beta-hydroxymethyl-3-epi-16-ene-24,24-difluoro-26,27-bis homo hybrid analog. The well-studied low calcemic, rapid acting agonist analogs 1alpha,25(OH)(2)lumisterol(3) (JN) and 1alpha,25(OH)(2)-7-dehydrocholesterol (JM) also protected skin cells from UV-induced cell loss and CPD damage to an extent comparable with that of 1,25(OH)(2)D(3). In contrast, the rapid response antagonist analog 1beta,25(OH)(2)D(3) (HL) completely abolished the photoprotective effects (reduced cell loss and reduced CPD damage) produced by treatment with 1,25(OH)(2)D(3), JN, JM and QW. Evidence for involvement of the nitric oxide pathway in the protection from CPD damage by 1,25(OH)(2)D(3) was obtained. These data provide further evidence for a role of the vitamin D pathway in the intrinsic skin defenses against UV damage. The data also support the hypothesis that the photoprotective effects of 1,25(OH)(2)D(3) are mediated via the rapid response pathway(s).

Oppressive limits: Callahan's foundation myth.

Daniel Callahan has not simply proposed alterations of important features of the health economy. He has constructed a blue print for society drawing on concepts of what is natural and appropriate to human beings. He is, in effect, establishing a new social order. Like any social order, Callahan's system has its justificatory schemes or founding myths. This paper offers a feminist examination of the functions that these four myths-the concept of a whole of life; the stages of life; a tolerable death; and a reconstruction of the meaning of the aged in terms of sacrifice-fulfill in Callahan's new social order. Callahan's concept of a whole of life reflects the power he assigns to nature, and the futility and harm he associates with attempts to repudiate biological imperatives. It introduces the stages of human life, tolerable death, and aging. The paper critically examines these concepts.

The role of the hospital in the evolution of elite cultures of medicine.

Rosenberg argues that a culture of medicine articulates a unique medical vision reflecting scientific ideologies and perceptions, professional values and rewards, career patterns, and the work context of each generation of physicians (Rosenberg, 1987, p. 7). It might be more accurate, however, to recognize multiple cultures of medicine. This paper traces American hospitals' contributions to three specific aspects of certain elite cultures of medicine. Its first section introduces and explores these aspects transhistorically. They are: a stratified paradigm of the physician-patient relationship, the equation of competence with specialization, and a pattern within a stratified model of medicine called the "Lone Ranger syndrome." A brief historical profile subsequently highlights changes in the structure and function of American hospitals in the 19th century. The three remaining sections chart the ebb and flow of the prominence and power of these specific aspects of elite cultures of medicine across three historical periods: the late 19th century, 1910-1930, and the post World War II era.