Blood lead and cadmium levels in a six hospital employee population. PESA study, 2009

IntroductionExposure to lead and cadmium is a public health problem due to the broad exposure to these toxic substances among the general population. The objective of this study is to determine blood lead and cadmium concentrations in a working population drawn from six university hospitals in Madrid, Getafe, Cartagena, Santiago de Compostela, Santander and Palma de Mallorca (Spain) and to identify associated factors.Materials and methods951 individuals participated in the study and were administered the standardized PESA^® questionnaire regarding exposure to lead and cadmium. The blood lead and cadmium concentrations were measured by electrothermal atomization atomic absorption spectrometry with Zeeman background correction in Perkin-Elmer spectrometers, guaranteeing the transferability of the results.ResultsThe median overall blood lead concentration was: 1.6 μg/dL (IQR: 0.9–2.7) and that of cadmium was: 0.21 μg/L (IQR: 0.10–0.50). There were significant differences in lead levels between men (2 μg/dL) and women (1.5 μg/dL), postmenopausal (2.6 μg/dL) and premenopausal women (1.1 μg/dL), and between participants who cooked in earthenware (2.1 μg/dL) and those who did not (1.5 μg/dL). The median of cadmium in women (0.24 μg/L) was higher than in men (0.11 μg/L) and was also higher in subjects who smoked (0.70 μg/L) than in non-smokers (0.13 μg/L).ConclusionsA reduction in blood lead and cadmium levels was observed with respect to previous studies carried out in Spain. Nevertheless, the results suggest there are certain factors which increase risk such as age, gender, menopause, age of housing, cooking in lead-glazed earthenware and exposure to cigarette smoke.     

Casearin X exhibits cytotoxic effects in leukemia cells triggered by apoptosis

Clerodane diterpenes have demonstrated cytotoxic, antiplasmodial and anti-ulcer properties. In the present work, we determined the cytotoxic effect of casearin L (Cas L), O (Cas O) and X (Cas X) and (?)-hardwickiic acid isolated from Casearia sylvestris leaves, and investigated the underlying mechanisms involved in in vitro cell death induced by Cas X in HL-60 leukemia cells (0.7, 1.5 and 3.0 μM). Cytotoxicity tests demonstrated that Cas X was the most active compound studied, showing greater cytotoxic effects against CEM and HL-60 lines (IC₅₀ of 0.4 μM) and human peripheral blood mononuclear cells (PBMC, IC₅₀ of 1.2 μM). After 24 h exposure, Cas X caused a decrease in 5-bromo-20-deoxyuridine (BrdU) incorporation (36.6 and 24.5% labeling at 0.7 and 1.5 μM, respectively), reduction in viability, and increase in apoptotic and necrotic leukemia cells in a dose-dependent manner evidenced by the trypan blue and AO/EB (acridine orange/ethidium bromide) assays. Moreover, Cas X-treated cells exhibited nuclear fragmentation and cytoplasmic vacuolization depending on the concentration tested. These characteristics of apoptosis or secondary necrosis were confirmed by flow cytometry which revealed DNA fragmentation, phosphatidylserine externalization, activation of the effector caspases 3/7 and mitochondrial depolarization. We then found evidence that Cas X causes cell death via apoptotic pathways, corroborating the potential of casearins as compounds with promising antitumor-related properties.     

The α-mangostin prevention on cisplatin-induced apoptotic death in LLC-PK1 cells is associated to an inhibition of ROS production and p53 induction

Cisplatin (CDDP) is a widely useful chemotherapeutic agent for the treatment of tumors including lung, ovary and testis. Acute renal injury, however, is the main side effect observed after CDDP treatment. This side effect is related to the apoptotic death in proximal tubular cells in the kidney and p53 protein has a central role in this process. On the other hand, α-mangostin (α-M), a xanthone derived from the pericarp of mangosteen, exerts a renoprotective effect against cisplatin-induced renal damage in rats. The aim of this study was to evaluate whether α-M protects proximal tubule renal epithelial cells (LLC-PK1) from CDDP-induced apoptotic death. Cells were co-incubated with 5 μM α-M and 100 μM CDDP for 24 h. It was found that α-M attenuated the following alterations: the apoptotic cell death, the increase in reactive oxygen species (ROS), the glutathione depletion and the increase in p53 expression induced by CDDP. In conclusion, the preventive effect of α-M on CDDP-induced apoptotic death is associated to the inhibition of p53 expression and ROS generation.     

