Magnesium deficiency with high calcium-to-magnesium ratio promotes a metastatic phenotype in the CT26 colon cancer cell line.

Magnesium (Mg) plays important roles in maintaining genomic stability and cellular redox. Mg also serves as nature's physiological calcium (Ca) channel antagonist, controlling intracellular Ca entry. Because Ca is the most important second messenger, its intracellular concentration is tightly regulated. Excess intracellular Ca can activate aberrant signaling pathways leading to the acquisition of pathological characteristics and cell injury. Several epidemiological studies have linked Mg deficiency (MgD) and increased Ca:Mg ratios with higher incidences of colon cancer and increased mortality. While it is estimated that less than 50% of the US population consumes the recommended daily allowance for Mg, Ca supplementation is widespread. Therefore, we studied the effect of MgD, with variable Ca:Mg ratios on cellular oxidative stress, cell migration, calpain activity, and associated signaling pathways using the CT26 colon cancer cell line. MgD (with Ca:Mg ratios >1) elevated intracellular Ca levels, calpain activity and TRPM7 expression, as well as oxidative stress and cell migration, consistent with observed degradation of full-length E-cadherin, ß-catenin, and N-terminal FAK. MgD was accompanied by enhanced degradation of IκBα and the transactivation domain containing the C-terminus of NF-κB p65 (RelA). MgD-exposed CT26 cells exhibited increased p53 degradation and aneuploidy, markers of genomic instability. By contrast, these pathological changes were not observed when CT26 were cultured under MgD conditions where the Ca:Mg ratio was kept at 1. Together, these data support that exposure of colon cancer cells to MgD with physiological Ca concentrations (or increasing Ca:Mg ratios) leads to the acquisition of a more aggressive, metastatic phenotype.

Magnesium improves cisplatin-mediated tumor killing while protecting against cisplatin-induced nephrotoxicity.

Approximately 30% of all cancer patients treated with cisplatin, a widely used broad-spectrum chemotherapeutic agent, experience acute kidney injury (AKI). Almost all patients receiving cisplatin have magnesium (Mg) losses, which are proposed to aggravate AKI. Currently, there are no methods to successfully treat or prevent cisplatin-AKI. Whereas Mg supplementation has been shown to reduce AKI in experimental models and several small clinical trials, the effects of Mg status on tumor outcomes in immunocompetent tumor-bearing mice and humans have not been investigated. The purpose of this study was to further examine the effects of Mg deficiency (±Mg supplementation) on cisplatin-mediated AKI and tumor killing in immunocompetent mice bearing CT26 colon tumors. Using a model where cisplatin alone (20 mg/kg cumulative dose) produced minimal kidney injury, Mg deficiency significantly worsened cisplatin-mediated AKI, as determined by biochemical markers (blood urea nitrogen and plasma creatinine) and histological renal changes, as well as markers of renal oxidative stress, inflammation, and apoptosis. By contrast, Mg supplementation blocked cisplatin-induced kidney injury. Using LLC-PK1 renal epithelial cells, we observed that Mg deficiency or inhibition of Mg uptake significantly enhanced cisplatin-induced cytotoxicity, whereas Mg supplementation protected against cytotoxicity. However, neither Mg deficiency nor inhibition of Mg uptake impaired cisplatin-mediated killing of CT26 tumor cells in vitro. Mg deficiency was associated with significantly larger CT26 tumors in BALB/c mice when compared with normal-fed control mice, and Mg deficiency significantly reduced cisplatin-mediated tumor killing in vivo. Finally, Mg supplementation did not compromise cisplatin's anti-tumor efficacy in vivo.

Magnesium protects against cisplatin-induced acute kidney injury without compromising cisplatin-mediated killing of an ovarian tumor xenograft in mice.

