The mitochondrial battlefield and membrane lipids during cell death signalling

This brief review focuses on current issues regarding the biochemical understanding of key reactions in the mitochondrial membrane damage induced by apoptosis signalling, especially in the pathway followed by death receptors in primary tissues. An overview of the emerging role of cardiolipin in apoptosis is also presented, discussing controversial issues and future directions of research. In addition, it is suggested that pro-apoptotic Bid may modulate key processes linking lipid constituents of mitochondrial membranes to lipid re-modelling that are altered early by death receptor signalling.     

Weekly oral alendronate in mevalonate kinase deficiency

Background

Mevalonate kinase deficiency (MKD) is caused by mutations in the MVK gene, encoding the second enzyme of mevalonate pathway, which results in subsequent shortage of downstream compounds, and starts in childhood with febrile attacks, skin, joint, and gastrointestinal symptoms, sometimes induced by vaccinations.

Methods

For a history of early-onset corticosteroid-induced reduction of bone mineral density in a 14-year-old boy with MKD, who also had presented three bone fractures, we administered weekly oral alendronate, a drug widely used in the management of osteoporosis and other high bone turnover diseases, which blocks mevalonate and halts the prenylation process.

Results

All of the patient’s MKD clinical and laboratory abnormalities were resolved after starting alendronate treatment.

Conclusions

This observation appears enigmatic, since alendronate should reinforce the metabolic block characterizing MKD, but is crucial because of the ultimate improvement shown by this patient. The anti-inflammatory properties of bisphosphonates are a new question for debate among physicians across various specialties, and requires further biochemical and clinical investigation.

     

Characterization of the rat alpha(1,3)galactosyltransferase: evidence for two independent genes encoding glycosyltransferases that synthesize Galalpha(1,3)Gal by two separate glycosylation pathways

The important xenoepitope Galalpha(1,3)Gal was thought to be exclusively synthesized by a single alpha(1,3)galactosyltransferase. However, the cloning of the distant family member rat iGb3 synthase, which is also capable of synthesizing Galalpha(1,3)Gal as the glycolipid structure iGb3, challenges the notion that alpha(1,3)galactosyltransferase is the sole Galalpha(1,3)Gal-synthesizing enzyme. We describe the cloning of the rat homolog of alpha(1,3)galactosyltransferase, showing that indeed the rat expresses two distinct alpha(1,3)galactosyltransferases, alpha(1,3)GT and iGb3 synthase. Rat alpha(1,3)galactosyltransferase shows a high amino acid sequence identity with the alpha(1,3)galactosyltransferase of mouse (90%), pig (76%), and ox (75%), in contrast to the low amino acid sequence identity (42%) with iGb3 synthase. The rat alpha(1,3)galactosyltransferase is expressed in heart, brain, spleen, kidney, and liver and has a similar intron/exon structure to the mouse alpha(1,3)galactosyltransferase. Transfection studies show that in contrast to the iGb3 synthase, rat alpha(1,3)galactosyltransferase can synthesize Galalpha(1,3)Gal on glycoproteins but cannot synthesize the glycolipid iGb3, defining two separate glycosylation pathways for the synthesis of Galalpha(1,3)Gal. Furthermore iGb3 synthase was found to be distinct from alpha(1,3)GT with its ability to synthesize poly-alpha-Gal glycolipid structures.     

Deregulation of transition metals homeostasis is a key feature of cadmium toxicity in Salmonella

Cadmium is a highly toxic metal whose presence in the environment represents a challenge for all forms of life. To improve our knowledge on cadmium toxicity, we have explored Salmonella Typhimurium responses to this metal. We have found that cadmium induces the concomitant expression of the cation efflux pump ZntA and of the high affinity zinc import system ZnuABC. This observation suggests that cadmium accumulation within the cell induces a condition of apparent zinc starvation, possibly due to the ability of this metal to compete with zinc for the metal binding site of proteins. This hypothesis is supported by the finding that strains lacking ZntA or ZnuABC are hyper-susceptible to cadmium and that the cadmium-induced growth defect of a znuABC mutant strain is largely relieved by zinc supplementation. A similar growth defect was observed for a mutant with impaired ability to acquire iron, whereas cadmium does not affect growth of a strain defective in manganese import. Cadmium also influences the expression and activity of the two cytoplasmic superoxide dismutases FeSOD and MnSOD, which are required to control cadmium-mediate oxidative stress. Exposure to cadmium causes a reduction of FeSOD activity in Salmonella wild type and the complete abrogation of its expression in the strain defective in iron import. In contrast, although MnSOD intracellular levels increase in response to cadmium, we observed discrepancies between protein levels and enzymatic activity which are suggestive of incorporation of non-catalytic metals in the active site or to cadmium-mediated inhibition of manganese import. Our results indicate that cadmium interferes with the ability of cells to manage transition metals and highlight the close interconnections between the homeostatic mechanisms regulating the intracellular levels of different metals.     