Trace elements in bipolar disorder

IntroductionTrace elements may play an important role in bipolar disorders. The objective of this study is to determine serum copper and zinc, blood lead and cadmium and urine lead, cadmium and thallium concentrations in patients diagnosed with bipolar disorders and to compare these levels with those of a healthy control group.Materials and methodsA total of 25 patients diagnosed with bipolar disorder and 29 healthy subjects participated in this study. Serum copper and zinc concentrations were measured using flame atomic absorption spectrometry; the blood lead and cadmium concentrations were measured by electrothermal atomization atomic absorption spectrometry with Zeeman background correction; urine lead, cadmium and thallium concentrations were measured by inductively coupled plasma mass spectrometry.ResultsMedian blood and urine lead and cadmium levels were significantly higher among the bipolar patients than among the control group: Blood lead (μg/dL): patient median: 3.00 (IQR: 1.40–4.20); control median (μg/dL): 2.20 (IQR: 0.90–3.00) p = 0.040. Blood cadmium (μg/L): patient median: 0.39 (IQR: 0.10–1.15); control median: 0.10 (IQR: 0.10–0.17) p < 0.001. The median of cadmium (μg/L) in patients who smoked (1.20 IQR: 0.44–2.30) was higher than that in non-smokers (0.12 IQR: 0.10–0.34) p < 0.001. There was a statistically significant increase (p = 0.001) in zinc levels among patients in the manic phase (mean 111.28, SD: 33.36 μg/dL) with respect to the control group (mean 86.07, SD: 12.39 μg/dL).ConclusionsThe results suggest that there could be higher levels of some toxic trace elements in the group of patients with bipolar disorder than in the healthy control group.     

A pharmaceutical preparation of Salvia miltiorrhiza protects cardiac myocytes from tumor necrosis factor-induced apoptosis and reduces angiotensin II-stimulated collagen synthesis in fibroblasts

Salvia miltiorrhiza is a medicinal herb commonly used in traditional Chinese medicine for the prevention and treatment of cardiovascular disease. This study investigated the effects of Cardiotonic Pill (CP), a pharmaceutical preparation of Salvia miltiorrhiza, on cardiac myocytes and fibroblasts with respect to the viability, proliferation, and collagen synthesis in these cells under various conditions. A cardiac myocyte line, H9c2, and primarily cultured fibroblasts from rat hearts were incubated with CP over a broad concentration range (50–800 μg/ml) under normal cultures, conditions of ischemia (serum-free culture), and stimulation by angiotensin II (AII, 100 nM), hydrogen peroxide (H₂O₂, 50–200 μM), or tumor necrosis factor α (TNFα, 40 ng/ml) for 24–48 h. Cell growth, apoptosis, DNA and collagen synthesis, and expression of relevant genes were assessed via cell number study, morphological examination, Annexin-V staining, flow-cytometry, [³H]-thymidine or [³H]-proline incorporation assay, and Western blotting analysis. It was found that (1) at therapeutic (50 μg/ml) and double therapeutic (100 μg/ml) concentrations, CP did not significantly affect normal DNA synthesis and cell growth in these cardiac cells, while at higher (over 4-fold therapeutic) concentrations (200–800 μg/ml), CP decreased DNA synthesis and cell growth and increased cell death; (2) CP treatment (50 μg/ml) significantly inhibited TNFα-induced apoptosis in myocytes, with 12.3±1.46% cells being apoptosis in CP treatment group and 37.0±7.34% in the control (p<0.01), and simultaneously, expression of activated (phosphorylated) Akt protein was increased by about 2 folds in the CP-treated cells; and (3) in cultured fibroblasts, CP significantly reduced AII-induced collagen synthesis in a concentration-dependent manner (by ～50% and ～90% reduction of AII-induced collagen synthesis at 50 and 100 μg/ml, respectively). Thus, Salvia miltiorrhiza preparation CP is physiologically active on cardiac cells. The actions by CP to reduce apoptotic damage in myocytes and collagen synthesis in fibroblasts may help to preserve the heart function and reduce heart failure risk. The actions by CP to inhibit DNA synthesis and cell growth, which occurred at over therapeutic doses, may weaken the ability of heart repair. Further studies are needed to identify the chemical compounds in this herbal product that are responsible for these observed physiological effects.     

