Immunomodulatory effect of vitamin D and its potential role in the prevention and treatment of thyroid autoimmunity: a narrative review.

The main role of vitamin D is to control mineral homeostasis. However, recent studies suggested the existence of a number of extraskeletal effects. Among the latter, preclinical studies provided consistent data on the involvement of vitamin D in innate and adaptive immunity and autoimmunity. Molecular biology studies showed that both vitamin D receptor and vitamin D enzymatic complexes are expressed in a large number of cells and tissues unrelated to mineral homeostasis. In contrast, only a few randomized clinical trials in humans investigated the possible role of vitamin D in the prevention or treatment of immunological disorders. In this regard, low serum vitamin D levels have been reported in observational trials in human autoimmune disorders. The aim of the present paper was to review the potential implications of vitamin D in immune modulation, with special focus on thyroid autoimmune disorders.

Co-targeting the IGF system and HIF-1 inhibits migration and invasion by (triple-negative) breast cancer cells.

BACKGROUND: Metastatic triple-negative breast cancer is mostly incurable, due to lack of suitable drug targets. The insulin-like growth factor (IGF) system could provide such a target, and IGF-1 receptor (IGF-1R)-directed agents are already available, but seem unable to control all the complexities of the system, including crosstalk with hypoxia-inducible pathways. METHODS: Migration of triple-negative MDA-231 breast cancer cells and its modulation by IGFs, the IGF-1R inhibitor NVP-AEW541 and the IGF-2-sequestering monoclonal antibody MAB292 were assessed by the scratch wound healing and Boyden chamber assays; the effect of topotecan (inhibiting hypoxia-inducible factor-1 (HIF-1)) under hypoxia was also evaluated. Constitutive as well as drug-modulated levels of components of the IGF and HIF-1 pathways were evaluated by western blotting and qPCR. RESULTS: IGF-induced migration of MDA-231 cells was not abrogated by the IGF-1R inhibitor NVP-AEW541, whereas IGF-2 sequestration by MAB292 significantly reduced cell migration. Under hypoxia, topotecan was also effective, likely by reducing HIF-1-induced IGF-2 release. Simultaneous targeting of IGF-1R and IGF-2 or HIF-1 completely abolished cell migration. CONCLUSIONS: IR activation may account for the failure of NVP-AEW541 to suppress MDA-231 cell migration. Ligand-targeting compounds, or co-inhibition of the IGF and HIF-1 systems, may prevent activation of compensatory signalling, thereby providing a valuable addition to IGF-1R inhibitor-based therapies.

Study of in vitro and in vivo effects of the piperidine nitroxide Tempol--a potential new therapeutic agent for gliomas.

The identification of novel therapeutic agents for the management of malignant gliomas represents an area of active research. Here, we show that Tempol (4-hydroxy-2,2,6,6-tetramethylpiperidine-1-oxyl; TPL), a stable nitroxide free radical, inhibits the growth of C6 glioma cells both in vitro and in vivo. Morphological features of apoptosis were apparent in C6 cells following in vitro treatment with TPL. Cell death was preceded by dose-dependent increase in p21(WAF1/CIP1) expression, without apparent stabilisation of the TP53 gene product. When C6 cells were grown as xenografts in nude mice, treatment with TPL induced a significant dose-dependent decrease in tumour growth, without signs of general or organ toxicity. Tumours from treated mice showed an increase in the number of apoptotic cells and a decrease in the rate of neo-vascularisation compared with tumours from control mice. Our findings suggest a potential use for TPL as a novel antiproliferative agent for the treatment of malignant gliomas.

Nitroxide TEMPOL impairs mitochondrial function and induces apoptosis in HL60 cells.

The piperidine nitroxide TEMPOL induces apoptosis in a number of tumor cell lines through free radical-dependent mechanisms. As mitochondria play a major role in apoptosis as both source and target for free radicals, the present study focuses on mitochondrial effects of TEMPOL in a human promyelocytic leukemic cell line (HL-60). On 24-h exposure to TEMPOL, the following alterations were observed: 1) decrease in both the intracellular and mitochondrial glutathione pools; 2) impairment of oxidative phosphorylation; and 3) decrease in mitochondrial membrane potential. In addition, TEMPOL was found to specifically target complex I of the respiratory chain, with minor effects on complexes II and IV, suggesting that mitochondrial effects might play a role in TEMPOL-induced oxidative stress and apoptosis, and that TEMPOL might sensitize tumor cells to the pro-apoptotic effects of cytotoxic agents.

Resistance of human leukemic cell lines to 1-beta-D-arabinofuranosylcytosine: characterization of an experimental model.

