ClC-2-like Chloride Current Alterations in a Cell Model of Spinal and Bulbar Muscular Atrophy, a Polyglutamine Disease.

Spinal and bulbar muscular atrophy (SBMA) is a neuromuscular disease caused by expansions of a polyglutamine (polyQ) tract in the androgen receptor (AR) gene. SBMA is associated with the progressive loss of lower motor neurons, together with muscle weakness and atrophy. PolyQ-AR is converted to a toxic species upon binding to its natural ligands, testosterone, and dihydrotestosterone (DHT). Our previous patch-clamp studies on a motor neuron-derived cell model of SBMA showed alterations in voltage-gated ion currents. Here, we identified and characterized chloride currents most likely belonging to the chloride channel-2 (ClC-2) subfamily, which showed significantly increased amplitudes in the SBMA cells. The treatment with the pituitary adenylyl cyclase-activating polypeptide (PACAP), a neuropeptide with a proven protective effect in a mouse model of SBMA, recovered chloride channel current alterations in SBMA cells. These observations suggest that the CIC-2 currents are affected in SBMA, an alteration that may contribute and potentially determine the pathophysiology of the disease.

Nutritional Guideline for the Management of Mexican Patients with CKD and Hyperphosphatemia.

Chronic kidney disease (CKD) represents a serious concern for the Mexican population since the main predisposing diseases (diabetes, hypertension, etc.) have a high prevalence in the country. The development of frequent comorbidities during CKD such as anemia, metabolic disorders, and hyperphosphatemia increases the costs, symptoms, and death risks of the patients. Hyperphosphatemia is likely the only CKD comorbidity in which pharmaceutical options are restricted to phosphate binders and where nutritional management seems to play an important role for the improvement of biochemical and clinical parameters. Nutritional interventions aiming to control serum phosphate levels need to be based on food tables, which should be specifically elaborated for the cultural context of each population. Until now, there are no available food charts compiling a high amount of Mexican foods and describing phosphorus content as well as the phosphate to protein ratio for nutritional management of hyperphosphatemia in CKD. In this work, we elaborate a highly complete food chart as a reference for Mexican clinicians and include charts of additives and drug phosphate contents to consider extra sources of inorganic phosphate intake. We aim to provide an easy guideline to contribute to the implementation of more nutritional interventions focusing on this population in the country.

Thermal stability of human matrix metalloproteinases.

Matrix metalloproteinases (MMP) are key players in the remodelling of the extracellular matrix under physiological and pathological conditions. Thermodynamic parameters of human recombinant metalloproteinases of the active (rMMP2, 3, 7, 8 and 9) and latent (rPro-MMP2, 3 and 9) forms were obtained by differential scanning calorimetry (DSC). Temperature by itself does not result in autocatalysis of recombinant MMP. The transitions observed by DSC correspond to structural domains of the monomeric protein. In this study, we show the domain organization of these proteins, where the thermal transition (Tm) of the main component is observed at 71.3 °C (ProMMP-2); 74.8 °C (ProMMP-8); 80.0 °C (ProMMP-3); 92.6 °C (ProMMP-9) and 98.3 °C (ProMMP-7). For MMP-3, this main Tm is related to the catalytic domain (CD). The isolated recombinant CD of MMP-3 unfolds as a single transition at Tm 83.4 °C, matching the more stable domain observed in the full-length active form of rMMP-3. The denaturation profile of rProMMP-3 shows the main transition at Tm 80 °C, a less stable domain before the propeptide domain (PD) cleavage. Our results indicate that the structural stability of MMP and particularly their CD are not substantially altered after cleavage of the PD. We propose that the thermodynamic parameters obtained by DSC are relevant for the functional study of MMP, particularly to reveal their contribution in complex biological samples in health and disease.

IL-1ß induced methylation of the estrogen receptor ERα gene correlates with EMT and chemoresistance in breast cancer cells.