The effects of angiotensin II and the oxidative stress mediator 4-hydroxynonenal on human osteoblast-like cell growth: possible relevance to otosclerosis

Otosclerosis is a complex disease characterized by an abnormal bone turnover of the otic capsule resulting in conductive hearing loss. Recent findings have shown that angiotensin II (Ang II), a major effector peptide of the renin–angiotensin system, plays an important role in the pathophysiology of otosclerosis, most likely by its proinflammatory effects on the bone cells. Because reactive oxygen species play a role both in inflammation and in the cellular signaling pathway of Ang II, the appearance of protein adducts of the “second messenger of free radicals,” the aldehyde 4-hydroxynonenal (HNE), in otosclerotic bone has been analyzed. Immunohistochemical analysis of HNE-modified proteins in tissue samples of the stapedial bones performed on 15 otosclerotic patients and 6 controls revealed regular HNE–protein adducts present in the subperiosteal parts of control bone specimens, whereas irregular areas of a pronounced HNE–protein adduct presence were found within stapedial bone in cases of otosclerosis. To study possible interference by HNE and Ang II in human bone cell proliferation, differentiation, and induction of apoptosis we used an in vitro model of osteoblast-like cells. HNE interacted with Ang II in a dose-dependent manner, both by forming HNE–Ang II adducts, as revealed by immunoblotting, and by modifying its effects on cultured cells. Namely, treatment with 0.1 nM Ang II and 2.5 μM HNE stimulated proliferation, whereas treatment with 10 μM HNE or in combination with Ang II (0.1, 0.5, and 1 nM) decreased cell proliferation. Moreover, 10 μM HNE alone and with Ang II (except if 1 nM Ang II was used) increased cellular differentiation and apoptosis. HNE at 5 μM did not affect differentiation nor significantly change apoptosis. On the other hand, when cells were treated with lower concentrations of HNE and Ang II we observed a decrease in cellular differentiation (combination of 1.0 or 2.5 μM HNE with 0.1 nM Ang II) and decrease in apoptosis (0.1 and 0.5 nM Ang II). Cellular necrosis was increased with 5 and 10 μM HNE if given alone or combined with Ang II, whereas 0.5 nM Ang II and combination of 1 μM HNE with Ang II (0.1 and 0.5 nM) reduced necrosis. These results indicate that HNE and Ang II might act mutually dependently in the regulation of bone cell growth and in the pathophysiology of otosclerosis.     