Recent changes in the distribution of carboxylesterase genes and associated chromosomal rearrangements in Greek populations of the tobacco aphid Myzus persicae nicotianae

We present data on the frequency of amplified E4 and FE4 carboxylesterase genes in Myzus persicae s.l. clones collected during the years 2002–2007 and 2012 in Greece. Most clones were of the tobacco aphid, Myzus persicae nicotianae. Samples from 2012 were genotyped with microsatellite DNA markers and a number of them were karyotyped. Aphid clones with amplified FE4 genes predominated in all years, whereas E4 was present in only 3.5% of all samples and always occurred in clones with FE4. Most of the clones examined showed high carboxylesterase activity levels (R2 resistant category). The results showed marked changes in the frequencies of the two carboxylesterase genes in the tobacco aphid populations compared to published data that were collected in Greece in the mid 1990s, when E4 was recorded on its own in 20% of all samples and in 32% of samples from tobacco. A parallel change in karyotype was also observed because the A1,3 translocation, which had a worldwide association with amplified E4 genes in the 1990s, was not detected in the clones analyzed in 2012. Possible causes for these changes are discussed, although selection as a result of pest management practices appears to be the major one. Novel chromosomal rearrangements were also found in M. persicae nicotianae clones. These rearrangements could be a result of clastogenic effects of nicotine, which could persist because of the holocentric nature of aphid chromosomes. The results are discussed in relation to rapid evolution events that have taken place in the tobacco aphid in Greece during the last two decades. © 2014 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 113 , 455–470.     

Pyridostigmine Restores Cardiac Autonomic Balance after Small Myocardial Infarction in Mice

The effect of pyridostigmine (PYR) - an acetylcholinesterase inhibitor - on hemodynamics and cardiac autonomic control, was never studied in conscious myocardial infarcted mice. Telemetry transmitters were implanted into the carotid artery under isoflurane anesthesia. Seven to ten days after recovery from the surgery, basal arterial pressure and heart rate were recorded, while parasympathetic and sympathetic tone (ΔHR) was evaluated by means of methyl atropine and propranolol. After the basal hemodynamic recording the mice were subjected to left coronary artery ligation for producing myocardial infarction (MI), or sham operation, and implantation of minipumps filled with PYR or saline. Separate groups of anesthetized (isoflurane) mice previously (4 weeks) subjected to MI, or sham coronary artery ligation, were submitted to cardiac function examination. The mice exhibited an infarct length of approximately 12%, no change in arterial pressure and increased heart rate only in the 1st week after MI. Vagal tone decreased in the 1^st week, while the sympathetic tone was increased in the 1^st and 4^th week after MI. PYR prevented the increase in heart rate but did not affect the arterial pressure. Moreover, PYR prevented the increase in sympathetic tone throughout the 4 weeks. Concerning the parasympathetic tone, PYR not only impaired its attenuation in the 1^st week, but enhanced it in the 4^th week. MI decreased ejection fraction and increased diastolic and systolic volume. Therefore, the pharmacological increase of peripheral acetylcholine availability by means of PYR prevented tachycardia, increased parasympathetic and decreased sympathetic tone after MI in mice.     

Proteomics in Parkinson's disease: An unbiased approach towards peripheral biomarkers and new therapies

Parkinson's disease is the most common neurodegenerative movement disorder, affecting about 6 million people worldwide with a slow progression of the symptoms. Its prevalence is expected to double in the most populated areas within the next two decades, according to increasing aged population. Consequently, Parkinson's disease is a socio-economic trouble and a major challenge for the public health system. Parkinson's disease treatment is merely symptomatic, as clinical symptoms appear when about 70% of the involved neurons are lost and potential disease-modifying/neuroprotective therapies would have no effect. In turn, the availability of an objective measure that allows early diagnosis would strongly impact on the costs that biotech- and pharma-companies will sustain in order to develop disease-modifying therapies. The establishment of suitable models to investigate the mechanisms of Parkinson's disease progression and, on the other hand, the discovery and validation of selective and specific molecular biomarkers for early and differential diagnosis are indeed two important goals for a better management of the disease. In this review, we focus on cellular and animal models of Parkinson's disease by describing their advantages and limitations as useful tools to identify pathogenetic pathways that deserve further exploitation. In parallel, we discuss how proteomics may provide a potent tool to observe altered pathways in models or altered biomarkers in patients with an unbiased, hypothesis-free approach.     

