Synthesis of vitamin E and aliphatic lipid vanadium(IV) and (V) complexes, and their cytotoxic properties.

Novel vitamin E chelate derivatives and their VIV/V complexes have been synthesized and characterized, and their anticancer properties have been evaluated. The new complexes have been designed to exhibit enhanced cytotoxicity by combining high lipophilicity with the properties of vanadium to induce the formation of reactive oxygen species (ROS). In particular, the ß-tocopherol derivatives with iminodiethanol (ß-tocDEA) and dipicolylamine (ß-tocDPA) as well their VV and VIV complexes, [VVO(ß-tocDEA] and [VIVO(ß-tocDPA] have been synthesized and characterized by Nuclear Magnetic Resonance (NMR), Ultra Violet-Visible (UV-Vis) and Electron Paramagnetic Resonance (EPR) spectroscopies. Although the ß-tocopherol compounds exhibit antioxidant activity their complexes induce formation of radicals. In addition, two vanadium amphiphilic complexes of 2,2'-((2-hydroxyoctadecyl)azanediyl)bis(ethan-1-ol) (C18DEA) and 1-(bis(pyridin-2-ylmethyl)amino)octadecan-2-ol (C18DPA) known to activate O2 and produce ROS were synthesized and characterized (C. Drouza, A. Dieronitou, I. Hadjiadamou, M. Stylianou, J. Agric. Food. Chem., vol. 65, 2017, pp. 4942-4951). The four amphiphilic vanadium complexes exhibit enhanced hydrolytic stability. All compounds found to be cytotoxic for cancer cells exhibiting activity similar or higher to cis-platin.

Effects of secondary biological treatment plant effluent administration, as drinking water, to rats' urogenital system in relation to cadmium and lead accumulation.

The aim of this study was to examine the effect of the secondary biological treatment plant effluent administration on the kidneys, urinary bladder, and testis of Wistar rats in relation to lead (Pb) and cadmium (Cd) accumulation, since such an effluent is used for irrigation of edible plants. Male Wistar rats, randomly assigned into 5 groups, were treated with domestic sewage effluent (DSE) for 24 months. Cadmium and lead concentrations in the DSE, rats' tissues, and urine were estimated by means of atomic spectroscopy. Lead was rapidly accumulated in high amounts in rats' kidney and to a lesser extent in the testis whereas Cd concentration was raised in all tissues examined. Deposition of Cd and Pd in the kidney of the rats resulted in profound damage over time. The results showed that long-term administration to DSE as drinking water exposes living organisms to urogenital stress related to heavy metal concentration and pH of the effluent.

Effects of α-Lipoic Acid, Carnosine, and Thiamine Supplementation in Obese Patients with Type 2 Diabetes Mellitus: A Randomized, Double-Blind Study.

Type 2 diabetes mellitus (T2DM) is evolving to an epidemic of the modern world. T2DM is associated with a number of pathological complications, including cardiovascular disease that is mostly promoted by the increased oxidative stress in type 2 diabetic patients. We performed a randomized double-blind placebo-controlled trial to investigate the effectiveness of an individualized oral supplementation with α-lipoic acid (ALA), carnosine, and thiamine. For that purpose, 82 obese type 2 diabetic patients were randomly assigned to 2 groups, and were either supplemented daily with 7 mg ALA/kg body weight, 6 mg carnosine/kg body weight, and 1 mg thiamine/kg body weight or placebo for 8 weeks. An array of biochemical tests including the estimation of oxidative stress and platelet aggregation were performed at baseline and at follow-up. Moreover, the antiplatelet activity of each of the supplement's components was determined ex vivo at human and washed rabbit platelets. Glucose and HbA1c levels were significantly reduced after supplementation (135.7 ± 19.5 mg/dL vs. 126.5 ± 16.8 mg/dL and 8.3% ± 0.3% vs. 6.03% ± 0.58%, respectively, P < .05); however, insulin was significantly increased (3.6 ± 0.7 µIU/mL vs. 6.8 ± 0.2 µIU/mL, P < .05). The patients treated with the supplement recorded higher follow-up values for HOMA-IR and HOMA-ß, and a significant drop in serum hydroperoxide level. Only ALA inhibited platelets aggregation ex vivo through ADP, platelet activating factor, arachidonic acid, epinephrine, collagen, and thrombin pathways. Daily supplementation with an individualized ALA, carnosine, and thiamine supplement effectively reduced glucose concentration in type 2 diabetic patients, probably by increasing insulin production from the pancreas. In addition to that, the reduction of oxidative stress and inhibition of platelet aggregation could potentially provide greater cardiovascular protection. Further studies are needed to fine-tune the supplementation dose-response effects in T2DM patients.

Zinc inhibits apoptosis and maintains NEP downregulation, induced by ropivacaine, in HaCaT cells.