6 Iodo-δ-lactone: A derivative of arachidonic acid with antitumor effects in HT-29 colon cancer cells

BackgroundIL-δ (5-hydroxy-6 iodo-8,11,14-eicosatrienoic delta lactone) an iodinated arachidonic acid (AA) derivative, is one of the iodolipids biosynthesized by the thyroid. Although IL-δ regulates several thyroid parameters such as cell proliferation and goiter growth it was found that this iodolipid inhibits the growth of other non thyroid cell lines.ObjectivesTo study the effect of IL-δ on cell proliferation and apoptosis in the colon cancer cell line HT-29.ResultsTreatment with IL-δ reduced cell viability in a concentration-dependent manner: 1 μM 20%, 5 μM 25%, 10 μM 31%, 50 μM 47% and caused a significant decrease of PCNA expression (25%). IL-δ had pro-apoptotic effects, evidenced by morphological features of programmed cell death such as pyknosis, karyorrhexis, cell shrinkage and cell blebbing observed by fluorescence microscopy, and an increase in caspase-3 activity and in Bax/Bcl-2 ratio (2.5 after 3 h of treatment). Furthermore, IL-δ increased ROS production (30%) and lipid peroxidation levels (19%), suggesting that apoptosis could be a result of increased oxidative stress. A maximum increase in c-fos and c-jun protein expression in response to IL-δ was observed 1 h after initiation of the treatment. IL-δ also induced a tumour growth delay of 70% compared to the control group in NIH nude mice implanted with HT-29 cells.ConclusionOur study shows that IL-δ inhibits cell growth and induces apoptosis in the colon cancer cell line, HT-29 and opens the possibility that IL-δ could be a potential useful chemotherapy agent.     

Synthesis of a terphenyl substituted 4-aza-2,3-didehydropodophyllotoxin analogues as inhibitors of tubulin polymerization and apoptosis inducers

A series of terphenyl based 4-aza-2,3-didehydropodophyllotoxin conjugates (8a–r) were synthesized by a straightforward one-step multicomponent synthesis that demonstrated anticancer activity against five human cancer cell lines (lung, colon, renal, prostate and cervical). All the tested compounds showed potent anticancer activity with IC₅₀ values ranging from 0.87 to 16.59 μM. Among them compounds 8n and 8p showed significant anticancer activity in lung cancer cells with IC₅₀ values 0.91 and 0.87 μM, respectively. Flow cytometric analysis revealed that these compounds induced cell cycle arrest in G₂/M phase in A549 cell line leading to caspase-3 dependent apoptotic cell death. The tubulin polymerization assay and immunofluorescence analysis showed that these compounds effectively inhibit microtubule assembly at both molecular and cellular levels in A549 cells. Further, Hoechst staining, DNA fragmentation analysis also suggested that these compounds induced cell death by apoptosis. Overall, the current study demonstrated that the synthesis of terphenyl based 4-aza-2,3-didehydropodophyllotoxin conjugates as promising anticancer agents with G₂/M cell cycle arrest and apoptotic-inducing activities via targeting tubulin.     

Synthesis and synergetic anti-tumor activity evaluation of dihydroartemisinin-organogermanium(IV) compound

Dihydroartemisinin (DHA), a semi-synthetic derivative of the herb artemisinin, has shown commendable bioactivity. In this paper, a novel dihydroartemisinin-organogermanium (DHA-Ge) compound was synthesized, characterized and its potential anti-tumor activity was evaluated by various methods. MTT results demonstrated that DHA-Ge could effectively inhibit the proliferation of HepG2 cells and showed their dose-dependent properties. The IC₅₀ value of inhibition effect on HepG2 cells of DHA-Ge was 10.23 μg/ml which was lower than 39.44 μg/ml of DHA. Flow cytometric results suggested that DHA-Ge could induce apoptosis of HepG2 cells and the apoptosis rate was 20.26% after 24 h treatment with 56.8 μg/ml DHA-Ge concentration. Atomic force microscopy images showed that HepG2 cells were collapsed and the cell nucleus were fragmented after 24 h treatment. All these results together showed that the DHA-Ge possessed desirable synergetic enhanced anti-tumor effects and could be developed as a suitable tumor therapeutic agent.     

Nitric oxide synthase activation and oxidative stress,but not intracellular zinc dyshomeostasis,regulate ultraviolet B light-induced apoptosis