1-beta-D-arabinofuranosylcytosine (ara-C) is an antimetabolite used for the treatment of acute myelogenous leukemia. The ability of ara-C to kill neoplastic cells has been correlated to the induction of apoptosis. The clinical use of ara-C is limited by the development of drug resistance. Alterations in drug-induced apoptosis play a critical role in ara-C resistance. In particular, the proto-oncogene bcl-2 has been implicated in this phenomenon. To better understand the molecular basis of the role of bcl-2 in ara-C resistance, we investigated the relationship between the cytotoxic effect of ara-C, the expression levels and the subcellular localization of bcl-2 in three human leukemic cell lines (HL-60, KG1, J111). We have also evaluated the effects of ara-C on the J111 leukemic cell line (showing the lowest levels of Bcl-2 and the highest sensitivity to ara-C) overexpressing the bcl-2 oncogene. The model we developed here will allow further studies on the role of post-translational events involving bcl-2 (such as translocation and/or phosphorylation) in the cellular response to ara-C treatment.

The nitroxide tempol induces oxidative stress, p21(WAF1/CIP1), and cell death in HL60 cells.

The antiproliferative effect of Tempol, a stable nitroxide free radical, was investigated on the p53-negative human leukemia cell line HL60. A concentration- and time-dependent inhibition of cell growth was observed that appears to be due to induction of apoptosis. Involvement of oxidative stress is indicated by a concentration-dependent increase in intracellular peroxides and a parallel decrease in total cellular glutathione; in addition, increased survival rates were observed in cells simultaneously treated with Tempol and the antioxidant N-acetylcysteine. Tempol did not affect the relative levels of Bax and Bcl2, whereas p21(WAF1/CIP1) was enhanced in a concentration- and time-dependent fashion; this effect was partially inhibited by N-acetylcysteine, was maintained for up to 8 h after Tempol removal, and seemed to depend on continuing protein synthesis. The increase in p21(WAF1/CIP1) was accompanied by a parallel accumulation of cells in the G(1) phase of the cycle and by a decrease in the 110 kDa form of pRb. Our results suggest that p53-independent induction of p21(WAF1/CIP1) mediates the antiproliferative effect of Tempol; on the basis of this observation, the nitroxide could be proposed as an useful adjunct to the treatment of p53-deficient tumors, which are often refractory to standard chemotherapy.

Antiproliferative effect of the piperidine nitroxide TEMPOL on neoplastic and nonneoplastic mammalian cell lines.

The stable nitroxide 4-hydroxy-2,2,6,6-tetramethylpiperidine-N-oxyl (TEMPOL) is widely used as a probe in biophysical studies and as an antioxidant in several experimental models. The potential cytotoxic effects of TEMPOL were tested on a panel of human and rodent cell lines, and the nitroxide proved to be significantly more effective in inhibiting the growth of neoplastic than nonneoplastic cell lines after a 96-h exposure. More detailed studies on MCF-7/WT cells indicate that at least 24 h are necessary for TEMPOL to induce irreversible cell damage, which seems to be related to the reactivity of the nitroxyl group. This observation, together with the antagonistic effect of N-acetylcysteine, suggests an involvement of free radical-mediated processes. Cell cycle studies indicate a biphasic effect of TEMPOL, with a short-term accumulation of the cells in the G1 phase and a later increase in G2/M phase; the pattern of DNA fragmentation observed in TEMPOL-treated cells points to an apoptotic mode of cell death. In conclusion, our data suggest that, while the possible cytotoxic effects of TEMPOL should not be overlooked when using this compound as a biophysical probe or antioxidant, these same properties could be exploited as a novel approach to cancer chemotherapy, especially in tumor cells exhibiting unfavorable characteristics, such as a multidrug-resistant phenotype or loss of hormone receptors.

Characterization of a clonal human colon adenocarcinoma line intrinsically resistant to doxorubicin.

Intrinsic low-level resistance to anti-cancer drugs is a major problem in the treatment of gastrointestinal malignancies. To address the problem presented by intrinsically resistant tumours, we have isolated two monoclonal lines from LoVo human colon adenocarcinoma cells: LoVo/C7, which is intrinsically resistant to doxorubicin (DOX); and LoVo/C5, which shows the same resistance index for DOX as the mixed parental cell population. For comparison, we have included in the study a LoVo-resistant line selected by continuous exposure to DOX and expressing a typical multidrug resistant (MDR) phenotype. In these cell lines we have studied the expression and/or activity of a number of proteins, including P-glycoprotein 170 (P-gp), multidrug resistance-associated protein (MRP), lung resistance-related protein (LRP), glutathione (GSH)-dependent enzymes and protein kinase C (PKC) isoforms, which have been implicated in anti-cancer drug resistance. Intracellular DOX distribution has been assessed by confocal microscopy. The results of the present study indicate that resistance in LoVo/C7 cells cannot be attributed to alterations in P-gp, LRP or GSH/GSH-dependent enzyme levels. Increased expression of MRP, accompanied by alterations in the subcellular distribution of DOX, has been observed in LoVo/C7 cells; changes in PKC isoform pattern have been detected in both intrinsically and pharmacologically resistant cells.