Nuria Olea-Herrero

Anti-tumoral action of cannabinoids on hepatocellular carcinoma: role of AMPK-dependent activation of autophagy.

Hepatocellular carcinoma (HCC) is the third cause of cancer-related death worldwide. When these tumors are in advanced stages, few therapeutic options are available. Therefore, it is essential to search for new treatments to fight this disease. In this study, we investigated the effects of cannabinoids – a novel family of potential anticancer agents – on the growth of HCC. We found that Δ9-tetrahydrocannabinol (Δ9-THC, the main active component of Cannabis sativa) and JWH-015 (a cannabinoid receptor 2 (CB2) cannabinoid receptor-selective agonist) reduced the viability of the human HCC cell lines HepG2 (human hepatocellular liver carcinoma cell line) and HuH-7 (hepatocellular carcinoma cells), an effect that relied on the stimulation of CB2 receptor. We also found that Δ9-THC- and JWH-015-induced autophagy relies on tribbles homolog 3 (TRB3) upregulation, and subsequent inhibition of the serine–threonine kinase Akt/mammalian target of rapamycin C1 axis and adenosine monophosphate-activated kinase (AMPK) stimulation. Pharmacological and genetic inhibition of AMPK upstream kinases supported that calmodulin-activated kinase kinase β was responsible for cannabinoid-induced AMPK activation and autophagy. In vivo studies revealed that Δ9-THC and JWH-015 reduced the growth of HCC subcutaneous xenografts, an effect that was not evident when autophagy was genetically of pharmacologically inhibited in those tumors. Moreover, cannabinoids were also able to inhibit tumor growth and ascites in an orthotopic model of HCC xenograft. Our findings may contribute to the design of new therapeutic strategies for the management of HCC.

Capsaicin, a component of red peppers, induces expression of androgen receptor via PI3K and MAPK pathways in prostate LNCaP cells.

In this study, capsaicin (trans‐8‐methyl‐N‐vanillyl‐6‐nonenamide) induced an increase in the cell viability of the androgen‐responsive prostate cancer LNCaP cells, which was reversed by the use of the TRPV1 antagonists capsazepine, I‐RTX and SB 366791. In further studies we observed that capsaicin induced a decrease in ceramide levels as well as Akt and Erk activation. To investigate the mechanism of capsaicin action we measured androgen (AR) receptor levels. Capsaicin induced an increase in the AR expression that was reverted by the three TRPV1 antagonists. AR silencing by the use of siRNA, as well as blocking the AR receptor with bicalutamide, inhibited the proliferative effect of capsaicin.

The vanilloid capsaicin induces IL-6 secretion in prostate PC-3 cancer cells.

Capsaicin (8-methyl-N-vanillyl-6-nonenamide), a constituent of green and red peppers, has been linked with suppression of tumorigenesis through a mechanism that is not well understood. In the present study, we examined the effects of capsaicin on the production of the cytokine interleukin (IL)-6 by PC-3 cells at both protein and mRNA levels which were evaluated by ELISA and real-time PCR, respectively. Capsaicin-treated PC-3 cells increased the synthesis and secretion of IL-6 which was abrogated by the transient receptor potential vanilloid receptor subtype 1 (TRPV1) antagonist capsazepine, as well as by inhibitors of PKC-α, phosphoinositol-3 phosphate kinase (PI-3K), Akt and extracellular signal-regulated protein kinase (ERK). We analyzed the role of capsaicin in the tumor necrosis factor (TNF)-α secretion by PC-3 cells which was increased at shorter times than IL-6 production. Furthermore, incubation of PC-3 cells with an anti-TNF-α antibody blocked the capsaicin-induced IL-6 secretion. These results raise the possibility that capsaicin-mediated IL-6 increase in prostate cancer PC-3 cells is regulated at least in part by TNF-α secretion and signaling pathway involving Akt, ERK and PKC-α activation.