AimsTo investigate the role of nitric oxide synthase (NOS) and intracellular free zinc ion (Zn²⁺) in regulation of ultraviolet B light (UVB)-induced cell damage and apoptosis.Main methodsReal-time confocal microscopy measurement was used to determine the changes of intracellular free zinc concentration under different conditions. Cell apoptotic death was determined using fluorescein isothiocyanate (FITC) conjugated-annexin V (ANX5)/PI labeling followed by flow cytometry. Western analysis was used to determine cell apoptosis and eNOS uncoupling.Key findingsUVB induced an elevation of Zn²⁺ within 2 min of exposure. The UVB-induced intracellular Zn²⁺ elevation was dependent on the increase of constitutive nitric oxide synthase (cNOS) activity and production of superoxide. Removal of Zn²⁺ with a lower concentration (< 25 μM) of N,N,N′,N′-tetrakis (2-pyridylmethyl) ethylenediamine (TPEN), a Zn²⁺-specific chelator, did not induce cell death or prevent cells from UVB-induced apoptosis. However, a higher [TPEN] (> 50 μM) was cytotoxic to cells, but prevented cells from further UVB-induced apoptosis. The higher [TPEN] also induced cNOS uncoupling. Furthermore, treating the cells with a membrane permeable superoxide dismutase (PEG-SOD) inhibited Zn²⁺ release and reduced apoptotic cell death after UVB treatment. The results demonstrated a complex and dynamic regulation of UVB-induced cell damage.SignificanceOur findings not only advance our understanding of the correlations between cNOS activation and Zn elevation, but also elucidated the role of cNOS in regulation of oxidative stress and apoptosis upon UVB-irradiation.     

Time-course correlation of early toxic events in three models of striatal damage: Modulation by proteases inhibition

Metabolic alterations in the nervous system can be produced at early stages of toxicity and are linked with oxidative stress, energy depletion and death signaling. Proteases activation is responsible for triggering deadly cascades during cell damage in toxic models. In this study we evaluated the early time-course of toxic events (oxidative damage to lipids, mitochondrial dysfunction and LDH leakage, all at 1, 3 and 6 h) in rat striatal slices exposed to quinolinic acid (QUIN, 100 μM) as an excitotoxic/pro-oxidant model, 3-nitropropionic acid (3-NP, 1 mM) as an inhibitor of mitochondrial succinate dehydrogenase, and a combined model produced by the co-administration of these two toxins at subtoxic concentrations (21 and 166 μM for QUIN and 3-NP, respectively). In order to further characterize a possible causality of caspases or calpains on the toxic mechanisms produced in these models, the broad calpain inhibitor IC1 (50 μM), and the pan-caspase inhibitor Z-VAD (100 μM) were tested. Lipid peroxidation (LP) was increased at all times and in all models evaluated. Both IC1 and Z-VAD exerted significant protection against LP in all models and at all times evaluated. Mitochondrial dysfunction (MD) was consistently affected by all toxic models at 3 and 6 h, but was mostly affected by 3-NP and QUIN at 1 h. IC1 differentially protected the slices against 3-NP and QUIN at 1 h and against QUIN at 3 h, while Z-VAD exhibited positive actions against QUIN and 3-NP at all times tested, and against their combination at 3 and 6 h. LDH leakage was enhanced at 1 and 3 h in all toxic models, but this effect was evident only for 3-NP + QUIN and 3-NP at 6 h. IC1 protected against LDH leakage at 1 h in 3-NP + QUIN and 3-NP models, at 3 h in all toxic models, and at 6 h in 3-NP + QUIN and 3-NP models. In turn, Z-VAD protected at 1 and 6 h in all models tested, and at 3 h in the combined and QUIN models. Our results suggest differential chronologic and mechanistic patterns, depending on the toxic insult. Although LP, MD and membrane cell rupture are shared by the three models, the occurrence of each event seems to obey to a selective recruitment of damaging signals, including a differential activation of proteases in time. Proteases activation is likely to be an up-stream event influencing oxidative stress and mitochondrial dysfunction in these toxic models.     

