Takehisa Matsukawa

Blood lead at currently acceptable levels may cause preterm labour

Objectives Although occupational and environmental exposures to lead have been dramatically reduced in recent decades, adverse pregnancy outcomes have been observed at ‘acceptable’ levels of blood lead concentrations (≤10 μg/dl). Methodology Blood samples were collected from 348 singleton pregnant women, aged 16–35 years, during the first trimester of pregnancy (8–12 weeks) for lead measurement by inductively coupled plasma–mass spectrometry. Subjects were followed up and divided into two groups (preterm and full-term deliveries) according to duration of gestation. Results The average (range) and geometric means of blood lead levels were 3.8 (1.0–20.5) and 3.5 μg/dl, respectively. Blood lead level was significantly (p<0.05) higher in mothers who delivered preterm babies than in those who delivered full-term babies (mean±SD: 4.46±1.86 and 3.43±1.22 μg/dl, respectively). Logistic regression analysis demonstrated that a 1 unit increase in blood lead levels led to an increased risk of preterm birth (OR 1.41, 95% CI 1.08 to 1.84). Conclusion Adverse pregnancy outcomes may occur at blood lead concentrations below the current acceptable level.

Osteopontin is involved in the development of acquired chemo-resistance of cisplatin in small cell lung cancer

Osteopontin (OPN) is a multi-functional cytokine involved in cell survival, migration and adhesion which is associated with tumorigenesis, progression and metastasis. However, the role of OPN in chemo-sensitivity of human lung cancer has not yet been elucidated. The purpose of this study is to investigate the role of OPN in chemo-sensitivity of lung cancer cells. We developed a stable OPN transfectant (SBC-3/OPN) and a control transfectant (SBC-3/NEO) from human small cell lung cancer cell line, SBC-3. SBC-3/OPN cells were more resistant to cisplatin than SBC-3/NEO cells. Multi-drug resistance-associated protein (MRP) does not appear to be involved in the development of acquired chemo-resistance, since MRP inhibitor did not alter chemo-sensitivity. After exposure to cisplatin, the apoptotic SBC-3/OPN cells were reduced in number compared to SBC-3/NEO cells. Treatment with cisplatin revealed that the expression of anti-apoptotic protein, bcl-2, was down-regulated in SBC-3/NEO cells, while that of SBC-3/OPN cells was not altered. In contrast, pro-apoptotic protein, bax, was not altered in both SBC-3/OPN and SBC-3/NEO cells, thus bcl-2/bax ratio was decreased in SBC-3/NEO but not altered in SBC-3/OPN cells. Activation of caspase-3 and caspase-9 was increased in SBC-3/NEO cells, but not in SBC-3/OPN cells. Our results suggest that OPN enhances chemo-resistance of cisplatin in SBC-3 cells by suppressing bcl-2 protein down-regulation, thereby blocking the caspase-9- and caspase-3-dependent cell apoptosis.

Chronic magnesium deficiency decreases tolerance to hypoxia/reoxygenation injury in mouse heart

AIMS Magnesium (Mg) deficiency has been reported to be associated with the development of the metabolic syndrome, cardiovascular diseases, and sudden death. We examined the influence of chronic Mg deficiency on cardiac tolerance to hypoxia/reoxygenation injury. MAIN METHODS Mice were fed an Mg-deficient diet for 4 weeks, and then their hearts were excised for Langendorff perfusion experiments. The levels of total Mg in the blood and heart were quantified by atomic absorption spectrometry. KEY FINDINGS In Mg-deficient mice, the Mg concentration in whole blood was markedly decreased; however, that in the heart remained unchanged. When the hearts of control mice were exposed to hypoxia/reoxygenation, removal of extracellular Mg from a normal Krebs solution containing 1.2 mM Mg resulted in a significant decrease in the recovery of the tension-rate product (TRP) upon reoxygenation. In Mg-deficient mice, the recovery of TRP in the heart was reduced significantly in the absence of extracellular Mg compared to that in controls. The addition of Mg to the perfusate did not improve TRP recovery. During hypoxia/reoxygenation, cardiac damage evaluated by myocardial aspartate amino transferase (AST) release was greater in hearts of Mg-deficient mice than in that of control mice. SIGNIFICANCE These results indicate that chronic Mg deficiency causes severe hypomagnesemia and a decrease in cardiac tolerance to hypoxia, without changing the intracellular Mg content. The decreased tolerance to hypoxia was not affected by the presence or absence of extracellular Mg, suggesting that some intracellular metabolic abnormalities develop in the cardiac myocytes of Mg-deficient mice.