Cisplatin, a commonly used chemotherapeutic for ovarian and other cancers, leads to hypomagnesemia in most patients and causes acute kidney injury (AKI) in 25-30% of patients. Previously, we showed that magnesium deficiency worsens cisplatin-induced AKI and magnesium replacement during cisplatin treatment protects against cisplatin-mediated AKI in non-tumor-bearing mice (Solanki MH, Chatterjee PK, Gupta M, Xue X, Plagov A, Metz MH, Mintz R, Singhal PC, Metz CN. Am J Physiol Renal Physiol 307: F369-F384, 2014). This study investigates the role of magnesium in cisplatin-induced AKI using a human ovarian tumor (A2780) xenograft model in mice and the effect of magnesium status on tumor growth and the chemotherapeutic efficacy of cisplatin in vivo. Tumor progression was unaffected by magnesium status in saline-treated mice. Cisplatin treatment reduced tumor growth in all mice, irrespective of magnesium status. In fact, cisplatin-treated magnesium-supplemented mice had reduced tumor growth after 3 wk compared with cisplatin-treated controls. While magnesium status did not interfere with tumor killing by cisplatin, it significantly affected renal function following cisplatin. Cisplatin-induced AKI was enhanced by magnesium deficiency, as evidenced by increased blood urea nitrogen, creatinine, and other markers of renal damage. This was accompanied by reduced renal mRNA expression of the cisplatin efflux transporter Abcc6. These effects were significantly reversed by magnesium replacement. On the contrary, magnesium status did not affect the mRNA expression of cisplatin uptake or efflux transporters by the tumors in vivo. Finally, magnesium deficiency enhanced platinum accumulation in the kidneys and renal epithelial cells, but not in the A2780 tumor cells. These findings demonstrate the renoprotective role of magnesium during cisplatin AKI, without compromising the chemotherapeutic efficacy of cisplatin in an ovarian tumor-bearing mouse model.

Omega-3 polyunsaturated fatty acids enhance cytokine production and oxidative stress in a mouse model of preterm labor.

OBJECTIVE: Omega-3 polyunsaturated fatty acid (ω-3 PUFA) supplementation during pregnancy remains controversial. We sought to examine the effects of ω-3 PUFA on inflammation and oxidative stress in vitro and in vivo using a model of preterm labor. METHODS: In vivo. Female Swiss Webster mice were fed a normal diet or a 5% fish oil (FO) diet for 3 weeks then mated with normal-fed males. On gestational day 15, dams were injected with either saline (n=10 per group) or lipopolysaccharide (LPS, intrauterine) (n=10 per group). Maternal plasma, amniotic fluid, placentas, and uteri were collected 4 h later and assessed for cytokines; maternal plasma and amniotic fluids were analyzed for oxidative stress. In vitro. RAW264.7 mouse macrophage-like cells were treated with either: vehicle, H2O2, docosahexaenoic acid (DHA), or eicosapentaenoic acid (EPA) (0, 0.1-100 µM) and analyzed for oxidative stress. RESULTS: In vivo. Administration of the 5% FO diet enhanced LPS-induced cytokines in the placenta (P<0.05-0.01) and increased tumor necrosis factor-α in the uterus (P<0.05) and amniotic fluid (P<0.01) when compared to LPS-treated normal-fed animals. Maternal plasma obtained from FO-fed dams showed higher LPS-induced oxidative stress than control-fed animals (P<0.035). However, no differences in oxidative stress were observed in the amniotic fluid. In vitro. Treatment of macrophage-like cells with ω-3 PUFA significantly and dose-dependently increased oxidative stress (P<0.001-0.0001). CONCLUSIONS: Supplementation with FO for prior to and during pregnancy significantly increased LPS-induced inflammation in the amniotic fluid, uterus, and placenta and significantly increased maternal systemic oxidative stress in vivo. Likewise, DHA and EPA induced oxidative stress in macrophage-like cells in vitro.

Sacral nerve stimulation reduces elevated urinary nerve growth factor levels in women with symptomatic detrusor overactivity.

OBJECTIVES: To investigate changes in urinary nerve growth factor (uNGF) in women with symptomatic detrusor overactivity (DO) following peripheral nerve evaluation (PNE) for sacral neuromodulation vs controls. STUDY DESIGN: There were 23 subjects with overactive bladder symptoms and DO who failed management with anticholinergics and 22 controls consented to participate in this prospective pilot study. Urine specimens were collected from controls at baseline for evaluation of uNGF and creatinine. Subjects were evaluated at baseline and 5 days after a trial of sacral nerve stimulation referred to as a PNE. Each visit included urine collection for uNGF and, Incontinence Quality of Life Questionnaire, Urinary Distress Inventory Questionnaire, postvoid residual volume, and a 3-day voiding diary. uNGF levels were measured by enzyme-linked immunosorbent assay and expressed as uNGF pg/creatinine mg. RESULTS: Subjects with DO had significantly higher baseline uNGF levels (corrected for creatinine) compared with controls (19.82 pg/mg vs 7.88 pg/mg, P < .002). Seventeen DO subjects underwent PNE and were evaluated at the end of the testing period. There was a significant improvement in quality of life scores for subjects after PNE compared with baseline (Urinary Distress Inventory Questionnaire: 7.0 vs 13.7, P < .001; Incontinence Quality of Life Questionnaire: 87.3 vs 52.8, P < .0001). Concordantly, uNGF levels significantly decreased from 17.23 pg/mg to 9.24 pg/mg (P < .02) after PNE. CONCLUSION: uNGF levels decrease with symptomatic response in DO subjects undergoing PNE. DO subjects had significantly higher uNGF at baseline vs controls, and uNGF levels significantly decreased after only 5 days of sacral nerve stimulation. These findings support a larger study to validate the use of uNGF as an objective tool to assess therapeutic outcome in patients undergoing PNE for sacral neuromodulation.