Levels of l-ascorbic acid and cadmium in the saphenous vein of patients with coronary artery disease are negatively correlated

BackgroundThe aim of this study was the simultaneous determination of levels of cadmium and l-ascorbic Acid (AA) in human saphenous vein (SV) used in coronary artery bypass grafting (CABG) and check whether there is a relationship between these levels.MethodsHuman SV were collected from 40 patients (20 men and 20 women; age, 40–75 years) at the time of routine coronary artery surgical revascularization. The concentration of cadmium in the tissue was determined according to the GF AAS—atomic absorption method. The concentration of AA was assayed in supernatant by FIA method with spectrophotometric detection.ResultsAA concentration (mean ± SD); men: 98,7 ± 13,18 μg/g tissue, women: 96,06 ± 11,98 μg/g tissue. Cadmium concentration(mean ± SD); men: 309 ± 103,71 ng/g tissue, women: 348,5 ± 255,71 ng/g tissue. Correlations among concentrations of AA and cadmium were insignificant negative in the group of men (Pearson r = −0,1504, p = 0,5269) and in the group women (Pearson r = −0339, p = 0144).ConclusionsNegative correlations among concentrations of AA and cadmium in human SV obtained in our study may indicate a protective effect of this vitamin in relation to toxic cadmium.     

Synthesis and biological evaluation of 1,2,3-triazole linked aminocombretastatin conjugates as mitochondrial mediated apoptosis inducers

A series of 1,2,3-triazole linked aminocombretastatin conjugates were synthesized and evaluated for cytotoxicity, inhibition of tubulin polymerization and apoptosis inducing ability. Most of the conjugates exhibited significant anticancer activity against some representative human cancer cell lines and two of the conjugates 6d and 7c displayed potent cytotoxicity with IC₅₀ values of 53 nM and 44 nM against A549 human lung cancer respectively, and were comparable to combretastatin A-4 (CA-4). SAR studies revealed that 1-benzyl substituted triazole moiety with an amide linkage at 3-position of B-ring of the combretastatin subunit are more active compared to 2-position. G2/M cell cycle arrest was induced by these conjugates 6d and 7c and the tubulin polymerization assay (IC₅₀ of 1.16 μM and 0.95 μM for 6d and 7c, respectively) as well as immunofluorescence analysis showed that these conjugates effectively inhibit microtubule assembly at both molecular and cellular levels in A549 cells. Colchicine competitive binding assay suggested that these conjugates bind at the colchicine binding site of tubulin as also observed from the docking studies. Further, mitochondrial membrane potential, ROS generation, caspase-3 activation assay, Hoechst staining and DNA fragmentation analysis revealed that these conjugates induce cell death by apoptosis.     

Enhanced cytotoxic activity of doxorubicin through the inhibition of autophagy in triple negative breast cancer cell line

BackgroundThe outcome of triple negative breast cancer is still poor and requires improvement with better therapy options. Autophagy has recently been shown to play a role in anticancer drug resistance. Therefore, we investigated if the effectiveness of doxorubicin was augmented by the inhibition of autophagy.MethodsMDA-MB-231 was used as a model cell line for triple negative breast cancer and 3-methyladenine was used as an inhibitor of autophagy. Cells were treated with 0.46–1.84 μM doxorubicin and 2.5–10 μM 3-methyladenine for 48 h. Cell death mode was examined with M30 and M65 ELISA assays. ROS level and LDH activity was examined and the cellular acidic compartment of cells was monitored by acridine orange staining. The expression of various autophagy and apoptosis related proteins/genes were evaluated with Western blotting and RT-qPCR respectively.ResultsSynergism was observed between the compounds (CI value < 1.0). RT-qPCR analysis revealed that the combination resulted in a down-regulation of autophagy-related genes. Moreover, the combination resulted in a different cell death modality, upregulating necroptosis-related genes. This suggests that the mode of cell death may switch from apoptosis to necroptosis, which is a more severe form of cell death, when autophagy is inhibited. These results were further confirmed at protein level by Western blotting.ConclusionInhibition of autophagy seems to sensitize triple negative breast cancer cells to doxorubicin, warranting further in vivo studies for the proof of this concept.General significanceAutophagy has a key role in drug resistance in MDA-MB-231 cells. Therefore combinatorial approaches may effectively overcome resistance.     

