Mitochondria are key organelles for cellular metabolism, and regulate several processes including cell death and macroautophagy/autophagy. Here, we show that mitochondrial respiratory chain (RC) deficiency deactivates AMP-activated protein kinase (AMPK, a key regulator of energy homeostasis) signaling in tissue and in cultured cells. The deactivation of AMPK in RC-deficiency is due to increased expression of the AMPKinhibiting protein FLCN (folliculin). AMPK is found to be necessary for basal lysosomal function, and AMPK deactivation in RC-deficiency inhibits lysosomal function by decreasing the activity of the lysosomal Ca 2+ channel MCOLN1 (mucolipin 1). MCOLN1 is regulated by phosphoinositide kinase PIKFYVE and its product PtdIns(3,5)P 2 , which is also decreased in RC-deficiency. Notably, reactivation of AMPK, in a PIKFYVEdependent manner, or of MCOLN1 in RC-deficient cells, restores lysosomal hydrolytic capacity. Building on these data and the literature, we propose that downregulation of the AMPK-PIKFYVE-PtdIns(3,5)P 2-MCOLN1 pathway causes lysosomal Ca 2+ accumulation and impaired lysosomal catabolism. Besides unveiling a novel role of AMPK in lysosomal function, this study points to the mechanism that links mitochondrial malfunction to impaired lysosomal catabolism, underscoring the importance of AMPK and the complexity of organelle cross-talk in the regulation of cellular homeostasis.

A B S T R A C TDoxorubicin (DOX) is one of the most widely used anti-neoplastic agents. However, treatment with DOX is associated with cumulative cardiotoxicity inducing progressive cardiomyocyte death. Sirtuin 3 (Sirt3), a mitochondrial deacetylase, regulates the activity of proteins involved in apoptosis, autophagy and metabolism. Our hypothesis is that pharmacological modulation by berberine (BER) pre-conditioning of Sirt3 protein levels decreases DOX-induced cardiotoxicity. Our results showed that DOX induces cell death in all experimental groups. Increase in Sirt3 content by transfection-mediated overexpression decreased DOX cytotoxicity, mostly by maintaining mitochondrial network integrity and reducing oxidative stress. p53 was upregulated by DOX, and appeared to be a direct target of Sirt3, suggesting that Sirt3-mediated protection against cell death could be related to this protein. BER pre-treatment increased Sirt3 and Sirt1 protein levels in the presence of DOX and inhibited DOX-induced caspase 9 and 3-like activation. Moreover, BER modulated autophagy in DOX-treated H9c2 cardiomyoblasts. Interestingly, mitochondrial biogenesis markers were upregulated in in BER/DOX-treated cells. Sirt3 over-expression contributes to decrease DOX cytotoxicity on H9c2 cardiomyoblasts, while BER can be used as a modulator of Sirtuin function and cell quality control pathways to decrease DOX toxicity.

Background Bisphenol A (BPA) is one of the most widely produced chemicals worldwide and is often used in the production of food and beverage containers. As a result of BPA contact with food, drink and toiletries, its ingestion and absorption by humans has been growing. The industrialization and modern lifestyles brought a constant exposure to several health-disturbing compounds and ushered a new era of chronic diseases. The endocrine disruptor potential of BPA is well known, but the research around its epigenotoxic effects raised further concerns whether chronic exposure to BPA can contribute to chronic human illness, including cancer in hormone-sensitive organs.

Although effective therapies to target mitochondria in the context of CVDs are not under widespread clinical use, the new strategies proposed constitute a real promise for the development of therapies which may effectively prevent CVDs in the near future.

Laboratory profile of young ovines was studied in order to evaluate and compare their antiserum production from natural and Cobalt-60 irradiated Crotalus durissus terrificus (C.d.t.) venoms. The parameters analyzed included complete blood count, and urea, creatinine, aspartate aminotransferase, total proteins, albumin and globulin serum measurements. Three groups of six animals each were used. Group 1 (G1) received natural C.d.t. venom; Group 2 (G2) received irradiated C.d.t. venom; and Group 3 (G3) was used as control and did not receive venom, only adjuvants, using seven venom inoculations. During the experimental period, animals were fortnightly weighed. According to clinical and weight evaluation, sheep in post-weaning phase showed no changes in their physiological profiles but had excellent weight gain. The parameters analyzed were not statistically different (p<5%) among the groups tested. The hyperimmunization process was successfully accomplished with the production of specific antibodies against Crotalus durissus terrificus venom. Results bring a new possibility of utilizing ovines in the commercial production of anticrotalic serum, which may be used to treat human and animal envenomation. Its production cost may be reduced by subsequent use of hyperimmunized sheep for human consumption

Neuro-hormonal regulation of cardiac function via cathecol-amines results in increased heart rate and contractility. A persistent adrenergic tone, however, is an insult to the heart, affecting its regular homeostasis, altering morphology and gene expression patterns, as well as inducing apoptosis of cardio-myocytes. At the same time as being the main oxygen consumers, mitochondria are also key to the energy production required for the heart to maintain its vital functions and to integrate a series of signaling pathways that define the life and death of the cell. As α-adrenergic receptors (α-AR) orchestrate multiple biochemical events that can either trigger or inhibit cell death, mitochondria can act as a referee in the entire process. In fact, α-AR subtypes α1 and α2 activate various down-stream pathways which differently modulate intracellular calcium levels and production of mitochondrial reactive oxygen species (ROS). The delicate balance between an adaptive (cardio-protective) response resulting in increased contractility and activation of survival pathways, vs. cell death caused by calcium and ROS-induced mitochondrial disruption, along with evidence of their clinical and potential therapeutic translations, are reviewed in this communication.

The adaptor protein p66Shc links membrane receptors to intracellular signaling pathways, with downstream consequences on mitochondrial metabolism and reactive oxygen species production. Moreover, p66Shc has also been implicated in cancer development, progression, and metastasis. Increased phosphorylation of serine 36 residue of p66Shc very often correlates with oxidative stress-associated pathologies. The pro-oxidative role of p66Shc also appears to be involved in chemical toxicity, being an important component of stress responses triggered by xenobiotics. Here, we present a protocol that can be used: (a) for isolation of mitochondrial, cytosolic, and mitochondrial-associated membrane fractions from adherent cells lines; (b) to perform p66Shc detection with specific antibodies in order to monitor its translocation between different cellular compartments in response to the oxidative stress; and (c) to modulate the p66Shc pathway with the use of pharmacological approaches or gene-silencing methods.

ABSTRACT:The unfavorable evolution of a young ovine during hyperimmunization process with Crotalus durissus terrificus venom was investigated in order to differentiate its origin between ophidic envenomation and copper toxicosis. Clinical, laboratory, necroscopic and histological exams as well as evaluation and measurement of heavy metals (copper) in the kidneys and in the liver were carried out. Blood counts revealed anemia and serological tests showed high levels of blood urea nitrogen, creatinine, aspartate aminotransferase, creatine phosphokinase, total bilirubin and indirect bilirubin; which indicates liver, kidney and skeletal muscle damages. At necropsy, the animal presented hepatopathy and nephropathy. Histological examination revealed renal and hepatic features that may imply copper intoxication. Copper levels were 237.8 µg/g in the liver and 51.2 µg/g in the kidneys. Although the amount of metal found in both organs was below the level that can cause death, according to the literature, anatomopathological signs were suggestive of copper intoxication. Therefore, the hypothesis of metal toxicosis during the hyperimmunization process became more consistent than the crotalic envenomation one.