Magnesium protects against cisplatin-induced acute kidney injury by regulating platinum accumulation.

Despite its success as a potent antineoplastic agent, ∼25% of patients receiving cisplatin experience acute kidney injury (AKI) and must discontinue therapy. Impaired magnesium homeostasis has been linked to cisplatin-mediated AKI, and because magnesium deficiency is widespread, we examined the effect of magnesium deficiency and replacement on cisplatin-induced AKI in physiologically relevant older female mice. Magnesium deficiency significantly increased cisplatin-associated weight loss and markers of renal damage (plasma blood urea nitrogen and creatinine), histological changes, inflammation, and renal cell apoptosis and modulated signaling pathways (e.g., ERK1/2, p53, and STAT3). Conversely, these damaging effects were reversed by magnesium. Magnesium deficiency alone significantly induced basal and cisplatin-mediated oxidative stress, whereas magnesium replacement attenuated these effects. Similar results were observed using cisplatin-treated LLC-PK1 renal epithelial cells exposed to various magnesium concentrations. Magnesium deficiency significantly amplified renal platinum accumulation, whereas magnesium replacement blocked the augmented platinum accumulation after magnesium deficiency. Increased renal platinum accumulation during magnesium deficiency was accompanied by reduced renal efflux transporter expression, which was reversed by magnesium replacement. These findings demonstrate the role of magnesium in regulating cisplatin-induced AKI by enhancing oxidative stress and thus promoting cisplatin-mediated damage. Additional in vitro experiments using ovarian, breast, and lung cancer cell lines showed that magnesium supplementation did not compromise cisplatin's chemotherapeutic efficacy. Finally, because no consistently successful therapy to prevent or treat cisplatin-mediated AKI is available for humans, these results support developing more conservative magnesium replacement guidelines for reducing cisplatin-induced AKI in cancer patients at risk for magnesium deficiency.

Maternal magnesium supplementation reduces intrauterine growth restriction and suppresses inflammation in a rat model.

OBJECTIVE: Intrauterine growth restriction (IUGR) is associated with increased inflammatory responses. We sought to investigate whether magnesium (Mg) attenuates inflammation and IUGR in a rat model. STUDY DESIGN: Pregnant Wistar rats (12 weeks, gestational day 18) were randomly assigned to 1 of 4 groups: normal diet with bilateral uterine artery ligation (BL) (n = 6) or sham surgery (SH) (n = 5); and Mg chloride (MgCl2) 1% (wt/vol) in the drinking water throughout gestation + BL (MgBL) (n = 6) or SH (MgSH) (n = 5). Dams were euthanized 24 hours postsurgery (gestational day 19). Maternal plasma, fetal plasma (pooled), individual amniotic fluid (AF) samples, and placentas (PL) were collected and assessed from live fetal pups only (BL, n = 36; SH, n = 20; MgBL, n = 20; MgSH, n = 20). All samples were analyzed for cytokines/chemokines (interleukin [IL]-6, IL-1ß, chemokine [C-X-C motif] ligand 1 [CXCL1], chemokine [C-C motif] ligand 2 [CCL2], and tumor necrosis factor [TNF-α] sensitivity <3 pg/mL) using a multiplex platform. Data were analyzed using Mann Whitney, analysis of variance, and Fisher exact tests. RESULTS: The incidence of IUGR (pup weight <10th percentile of SH) in the MgBL group was significantly lower (31%) than the BL group (86.3%) (relative risk, 0.36; 95% confidence interval, 0.2-0.6; P < .0001). BL significantly increased AF levels of IL-6, IL-1ß, TNF-α (P < .05), and CCL2 (P < .001) vs SH and PL levels of IL-6, IL-1ß, CCL2 and CXCL1 (P < .001), and TNF-α (P < .05) vs SH. Maternal MgCl2 supplementation significantly decreased IL-1ß, TNF-α, and CCL2 levels in AF and IL-1ß in PL tissues of MgBL vs BL rats (P < .0001). CONCLUSION: Maternal oral MgCl2 supplementation reduced BL-induced IUGR by 64% and suppressed cytokine/chemokine levels in the AF and PL.