Simultaneous determination of the enantiomers of leucine and [2H7]leucine in plasma by capillary gas chromatography–mass spectrometry

A method for the stereoselective assay of D- and L-enantiomers of both leucine and [2H7]leucine in rat plasma was developed using gas chromatography-mass spectrometry-selected-ion monitoring. DL-[2H3]leucine was used as an internal standard. The method involved purification by cation-exchange chromatography using BondElut SCX cartridge and derivatization with hydrochloric acid in methanol to form methyl ester followed by subsequent chiral derivatization with (+)-alpha-methoxy-alpha-trifluoromethylphenylacetyl chloride to form diastereomeric amide. The derivatization made the separation of the leucine enantiomers possible with good gas chromatographic behavior. Quantitation was performed by selected-ion monitoring of the quasi-molecular ions of the diastereomers on the chemical ionization method. The sensitivity, specificity, accuracy and reproducibility of the method were demonstrated to be satisfactory for application to pharmacokinetic studies of leucine enantiomers.

Sevoflurane suppresses tumour necrosis factor-α-induced inflammatory responses in small airway epithelial cells after anoxia/reoxygenation

BACKGROUND Lung ischaemia-reperfusion (I/R) injury is correlated with poor clinical outcome. The inflammatory cytokines interleukin (IL)-6, IL-8, and monocyte chemotactic protein-1 (MCP-1) are produced by pulmonary epithelial cells during lung transplantation and are considered to be involved in I/R injury. The volatile anaesthetic sevoflurane has been shown to exert a protective effect on I/R injury in various organs. We investigated the effect of sevoflurane on the inflammatory functions of pulmonary epithelial cells in vitro. METHODS Human normal small airway epithelial cells (SAEC) were incubated under anoxic conditions for 24 h with or without sevoflurane and then stimulated with tumour necrosis factor (TNF)-α under hyperoxic conditions for 5 h with or without sevoflurane. After incubation, IL-6, IL-8, and MCP-1 mRNA expression was analysed by quantitative real-time RT-PCR. The production of IL-6, IL-8, and MCP-1 was assayed by enzyme-linked immunosorbent assay, the effects of sevoflurane on inflammatory gene expression were examined by DNA microarray analysis, and the effects of sevoflurane on NF-κB-mediated inflammatory cytokine production were examined by immunoblotting. RESULTS Sevoflurane suppressed TNF-α-induced IL-6, IL-8, and MCP-1 gene expression and the production of IL-6 and IL-8 in SAEC under anoxia/reoxygenation conditions. DNA microarray analysis indicated that sevoflurane modulated NF-κB-related gene expression. Sevoflurane significantly inhibited TNF-α-induced translocation of p65 NF-κB into the nucleus. Sevoflurane enhanced TNF-α-induced gene expression of inhibitor κB (IκB) but not of NF-κB. CONCLUSIONS Sevoflurane suppressed the NF-κB-mediated production of pulmonary epithelial cell-derived inflammatory cytokines, including IL-6 and IL-8, which are capable of causing I/R injury.

Low level prenatal blood lead adversely affects early childhood mental development.

The effect of prenatal lead exposure on child development has been a topic of public health concern for decades. To estimate prenatal lead exposure effects on early childhood development, maternal blood (n = 364) and umbilical cord blood (n = 224) samples were collected during pregnancy and at delivery. Mental development was assessed using the Harold Ireton Early Child Development Inventory from 174 children. Maternal whole blood lead levels in the first trimester were significantly higher in children with developmental scores <20% than in those with normal scores (mean ± standard deviation: 6.3 ± 1.9 vs 4.0 ± 2.4 µg/dL, respectively, P = .01). Maternal blood lead levels in the first trimester were also inversely associated with the development scores (r = –0.155, P = .041). Logistic regression analysis showed a significant relationship between increasing maternal blood lead levels in the first trimester with low development scores (odds ratio = 1.74, 95% confidence interval = 1.18-2.57, P = .005). The findings of the present study showed a relatively low level of prenatal lead exposure (mean < 6.5 µg/dL) associated with lower developmental scores in early childhood.

Simultaneous determination of selenomethionine enantiomers in biological fluids by stable isotope dilution gas chromatography–mass spectrometry

A method for the stereoselective determination of D- and L-enantiomers of selenomethionine in mouse plasma was developed using gas chromatography-mass spectrometry with selected-ion monitoring (GC-MS-SIM). DL-[(2)H(3,)(82)Se]selenomethionine was used as analytical internal standard to account for losses associated with the extraction, derivatization and chromatography. Selenomethionine enantiomers in mouse plasma were purified by cation-exchange chromatography using BondElut SCX cartridge and derivatized with HCl in methanol to form methyl ester followed by subsequent N-acylation with optically active (+)-α-methoxy-α-trifluoromethylphenylacetyl chloride to form diastereomeric amide. Quantification was performed by SIM of the molecular-related ions of the diastereomers on the chemical ionization mode. The intra- and inter-day precision for D- and L-selenomethionine spiked to mouse plasma gave good reproducibility with relative standard deviation of 3% and 3% for D-selenomethionine and 6% and 3% for L-selenomethionine, respectively. The estimated amounts were in good agreement with the actual amounts spiked, the intra- and inter-day relative error being 5% and 2% for D-selenomethionine and 2% and 1% for L-selenomethionine, respectively. The present method is sensitive enough to determine pharmacokinetics of selenomethionine enantiomers.