Zinc (Zn), a cell-protective metal against various toxic compounds, is the key agent for neutral endopeptidase (NEP) functional structure. NEP is a zinc metalloenzyme which degrades endogenous opioids and is expressed in human keratinocytes (HaCaT). Ropivacaine, a widely used opiate local anaesthetic, exerts cell toxic and apoptotic effects against HaCaT cells. The aim of the present study is to investigate whether zinc modulates the effects of ropivacaine on proliferation, viability, apoptosis and NEP expression in HaCaT cells. To investigate the role of ropivacaine in NEP function, HaCaT cells overexpressing NEP were generated via cell transfection with plasmids carrying NEP cDNA. Ropivacaine's anti-proliferative effect was tested by Neubauer's chamber cell counting, and induction of cell death was demonstrated by trypan blue exclusion assay. Apoptosis due to ropivacaine was tested via DNA fragmentation and poly-ADP-ribose-polymerase (PARP) cleavage. NEP and PARP expression was performed by western blot analysis. Results showed that zinc (15 µΜ) inhibited proliferation and cell death induction by ropivacaine (0.5, 1 and 2 mM) (p < 0.05) as well as apoptosis induced by the drug (0.5 and 1 mM) in HaCaT cells. Ropivacaine (1.0, 2.0 and 5.0 mM) downregulated NEP expression in the presence of zinc (15 µΜ) while NEP overexpression enhanced ropivacaine's apoptotic effect. In conclusion, the abilities of zinc to inhibit the toxic and apoptotic effects of ropivacaine, to maintain NEP downregulation induced by the drug and, consequently, to enhance its anaesthetic result suggest that zinc may have a significant role in pain management and tissue protection.

Anticancer and cytotoxic effects of a triorganotin compound with 2-mercapto-nicotinic acid in malignant cell lines and tumor bearing Wistar rats.

Nowadays, investigation for possible therapeutic applications of various metal-based drugs attracts the scientific interest worldwide. The triorganotin compound bis[triphenyltin(IV)](3-carboxy-pyridine-2-thionato) (SnMNA), was tested for its anti-proliferative and antitumor activities. Cytotoxic activity was assessed by Trypan blue and 3-(4.5-dimethylthiazol-2-yl)-2.5-diphenyltetrazolium bromide assay (MTT). SnMNA exhibited potent cytotoxic effects against leiomyosarcoma cells (LMS) and human breast adenocarcinoma cells (MCF-7), which is 200 times stronger than that of cisplatin. Moreover, SnMNA induced significant apoptosis in LMS and MCF-7 cells characterized by flow cytometry analysis and DNA fragmentation. Acute and chronic toxicity studies on Wistar rats caused kidney and lung toxicity at a single dose of 80mg/kgBody Weight (BW) or four repeated doses of 8mg/kgBW once per week. Furthermore, antitumor activity studies on sarcoma bearing Wistar rats revealed that SnMNA complex at four repeated doses of 5.4mg/kgBW every three days prolonged mean survival time of the animal at 200% and decreased mean tumor growth rate (MTGR) compared to the control group (p<0.05). It is noteworthy to mention that the 30% (3 out of 10) of the bearing animals were totally cured. These findings indicate that SnMNA might be a promising new antitumor agent.

Vanadium-induced apoptosis of HaCaT cells is mediated by c-fos and involves nuclear accumulation of clusterin.

Vanadium exerts a variety of biological effects, including antiproliferative responses through activation of the respective signaling pathways and the generation of reactive oxygen species. As epidermal cells are exposed to environmental insults, human keratinocytes (HaCaT) were used to investigate the mechanism of the antiproliferative effects of vanadyl(IV) sulfate (VOSO(4)). Treatment of HaCaT cells with VOSO(4) inhibited proliferation and induced apoptosis in a dose-dependent manner. Inhibition of proliferation was associated with downregulation of cyclins D1 and E, E2F1, and the cyclin-dependent kinase inhibitors p21(Cip1/Waf1) and p27(Kip1). Induction of apoptosis correlated with upregulation of the c-fos oncoprotein, changes in the expression of clusterin (CLU), an altered ratio of antiapoptotic to proapoptotic Bcl-2 protein family members, and poly(ADP-ribose) polymerase-1 cleavage. Forced overexpression of c-fos induced apoptosis in HaCaT cells that correlated with secretory CLU downregulation and upregulation of nuclear CLU (nCLU), a pro-death protein. Overexpression of Bcl-2 protected HaCaT cells from vanadium-induced apoptosis, whereas secretory CLU overexpression offered no cytoprotection. In contrast, nCLU sensitized HaCaT cells to apoptosis. Our data suggest that vanadium-mediated apoptosis was promoted by c-fos, leading to alterations in CLU isoform processing and induction of the pro-death nCLU protein.

Bcl-2 blocks 2-methoxyestradiol induced leukemia cell apoptosis by a p27(Kip1)-dependent G1/S cell cycle arrest in conjunction with NF-kappaB activation.