Inflammation has been recently acknowledged as a key participant in the physiopathology of oncogenesis and tumor progression. The inflammatory cytokine IL-1ß has been reported to induce the expression of markers associated with malignancy in breast cancerous cells through Epithelial-Mesenchymal Transition (EMT). Aggressive breast cancer tumors classified as Triple Negative do not respond to hormonal treatment because they lack three crucial receptors, one of which is the estrogen receptor alpha (ERα). Expression of ERα is then considered a good prognostic marker for tamoxifen treatment of this type of cancer, as the binding of this drug to the receptor blocks the transcriptional activity of the latter. Although it has been suggested that inflammatory cytokines in the tumor microenvironment could regulate ERα expression, the mechanism(s) involved in this process have not yet been established. We show here that, in a cell model of breast cancer cells (6D cells), in which the inflammatory cytokine IL-1ß induces EMT by activation of the IL-1ß/IL-1RI/ß-catenin pathway, the up regulation of TWIST1 leads to methylation of the ESR1 gene promoter. This epigenetic modification produced significant decrease of the ERα receptor levels and increased resistance to tamoxifen. The direct participation of IL-1ß in these processes was validated by blockage of the cytokine-induced signaling pathway by wortmannin inactivation of the effectors PI3K/AKT. These results support our previous reports that have suggested direct participation of the inflammatory cytokine IL-1ß in the transition to malignancy of breast cancer cells.

Altered ionic currents and amelioration by IGF-1 and PACAP in motoneuron-derived cells modelling SBMA.

Spinal and bulbar muscular atrophy (SBMA), also known as Kennedy's disease, is a motor neuron disease caused by the expansion of a polymorphic CAG tandem repeat encoding a polyglutamine (polyQ) tract in the androgen receptor (AR) gene. SBMA is triggered by the binding of mutant AR to its natural ligands, testosterone and dihydrotestosterone (DHT). To investigate the neuronal alterations of motor neuron cell models of SBMA, we applied patch-clamp methods to verify how polyQ expansions in the AR alter cell ionic currents. We used mouse motoneuron-derived MN-1 cells expressing normal AR (MN24Q) and mutant AR (MN100Q treated cells with vehicle EtOH and DHT). We observed a reduction of the current flux mainly at depolarizing potentials in the DHT-treated cells, while the dissection of macroscopic currents showed single different cationic currents belonging to voltage-gated channels. Also, we treated the cells with IGF-1 and PACAP, which have previously been shown to protect MN-1 cells from the toxicity of mutant AR, and we found an amelioration of the altered currents. Our results suggest that the electrophysiological correlate of SBMA is a suitable reference point for the identification of disease symptoms and for future therapeutic targets.

KV10.1 K(+)-channel plasma membrane discrete domain partitioning and its functional correlation in neurons.

KV10.1 potassium channels are implicated in a variety of cellular processes including cell proliferation and tumour progression. Their expression in over 70% of human tumours makes them an attractive diagnostic and therapeutic target. Although their physiological role in the central nervous system is not yet fully understood, advances in their precise cell localization will contribute to the understanding of their interactions and function. We have determined the plasma membrane (PM) distribution of the KV10.1 protein in an enriched mouse brain PM fraction and its association with cholesterol- and sphingolipid-rich domains. We show that the KV10.1 channel has two different populations in a 3:2 ratio, one associated to and another excluded from Detergent Resistant Membranes (DRMs). This distribution of KV10.1 in isolated PM is cholesterol- and cytoskeleton-dependent since alteration of those factors changes the relationship to 1:4. In transfected HEK-293 cells with a mutant unable to bind Ca(2+)/CaM to KV10.1 protein, Kv10.1 distribution in DRM/non-DRM is 1:4. Mean current density was doubled in the cholesterol-depleted cells, without any noticeable effects on other parameters. These results demonstrate that recruitment of the KV10.1 channel to the DRM fractions involves its functional regulation.