Oral administration of bisphenol A induces high blood pressure through angiotensin II/CaMKII-dependent uncoupling of eNOS

Bisphenol A (BPA) is found in human urine and fat tissue. Higher urinary BPA concentrations are associated with arterial hypertension. To shed light on the underlying mechanism, we orally administered BPA (4 nM to 400 μM in drinking water) to 8‐wk‐old CD11 mice over 30 d. Mice developed dosage‐dependent high blood pressure (systolic 130±12 vs. 170±12 mmHg; EC50 0.4 μM), impairment of acetylcholine (AcH)‐induced carotid relaxation (0.66±0.08 vs. 0.44±0.1 mm), a 1.7‐fold increase in arterial angiotensin II (AngII), an 8.7‐fold increase in eNOS mRNA and protein, and significant eNOS‐dependent superoxide and peroxynitrite accumulation. AngII inhibition with 0.5 mg/ml losartan reduced oxidative stress and normalized blood pressure and endothelium‐dependent relaxation, which suggests that AngII uncouples eNOS and contributes to the BPA‐induced endothelial dysfunction by promoting oxidative and nitrosative stress. Microarray analysis of mouse aortic endothelial cells revealed a 2.5‐fold increase in expression of calcium/calmodulin‐dependent protein kinase II‐α (CaMKII‐α) in response to 10 nM BPA, with increased expression of phosphorylated‐CaMKII‐α in carotid rings of BPA‐exposed mice, whereas CaMKII‐α inhibition with 100 nM autocamptide‐2‐related inhibitor peptide (AIP) reduced BPA‐mediated increase of superoxide. Administration of CaMKII‐α inhibitor KN 93 reduced BPA‐induced blood pressure and carotid blood velocity in mice, and reverted BPA‐mediated carotid constriction in response to treatment with AcH. Given that CaMKII‐α inhibition prevents BPA‐mediated high blood pressure, our data suggest that BPA regulates blood pressure by inducing AngII/CaMKII‐α uncoupling of eNOS.—Saura, M., Marquez, S., Reventun, P., Olea‐Herrero, N., Arenas, M.I., Moreno‐Gómez‐Toledano, R., Gómez‐Parrizas, M., Muñóz‐Moreno, C., González‐Santander, M., Zaragoza, C., Bosch, R.J. Oral administration of bisphenol A induces high blood pressure through angiotensin II/CaMKII‐dependent uncoupling of eNOS. FASEB J. 28, 4719–4728 (2014). www.fasebj.org

Bisphenol‐A Induces Podocytopathy With Proteinuria in Mice

Bisphenol‐A, a chemical used in the production of the plastic lining of food and beverage containers, can be found in significant levels in human fluids. Recently, bisphenol‐A has been associated with low‐grade albuminuria in adults as well as in children. Since glomerular epithelial cells (podocytes) are commonly affected in proteinuric conditions, herein we explored the effects of bisphenol‐A on podocytes in vitro and in vivo. On cultured podocytes we first observed that bisphenol‐A—at low or high concentrations—(10 nM and 100 nM, respectively) was able to induce hypertrophy, diminish viability, and promote apoptosis. We also found an increase in the protein expression of TGF‐β1 and its receptor, the cyclin‐dependent kinase inhibitor p27Kip1, as well as collagen‐IV, while observing a diminished expression of the slit diaphragm proteins nephrin and podocin. Furthermore, mice intraperitoneally injected with bisphenol‐A (50 mg/Kg for 5 weeks) displayed an increase in urinary albumin excretion and endogenous creatinine clearance. Renal histology showed mesangial expansion. At ultrastructural level, podocytes displayed an enlargement of both cytoplasm and foot processes as well as the presence of condensed chromatin, suggesting apoptosis. Furthermore, immunohistochemistry for WT‐1 (specific podocyte marker) and the TUNEL technique showed podocytopenia as well as the presence of apoptosis, respectively. In conclusion, our data demonstrate that Bisphenol‐A exposure promotes a podocytopathy with proteinuria, glomerular hyperfiltration and podocytopenia. Further studies are needed to clarify the potential role of bisphenol‐A in the pathogenesis as well as in the progression of renal diseases. J. Cell. Physiol. 229: 2057–2066, 2014.