2-Methoxyestradiol (2-ME2) induces leukemia cells to undergo apoptosis in association with Bcl-2 inactivation but the mechanisms whereby Bcl-2 contributes to protection against programmed cell death in this context remain unclear. Here we showed that 2-ME2 inhibited the proliferation of Jurkat leukemia cells by markedly suppressing the levels of cyclins D3 and E, E2F1 and p21(Cip1/Waf1) and up-regulating p16(INK4A). Further, 2-ME2 induced apoptosis of Jurkat cells in association with down-regulation and phosphorylation of Bcl-2 (as mediated by JNK), up-regulation of Bak, activation of caspases-9 and -3 and PARP-1 cleavage. To determine the importance and mechanistic role of Bcl-2 in this process, we enforced its expression in Jurkat cells by retroviral transduction. Enforcing Bcl-2 expression in Jurkat cells abolished 2-ME2-induced apoptosis and instead produced a G1/S phase cell cycle arrest in association with markedly increased levels of p27(Kip1). Bcl-2 and p27(Kip1) were localized mainly in the nucleus in these apoptotic resistant cells. Interestingly, NF-kappaB activity and p50 levels were increased by 2-ME2 and suppression of NF-kappaB signaling reduced p27(Kip1) expression and sensitized cells to 2-ME2-induced apoptosis. Importantly, knocking-down p27(Kip1) in Jurkat Bcl-2 cells sensitized them to spontaneous and 2-ME2-induced apoptosis. Thus, Bcl-2 prevented the 2-ME2-induced apoptotic response by orchestrating a p27(Kip1)-dependent G1/S phase arrest in conjunction with activating NF-kappaB. Thus, we achieved a much better understanding of the penetrance and mechanistic complexity of Bcl-2 dependent anti-apoptotic pathways in cancer cells and why Bcl-2 inactivation is so critical for the efficacy of apoptosis and anti-proliferative inducing drugs like 2-ME2.

Bcl-2 but not clusterin/apolipoprotein J protected human diploid fibroblasts and immortalized keratinocytes from ceramide-induced apoptosis: role of p53 in the ceramide response.

The role of clusterin/apolipoprotein J (Clu/ApoJ) and Bcl-2 on C(2)-ceramide-induced apoptosis of embryonic human diploid fibroblasts, MRC-5 and immortalized adult skin keratinocytes, HaCaT was investigated. C(2)-ceramide-induced apoptosis of HaCaT in a time- and dose-dependent manner, while in MRC-5 only at higher concentrations. There was a dose-dependent accumulation of Clu/ApoJ and downregulation of Bcl-2 which correlated with C(2)-ceramide-induced apoptosis of MRC-5. While overexpression of Bcl-2 suppressed C(2)-ceramide-mediated apoptosis in both cell types, Clu/ApoJ failed to do so, accessed by morphological changes, DNA fragmentation and PARP cleavage. There was no change in the expression of endogenous p53 or p21(Waf1/Cip1) upon C(2)-ceramide treatment of MRC-5. However, mutant p53(143ala) increased the sensitivity of MRC-5 to C(2)-ceramide-induced apoptosis by markedly downregulating Bcl-2, pointing to a role for p53. These results suggested that whereas downregulation of Bcl-2 may be a crucial factor involved in C(2)-ceramide-induced apoptosis, accumulation of Clu/ApoJ may be a signal of stress response. Moreover, the ceramide-activated apoptotic pathway may be regulated by p53.

Ectopic expression of clusterin/apolipoprotein J or Bcl-2 decreases the sensitivity of HaCaT cells to toxic effects of ropivacaine.

Local anesthetics inhibit cell proliferation and induce apoptosis in various cell types. Ropivacaine, a unique, novel tertiary amine-type anesthetic, was shown to inhibit the proliferation of several cell types including keratinocytes. We found that Ropivacaine could inhibit the proliferation and induce apoptosis in an immortalized human keratinocyte line, HaCaT, in a dose- and time-dependent manner and with the deprivation of serum. The dose-dependent induction of apoptosis by ropivacaine was demonstrated by DNA fragmentation analysis and the proteolytic cleavage of a caspase-3 substrate-poly (ADP-ribose) polymerase (PARP). In addition, ropivacaine downregulated the expression of clusterin/ apoliporotein J, a protein with anti-apoptotic properties, in a dose-dependent manner, which well correlated with the induction of apoptosis of HaCaT cells. To investigate the role of clusterin/apoliporotein J in ropivacaine-induced apoptosis, HaCaT cells overexpressing clusterin/apoliporotein J were generated and compared to cells expressing the well established anti-apoptotic Bcl-2 protein. Ectopic overexpression of the secreted form of clusterin/apoliporotein J or Bcl-2 decreased the sensitivity of HaCaT cells to toxic effects of ropivacaine as demonstrated by DNA fragmentation, the proteolytic cleavage of PARP and by a reduction in procaspase-3 expression. Furthermore, the downregulation of endogenous clusterin/apolipoprotein J levels by ropivacaine suggested that this might be one mechanism by which ropivacaine induced cell death in HaCaT cells. In conclusion, the ability of ropivacaine to induce antiproliferative responses and to suppress the expression of the anti-apoptotic protein clusterin/apolipoprotein J, combined with previously reported anti-inflammatory activity and analgesic property of the drug, suggests that ropivacaine may have potential utility in the local treatment of tumors.