Ricin A-chain requires c-Jun N-terminal kinase to induce apoptosis in nontransformed epithelial cells

Ricin is a toxin isolated from castor beans that has potential as a weapon of bioterrorism. This glycoprotein consists of an A-chain (RTA) that damages the ribosome and inhibits protein synthesis and a B-chain that plays a role in cellular uptake. Ricin activates the c-Jun N-terminal kinase (JNK) and p38 signaling pathways; however, a role for these pathways in ricin-induced cell death has not been investigated. Our goals were to determine if RTA alone could activate apoptosis and if the JNK and p38 pathways were required for this response. Comparable caspase activation was observed with both ricin and RTA treatment in the immortalized, nontransformed epithelial cell line, MAC-T. Ribosome depurination and inhibition of protein synthesis were induced in 2–4 h with 1 μg/ml RTA and within 4–6 h with 0.1 μg/ml RTA. Apoptosis was not observed until 4 h of treatment with either RTA concentration. RTA activated JNK and p38 in a time- and concentration-dependent manner that preceded increases in apoptosis. Inhibition of the JNK pathway reduced RTA-induced caspase activation and poly(ADP-ribose) polymerase cleavage. In contrast, inhibition of the p38 pathway had little effect on RTA-induced caspase 3/7 activation. These studies are the first to demonstrate a role for the JNK signaling pathway in ricin-induced cell death. In addition, the MAC-T cell line is shown to be a sensitive in vitro model system for future studies using RTA mutants to determine relationships between RTA-induced depurination, ribotoxic stress, and apoptosis in normal epithelial cells.     

Protection of oxidative stress induced apoptosis in osteosarcoma cells by dihydromyricetin through down-regulation of caspase activation and up-regulation of BcL-2

Current study was aimed to investigate the effect of dihydromyricetin on hydrogen peroxide induced oxidative stress in the osteosarcoma cells. MTT assay showed that hydrogen peroxide treatment at a concentration of 100 μM caused a significant (p < 0.005) reduction in the viability of MG63 cells. However, reduction in cell viability caused by 100 μM concentration of hydrogen peroxide was completely prevented on incubation with 30 μM dose of dihydromyricetin. Treatment with 100 μM concentration of hydrogen peroxide for 24 h led to condensation of chromatin material, rounding of cell shape and detachment of cells. The results from flow cytometry using annexin V-FITC and PI double staining showed apoptosis induction in 47.84 ± 5.21% cells on treatment with 100 μM concentration of hydrogen peroxide compared to 2.32 ± 0.54% in controlcells. The apoptotic alterations in MG63 cell morphology were prevented significantly on pre-treatment with 30 μM doses of dihydromyricetin for 48 h. Annexin V-FITC and PI staining showed reduction of hydrogen peroxide induced apoptotic cell percentage to 3.07 ± 0.86% on pre-treatment of MG63 cells with 30 μM dose of dihydromyricetin. Western blot analysis showed a significant increase in the activation of caspase-3 and -9 on treatment of MG63 cells for 24 h with 100 μM concentration of hydrogen peroxide. The expression level of Bcl-2 was decreased significantly by 100 μM concentration of hydrogen peroxide in MG63 cells. However, pre-treatment of MG63 cells with 30 μM dose of dihydromyricetin for 48 h significantly prevented hydrogen peroxide induced increase in caspase-3 and -9 levels and reduction in Bcl-2 level. Thus dihydromyricetin prevents hydrogen peroxide induced reduction in viability and induction of apoptosis in MG63 cells through down-regulation of caspase activation and up-regulation of Bcl-2 levels.     

Gender difference in blood cadmium concentration in the general population: Can it be explained by iron deficiency?

