Coupling mitogenesis and mitophagy for longevity

Maintenance of mitochondrial function and energy homeostasis requires both generation of newly synthesized and elimination of dysfunctional mitochondria. Impaired mitochondrial function and excessive mitochondrial content are major characteristics of aging and several human pathophysiological conditions, highlighting the pivotal role of the coordination between mitochondrial biogenesis and mitophagy. However, the cellular and molecular underpinnings of mitochondrial mass homeostasis remain obscure. In our recent study, we demonstrate that DCT-1, the Caenorhabditis elegans homolog of mammalian BNIP3 and BNIP3L/NIX, is a key mediator of mitophagy promoting longevity under stress. DCT-1 acts downstream of the PINK-1-PDR-1/Parkin pathway and is ubiquitinated upon mitophagy-inducing conditions to mediate the removal of damaged mitochondria. Accumulation of damaged mitochondria triggers SKN-1 activation, which initiates a bipartite retrograde signaling pathway stimulating the coordinated induction of both mitochondrial biogenesis and mitophagy genes. Taken together, our results unravel a homeostatic feedback loop that allows cells to adjust their mitochondrial population in response to environmental and intracellular cues. Age-dependent decline of mitophagy both inhibits removal of dysfunctional or superfluous mitochondria and impairs mitochondrial biogenesis resulting in progressive mitochondrial accretion and consequently, deterioration of cell function.     

Estimating dung decay rates of roe deer (<Emphasis Type="Italic">Capreolus capreolus</Emphasis>) in different habitat types of a Mediterranean ecosystem: an information theory approach

For elusive species living in concealing habitats (e.g. deer in a forest habitat), indirect methods such as faecal pellet counts are considered more practical means of estimating population density and abundance. Accurate estimation of deer density using the faecal standing crop (FSC) method necessitates the reliable estimation of the mean time to decay of pellet groups present during the survey. Mean time to decay is generally habitat specific, and separate estimations should be made for each habitat type in the study area. In a confined mountainous area of Greece, the habitat-specific mean time to decay of roe deer pellet groups was estimated by locating and marking fresh pellet groups on several dates in the lead up to an FSC survey and returning to the marked signs at the time of the survey to record whether or not each pellet group had survived. Several logistic models were fitted to the data, and estimations were based on a multi-model inference (MMI) approach according to information theory. The highest mean time to decay was estimated in coniferous forests, while mid-ranged values were found in maquis shrubs, and the lowest mean time to decay was observed in open areas. MMI by model averaging, based on Akaike weights, is recommended for making robust parameter estimations and for dealing with uncertainty in model selection.     

Association of MT1A haplotype with cardiovascular disease and antioxidant enzyme defense in elderly Greek population: comparison with an Italian cohort

Metallothioneins (MT), the antioxidant zinc-binding proteins, seem to mediate cardioprotection. It has been postulated that zinc homeostasis and MT function may be altered, as a consequence of oxidative stress, in cardiovascular disease (CVD), with a potential implication of MT genetic polymorphisms. The present study explores the role of +647A/C and +1245A/G MT1A polymorphisms on the susceptibility to CVD, zinc status and enzyme antioxidant activity, in the Greek and Italian populations. The country selection was based on the lower zinc status and the reduced zinc dietary intake in Greece than in Italy despite the similar Mediterranean dietary pattern. A total of 464 old, healthy control subjects and 369 old CVD patients more than 70 years of age were studied. Logistic regression model indicated that +1245 MT1A G+ genotype significantly increased the risk of CVD in Greece (34.4% vs. 23.2%; odds ratio=1.88, 95% confidence interval=1.14–3.08; P=.013) but not in Italy. Haplotype analysis showed an increment of CG haplotype frequency in CVD Greek patients (17.4% vs. 10.6%, P<.05). Differential country-related frequency distribution was also recorded. Applying a multivariate regression model, +647/+1245 MT1A haplotype was associated with a modulation of enzyme antioxidant activities in both countries. Decreased plasma zinc and reduced intracellular Zn release, as well as increased enzyme antioxidant activity, were more apparent in Greek healthy donors than in Italy. In conclusion, +1245 MT1A polymorphism and +647/+1245 MT1A haplotype are implicated in CVD in Greece but not in Italy, suggesting a role of gene–diet interaction in the disease predisposition.     

Opioids transiently prevent activation of apoptotic mechanisms following short periods of serum withdrawal

Opioids exert a proapoptotic effect on several normal and tumoral cells. The aim of the present article was to examine the effect of opioids on the PC12 rat pheochromocytoma cell line, a model for the study of chromaffin cell apoptosis. These cells produce delta- and kappa-opioid agonists and their receptors. Our results were as follows: The kappa- and delta2-opioid receptor agonists had a rapid but transient effect on apoptosis at 3 h, whereas mu opioids did not. The effect of opioids was reversible by the opioid antagonists naloxone and nor-binaltorphimine. The effect of opioids was protective, suppressing serum deprivation-induced apoptosis to approximately 50% of controls. The protective effect of opioids on PC12 apoptosis was measurable only under serum deprivation. The effect of opioids was remarkably reproducible and highly constant in timing, which did not appear to depend on the duration of the preceding serum deprivation. Finally, opioids prevented the elevation of the Bcl-2 and Bak proteins following serum deprivation to the levels attained by serum supplementation. Our combined data suggest that opioids protect PC12 cells from entering a state of induced apoptosis following serum deprivation.     