Human Chondrocytes Respond Discordantly to the Protein Encoded by the Osteoarthritis Susceptibility Gene GDF5

A genetic deficit mediated by SNP rs143383 that leads to reduced expression of GDF5 is strongly associated with large-joint osteoarthritis. We speculated that this deficit could be attenuated by the application of exogenous GDF5 protein and as a first step we have assessed what effect such application has on primary osteoarthritis chondrocyte gene expression. Chondrocytes harvested from cartilage of osteoarthritic patients who had undergone joint replacement were cultured with wildtype recombinant mouse and human GDF5 protein. We also studied variants of GDF5, one that has a higher affinity for the receptor BMPR-IA and one that is insensitive to the GDF5 antagonist noggin. As a positive control, chondrocytes were treated with TGF-β1. Chondrocytes were cultured in monolayer and micromass and the expression of genes coding for catabolic and anabolic proteins of cartilage were measured by quantitative PCR. The expression of the GDF5 receptor genes and the presence of their protein products was confirmed and the ability of GDF5 signal to translocate to the nucleus was demonstrated by the activation of a luciferase reporter construct. The capacity of GDF5 to elicit an intracellular signal in chondrocytes was demonstrated by the phosphorylation of intracellular Smads. Chondrocytes cultured with TGF-β1 demonstrated a consistent down regulation of MMP1, MMP13 and a consistent upregulation of TIMP1 and COL2A1 with both culture techniques. In contrast, chondrocytes cultured with wildtype GDF5, or its variants, did not show any consistent response, irrespective of the culture technique used. Our results show that osteoarthritis chondrocytes do not respond in a predictable manner to culture with exogenous GDF5. This may be a cause or a consequence of the osteoarthritis disease process and will need to be surmounted if treatment with exogenous GDF5 is to be advanced as a potential means to overcome the genetic deficit conferring osteoarthritis susceptibility at this gene.     

Sex allocation in yellow-legged gulls (Larus michahellis) depends on nutritional constraints on production of large last eggs

Male and female offspring can differ in their susceptibility to pre-natal (e.g. egg quality) and post-natal (e.g. sib–sib competition) conditions, and parents can therefore increase their individual fitness by adjusting these maternal effects according to offspring sex. In birds, egg mass and laying/hatching order are the main determinants of offspring viability, but these effects can act differently on each sex. In a previous study, relatively large last-laid (c-)eggs of yellow-legged gulls (Larus michahellis) were more likely to carry a female embryo. This suggests compensatory allocation of maternal resources to daughters from c-eggs, which suffer reduced viability. In the present study, we supplemented yellow-legged gulls with food during the laying period to experimentally test whether their nutritional conditions were responsible for the observed covariation between c-egg sex and mass. As predicted, food supplementation enhanced female c-eggs'' mass more than that of male c-eggs. Thus, this experiment indicates that mothers strategically allocated their resources to c-eggs, possibly in order to compensate for the larger susceptibility of daughters to hatching (and laying) order. The results also suggested that mothers decided on resource allocation depending on the sex of already ovulated c-eggs, rather than ovulating ova of either sex depending on food availability.     

Contribution of global rare copy-number variants to the risk of sporadic congenital heart disease

Previous studies have shown that copy-number variants (CNVs) contribute to the risk of complex developmental phenotypes. However, the contribution of global CNV burden to the risk of sporadic congenital heart disease (CHD) remains incompletely defined. We generated genome-wide CNV data by using Illumina 660W-Quad SNP arrays in 2,256 individuals with CHD, 283 trio CHD-affected families, and 1,538 controls. We found association of rare genic deletions with CHD risk (odds ratio [OR] = 1.8, p = 0.0008). Rare deletions in study participants with CHD had higher gene content (p = 0.001) with higher haploinsufficiency scores (p = 0.03) than they did in controls, and they were enriched with Wnt-signaling genes (p = 1 × 10⁻⁵). Recurrent 15q11.2 deletions were associated with CHD risk (OR = 8.2, p = 0.02). Rare de novo CNVs were observed in ∼5% of CHD trios; 10 out of 11 occurred on the paternally transmitted chromosome (p = 0.01). Some of the rare de novo CNVs spanned genes known to be involved in heart development (e.g., HAND2 and GJA5). Rare genic deletions contribute ∼4% of the population-attributable risk of sporadic CHD. Second to previously described CNVs at 1q21.1, deletions at 15q11.2 and those implicating Wnt signaling are the most significant contributors to the risk of sporadic CHD. Rare de novo CNVs identified in CHD trios exhibit paternal origin bias.