IntroductionGender differences in blood cadmium concentrations and the effect of iron deficiency on blood cadmium levels were analyzed in a representative sample of Koreans assessed in the Korean National Health and Nutritional Examination Survey (KNHANES) 2008–2011.MethodsA rolling sampling design was used to perform a complex, stratified, multistage probability cluster survey of a representative sample of the non-institutionalized civilian population in South Korea. Serum ferritin was categorized as low (<15.0 μg/L), low normal (15.0–<30.0 μg/L for females and 15.0–<50.0 μg/L for males), and normal (≥30.0 μg/L for females and ≥50.0 μg/L for males), and its association with blood cadmium levels was assessed after adjustment for various demographic and lifestyle factors.ResultsThe geometric mean (GM) of the blood cadmium level was significantly higher in females than in males, and significantly higher in older individuals for both genders. After controlling for covariates, multiple regression analysis with interaction terms showed that blood cadmium was correlated with serum ferritin levels only in pre-menopausal females.DiscussionIron deficiency is associated with blood cadmium levels in a representative sample of pre-menopausal females, as evaluated in KNHANES. Gender differences in blood cadmium concentration may not be due solely to an iron deficiency-associated increase in blood cadmium.     

3′,4′-Bis-difluoromethoxycinnamoylanthranilate (FT061): An orally-active antifibrotic agent that reduces albuminuria in a rat model of progressive diabetic nephropathy

Cinnamoylanthranilates including tranilast have been identified as promising antifibrotics that can reduce fibrosis occurring in the kidney during diabetes, thereby delaying and/or preventing kidney dysfunction. Structure–activity relationships aimed at improving potency and metabolic stability have led to the discovery of FT061. This compound, which bears a bis-difluoromethoxy catechol, attenuates TGF-β-stimulated production of collagen in cultured renal mesangial cells (approx 50% at 3 μM). When dosed orally at 20 mg/kg to male Sprague Dawley rats, FT061 exhibited a high bioavailability (73%), C_max of 200 μM and T_max of 150 min, and a half-life of 5.4 h. FT061 reduced albuminuria when orally dosed in rats at 200 mg kg/day in a late intervention study of a rat model of progressive diabetic nephropathy.     

Cyathula prostrata ethanol extract activates the extrinsic pathway of apoptosis in HeLa and U937 cell lines

Cyathula prostrata is a medicinal plant that is used in tropical Africa, Asia and Australia to treat many ailments including rheumatic fever, dysentery, wounds and eye trouble (Burkill, 1995). Sowemimo et al. (2009) reported cytotoxicity against a cervical cancer cell line, HeLa, at a concentration of 250 μg/mL. Due to the reported cytotoxicity, the mode of cell death caused by an ethanolic C. prostrata extract was investigated. Dose–response assays were carried out using HeLa and U937 cell lines and IC₅₀ values of 100.8 μg/mL and 64.43 μg/mL, respectively, were achieved. Cisplatin was used as a positive control for all experiments. Cytotoxicity of the plant extract was not evident when treating normal peripheral blood mononuclear cells. Progression of cells through the cell cycle, apoptosis and mitochondrial membrane potential was investigated. Arrest of cells in the G0/G1 phase of the cell cycle was evident after 24 h of exposure to the plant extract in both cell lines, but not due to increases in p21 levels. Caspase 8 activation was evident and no depolarisation of the mitochondrial membrane and cytochrome c release was seen in both cell lines. Phosphatidylserine translocation was investigated and confirmed the onset of apoptosis. Thus, it is deduced that exclusive activation of the extrinsic pathway of apoptosis is caused by the treatment of HeLa and U937 cells with ethanolic C. prostrata.     

Apoptotic effects and glucose-6-phosphate dehydrogenase responses in liver and gill tissues of rainbow trout treated with chlorpyrifos

We investigated apoptotic effects and changes in glucose-6-phosphate dehydrogenase (G6PD) enzyme activity in liver and gill tissues of fish exposed to chlorpyrifos. Three different chlorpyrifos doses (2.25, 4.5 and 6.75 μg/L) were administrated to rainbow trout at different time intervals (24, 48, 72 and 96 h). Acute exposure to chlorpyrifos showed time dependent decrease in G6PD enzyme activity at all concentrations (p < 0.05). Immunohistochemical results showed that chlorpyrifos caused mucous cell loss in gill tissue and apoptosis via caspase-3 activation in fish. The present study suggested that chlorpyrifos inhibits G6PD enzyme and causes mucous cell loss in gill and apoptosis in gill and liver tissues.