Crystal and microparticle effects on MDCK cell superoxide production: oxalate-specific mitochondrial membrane potential changes

We have previously shown that crystals of calcium oxalate (COM) elicit a superoxide (O₂⁻) response from mitochondria. We have now investigated: (i) if other microparticles can elicit the same response, (ii) if processing of crystals is involved, and (iii) at what level of mitochondrial function oxalate acts. O₂⁻ was measured in digitonin-permeabilized MDCK cells by lucigenin (10 μM) chemiluminescence. [¹⁴C]-COM dissociation was examined with or without EDTA and employing alternative chelators. Whereas mitochondrial O₂⁻ in COM-treated cells was three- to fourfold enhanced compared to controls, other particulates (uric acid, zymosan, and latex beads) either did not increase O₂⁻ or were much less effective (hydroxyapatite +50%, p < 0.01), with all at 28 μg/cm². Free oxalate (750 μM), at the level released from COM with EDTA (1 mM), increased O₂⁻ (+50%, p < 0.01). Omitting EDTA abrogated this signal, which was restored completely by EGTA and partially by ascorbate, but not by desferrioxamine or citrate. Omission of phosphate abrogated O₂⁻, implicating phosphate-dependent mitochondrial dicarboxylate transport. COM caused a time-related increase in the mitochondrial membrane potential (Δψ_m) measured using TMRM fluorescence and confocal microscopy. Application of COM to Fura 2-loaded cells induced rapid, large-amplitude cytosolic Ca²⁺ transients, which were inhibited by thapsigargin, indicating that COM induces release of Ca²⁺ from internal stores. Thus, COM-induced mitochondrial O₂⁻ requires the release of free oxalate and contributes to a synergistic response. Intracellular dissociation of COM and the mitochondrial dicarboxylate transporter are important in O₂⁻ production, which is probably regulated by Δψ_m.     

Carbonic Anhydrase Protects Fatty Liver Grafts against Ischemic Reperfusion Damage

Carbonic anhydrases (CAs) are ubiquitous metalloenzymes that catalyze the reversible hydration of carbon dioxide to bicarbonate and a proton. CAs are involved in numerous physiological and pathological processes, including acid-base homeostasis, electrolyte balance, oxygen delivery to tissues and nitric oxide generation. Given that these processes are found to be dysregulated during ischemia reperfusion injury (IRI), and taking into account the high vulnerability of steatotic livers to preservation injury, we hypothesized a new role for CA as a pharmacological agent able to protect against ischemic damage. Two different aspects of the role of CA II in fatty liver grafts preservation were evaluated: 1) the effect of its addition to Institut Georges Lopez (IGL-1) storage solution after cold ischemia; 2) and after 24h of cold storage followed by two hours of normothermic ex-vivo perfusion. In all cases, liver injury, CA II protein concentration, CA II mRNA levels and CA II activity were determined. In case of the ex-vivo perfusion, we further assessed liver function (bile production, bromosulfophthalein clearance) and Western blot analysis of phosphorylated adenosine monophosphate activated protein kinase (AMPK), mitogen activated protein kinases family (MAPKs) and endoplasmic reticulum stress (ERS) parameters (GRP78, PERK, IRE, eIF2α and ATF6). We found that CA II was downregulated after cold ischemia. The addition of bovine CA II to IGL-1 preservation solution efficiently protected steatotic liver against cold IRI. In the case of reperfusion, CA II protection was associated with better function, AMPK activation and the prevention of ERS and MAPKs activation. Interestingly, CA II supplementation was not associated with enhanced CO₂ hydration. The results suggest that CA II modulation may be a promising target for fatty liver graft preservation.     

The effect of addition of olive oil and "Aceto balsamico di Modena" wine vinegar in conjunction with active atmosphere packaging on the microbial and sensory quality of "Lollo Verde" lettuce and rocket salad

Fresh rocket “Eruca Sativa” and lettuce “Lollo Verde” leaves were stored with the addition of olive oil and wine vinegar “Aceto balsamico di Modena” under modified atmosphere packaging (MAP) (5% O₂/10% CO₂/85% N₂ for MAP A and 2% O₂/5% CO₂/93% N₂ for MAP B). The microbial (mesophilic, psychrotrophic bacteria and Enterobacteriacae), physical (color and firmness) and sensory parameters of samples were studied in relation to storage time (up to 10 days at 5 ± 1 °C). The effect of wine vinegar and the application of both MAP treatments reduced the growth of all bacteria populations (p < 0.05). Samples with olive oil stored under MAP A gave the best score for overall impression (3 and 2.1 for MAP A and B respectively at the 9th day of storage) while the addition of vinegar limited sensory shelf-life to 3 days (p < 0.05). Firmness was negatively affected by wine vinegar while samples with olive oil stored under MAP A maintained firmness close to normal. Color attributes were maintained better under both MAP treatments (p < 0.05).