The cannabinoid R+ methanandamide induces IL-6 secretion by prostate cancer PC3 cells.

In the present study, we have investigated the effect of the cannabinoid R(+)methanandamide (MET) in the androgen-resistant prostate cancer PC3 cells. MET induced a dose-dependent decrease in PC3 cell viability as well as a dose-dependent increase in the secretion of the cytokine IL-6. Looking deeper into the mechanisms involved, we found that MET-induced de novo synthesis of the lipid mediator ceramide that was blocked by the ceramide synthase inhibitor Fumonisin B1. Pre-incubation of cells with the cannabinoid receptor CB2 antagonist SR 144528 (SR2), but not the CB1 antagonist Rimonabant or the TRPV1 antagonist capsazepine, partially prevented the anti-proliferative effect, the ceramide accumulation, and the IL-6-induced secretion, suggesting a CB2 receptor-dependent mechanism. Fumonisin B1 did not have any effect in the IL-6 secretion increase induced by MET. However, even an incomplete down-regulation of (i.e., not a total silencing of) ceramide kinase expression by specific siRNA prevented the MET-induced IL-6 secretion. These results suggest that MET regulates ceramide metabolism in prostate PC3 cells which is involved in cell death as well as in IL-6 secretion. Our findings also suggest that CB2 agonists may offer a novel approach in the treatment of prostate cancer by decreasing cancer epithelial cell proliferation. However, the interaction of prostate cancer cells with their surrounding, and in particular with the immune system in vivo, needs to be further explored.

El bisfenol A: un factor ambiental implicado en el daño nefrovascular

Ricardo J. Bosch, Borja Quiroga, Carmen Muñoz-Moreno, Nuria Olea-Herrero, María Isabel Arenas , Marta González-Santander, Paula Reventún , Carlos Zaragoza , Gabriel de Arriba y Marta Saura e a Laboratorio de Fisiología Renal y Nefrología Experimental, Unidad de Fisiología, Departamento de Biología de Sistemas, Universidad de Alcalá, Alcalá de Henares, España b Servicio de Nefrología, Hospital Universitario de Guadalajara, Guadalajara, España c Unidad de Biología Celular, Departamento de Biomedicina y Biotecnología, Universidad de Alcalá, Alcalá de Henares, España d Departamento de Medicina y Especialidades Médicas, Universidad de Alcalá, Alcalá de Henares, España e Laboratorio de Fisiopatología de la Pared Vascular, Unidad de Fisiología, Departamento de Biología de Sistemas, Universidad de Alcalá, Alcalá de Henares, España f Laboratorio de Fisiopatología Cardiovascular, Unidad de Investigación Translacional, Facultad de Medicina, Universidad Francisco de Vitoria, Madrid, España

Online Hemodiafiltration Reduces Bisphenol A Levels

Several uremic toxins have been identified and related to higher rates of morbidity and mortality in dialysis patients. Bisphenol A (BPA) accumulates in patients with chronic kidney disease. The aim of this study is to demonstrate the usefulness of online hemodiafiltration (OL‐HDF) in reducing BPA levels. Thirty stable hemodialysis patients were selected to participate in this paired study. During three periods of 3 weeks each, patients were switched from high‐flux hemodialysis (HF‐HD) to OL‐HDF, and back to HF‐HD. BPA levels were measured in the last session of each period (pre‐ and post‐dialysis) using ELISA and HPLC. Twenty‐two patients (mean age 73 ± 14 years; 86.4% males) were included. Measurements of BPA levels by HPLC and ELISA assays showed a weak but significant correlation (r = 0.218, P = 0.012). BPA levels decreased in the OL‐HDF period of hemodialysis, in contrast to the HF‐HD period when they remained stable (P = 0.002). In conclusion, OL‐HDF reduced BPA levels in dialysis patients.