Gas chromatographic-mass spectrometric determination of α-ketoisocaproic acid and [2H7]α-ketoisocaproic acid in plasma after derivatization with N-phenyl-1,2-phenylenediamine

A method for determination of alpha-ketoisocaproic acid (KIC) and [4,5,5,5,6,6,6-2H7]alpha-ketoisocaproic acid ([2H7]KIC) in rat plasma was developed using gas chromatography-mass spectrometry-selected ion monitoring (GC-MS-SIM). [5,5,5-2H3]alpha-Ketoisocaproic acid ([2H3]KIC) was used as an analytical internal standard to account for losses associated with the extraction, derivatization and chromatography. The keto acids were extracted by cation-exchange chromatography using BondElut SCX cartridge and derivatized with N-phenyl-1,2-phenylenediamine to form N-phenylquinoxalinone derivatives. Quantitation was performed by SIM of the respective molecular ions at m/z 278, 281 and 285 for the derivatives of KIC, [2H3]KIC and [2H7]KIC on the electron impact method. The limit of detection was found to be 70 fmol per injection (S/N=3) and the limit of quantitation for [2H7]KIC was around 50 nM in rat plasma. Endogenous KIC concentrations in 50 microl of rat plasma were measured with relative intra- and inter-day precision of 4.0% and 3.3%, respectively. The intra- and inter-day precision for [2H7]KIC spiked to rat plasma in the range of 0.1 to 10 microM gave good reproducibility with relative standard deviation (RSD) of 6.5% and 5.4%, respectively. The intra- and inter-day relative errors (RE) for [2H7]KIC were less than 6.4% and 3.8%, respectively. The method was applied to determine the plasma concentration of [2H7]KIC after an intravenous administration of [2H7]KIC in rat.

Sex Differences in Shotgun Proteome Analyses for Chronic Oral Intake of Cadmium in Mice

Environmental diseases related to cadmium exposure primarily develop owing to industrial wastewater pollution and/or contaminated food. In regions with high cadmium exposure in Japan, cadmium accumulation occurs primarily in the kidneys of individuals who are exposed to the metal. In contrast, in the itai-itai disease outbreak that occurred in the Jinzu River basin in Toyama Prefecture in Japan, cadmium primarily accumulated in the liver. On the other hand, high concentration of cadmium caused renal tubular disorder and osteomalacia (multiple bone fracture), probably resulting from the renal tubular dysfunction and additional pathology. In this study, we aimed to establish a mouse model of chronic cadmium intake. We administered cadmium-containing drinking water (32 mg/l) to female and male mice ad libitum for 11 weeks. Metal analysis using inductively coupled plasma mass spectrometry revealed that cadmium accumulated in the kidneys (927 x 10 + 185 ng/g in females and 661 x 10 + 101 ng/g in males), liver (397 x 10 + 199 ng/g in females and 238 x 10 + 652 ng/g in males), and thyroid gland (293 + 93.7 ng/g in females and 129 + 72.7 ng/g in males) of mice. Female mice showed higher cadmium accumulation in the kidney, liver, and thyroid gland than males did (p = 0.00345, p = 0.00213, and p = 0.0331, respectively). Shotgun proteome analyses after chronic oral administration of cadmium revealed that protein levels of glutathione S-transferase Mu2, Mu4, and Mu7 decreased in the liver, and those of A1 and A2 decreased in the kidneys in both female and male mice.

Cross-sectional Study on the Effects of Socioeconomic Factors on Lead Exposure in Children by Gender in Serpong, Indonesia

To elucidate the socioeconomic factors influencing lead exposure in elementary school children by gender, 108 children (56 male, 52 female), aged 6–7 years, were randomly selected from 39 elementary state schools in Serpong, Banten, Indonesia. Their parents were interviewed to obtain information on sociodemographic characteristics. Their blood lead (BPb) levels were measured by atomic absorption spectrophotometry. BPb concentrations were significantly higher in males than in females, i.e., 6.8 ± 2.0 (2.9–12.5) µg/dL and 5.9 ± 1.9 (3.1–11.7) µg/dL, respectively (p < 0.05). Lower socioeconomic status and well water use were associated with increased BPb concentrations, especially in females. The proportion of well water use was related to lower socioeconomic status. Lower socioeconomic status linked with well water drinking seemed to be associated with increased lead exposure in children in Serpong. Their exposure levels possibly varied according to gender differences in behavior. An intervention should be instituted among children in Serpong with BPb concentrations of 10 µg/dL or above.