Continent-wide declines in shallow reef life over a decade of ocean warming.

Human society is dependent on nature1,2, but whether our ecological foundations are at risk remains unknown in the absence of systematic monitoring of species' populations3. Knowledge of species fluctuations is particularly inadequate in the marine realm4. Here we assess the population trends of 1,057 common shallow reef species from multiple phyla at 1,636 sites around Australia over the past decade. Most populations decreased over this period, including many tropical fishes, temperate invertebrates (particularly echinoderms) and southwestern Australian macroalgae, whereas coral populations remained relatively stable. Population declines typically followed heatwave years, when local water temperatures were more than 0.5 °C above temperatures in 2008. Following heatwaves5,6, species abundances generally tended to decline near warm range edges, and increase near cool range edges. More than 30% of shallow invertebrate species in cool latitudes exhibited high extinction risk, with rapidly declining populations trapped by deep ocean barriers, preventing poleward retreat as temperatures rise. Greater conservation effort is needed to safeguard temperate marine ecosystems, which are disproportionately threatened and include species with deep evolutionary roots. Fundamental among such efforts, and broader societal needs to efficiently adapt to interacting anthropogenic and natural pressures, is greatly expanded monitoring of species' population trends7,8.

Global conservation outcomes depend on marine protected areas with five key features.

In line with global targets agreed under the Convention on Biological Diversity, the number of marine protected areas (MPAs) is increasing rapidly, yet socio-economic benefits generated by MPAs remain difficult to predict and under debate. MPAs often fail to reach their full potential as a consequence of factors such as illegal harvesting, regulations that legally allow detrimental harvesting, or emigration of animals outside boundaries because of continuous habitat or inadequate size of reserve. Here we show that the conservation benefits of 87 MPAs investigated worldwide increase exponentially with the accumulation of five key features: no take, well enforced, old (>10 years), large (>100 km(2)), and isolated by deep water or sand. Using effective MPAs with four or five key features as an unfished standard, comparisons of underwater survey data from effective MPAs with predictions based on survey data from fished coasts indicate that total fish biomass has declined about two-thirds from historical baselines as a result of fishing. Effective MPAs also had twice as many large (>250 mm total length) fish species per transect, five times more large fish biomass, and fourteen times more shark biomass than fished areas. Most (59%) of the MPAs studied had only one or two key features and were not ecologically distinguishable from fished sites. Our results show that global conservation targets based on area alone will not optimize protection of marine biodiversity. More emphasis is needed on better MPA design, durable management and compliance to ensure that MPAs achieve their desired conservation value.

Integrating abundance and functional traits reveals new global hotspots of fish diversity.

Species richness has dominated our view of global biodiversity patterns for centuries. The dominance of this paradigm is reflected in the focus by ecologists and conservation managers on richness and associated occurrence-based measures for understanding drivers of broad-scale diversity patterns and as a biological basis for management. However, this is changing rapidly, as it is now recognized that not only the number of species but the species present, their phenotypes and the number of individuals of each species are critical in determining the nature and strength of the relationships between species diversity and a range of ecological functions (such as biomass production and nutrient cycling). Integrating these measures should provide a more relevant representation of global biodiversity patterns in terms of ecological functions than that provided by simple species counts. Here we provide comparisons of a traditional global biodiversity distribution measure based on richness with metrics that incorporate species abundances and functional traits. We use data from standardized quantitative surveys of 2,473 marine reef fish species at 1,844 sites, spanning 133 degrees of latitude from all ocean basins, to identify new diversity hotspots in some temperate regions and the tropical eastern Pacific Ocean. These relate to high diversity of functional traits amongst individuals in the community (calculated using Rao's Q), and differ from previously reported patterns in functional diversity and richness for terrestrial animals, which emphasize species-rich tropical regions only. There is a global trend for greater evenness in the number of individuals of each species, across the reef fish species observed at sites ('community evenness'), at higher latitudes. This contributes to the distribution of functional diversity hotspots and contrasts with well-known latitudinal gradients in richness. Our findings suggest that the contribution of species diversity to a range of ecosystem functions varies over large scales, and imply that in tropical regions, which have higher numbers of species, each species contributes proportionally less to community-level ecological processes on average than species in temperate regions. Metrics of ecological function usefully complement metrics of species diversity in conservation management, including when identifying planning priorities and when tracking changes to biodiversity values.

Fishing-gear restrictions and biomass gains for coral reef fishes in marine protected areas.

Considerable empirical evidence supports recovery of reef fish populations with fishery closures. In countries where full exclusion of people from fishing may be perceived as inequitable, fishing-gear restrictions on nonselective and destructive gears may offer socially relevant management alternatives to build recovery of fish biomass. Even so, few researchers have statistically compared the responses of tropical reef fisheries to alternative management strategies. We tested for the effects of fishery closures and fishing gear restrictions on tropical reef fish biomass at the community and family level. We conducted 1,396 underwater surveys at 617 unique sites across a spatial hierarchy within 22 global marine ecoregions that represented 5 realms. We compared total biomass across local fish assemblages and among 20 families of reef fishes inside marine protected areas (MPAs) with different fishing restrictions: no-take, hook-and-line fishing only, several fishing gears allowed, and sites open to all fishing gears. We included a further category representing remote sites, where fishing pressure is low. As expected, full fishery closures, (i.e., no-take zones) most benefited community- and family-level fish biomass in comparison with restrictions on fishing gears and openly fished sites. Although biomass responses to fishery closures were highly variable across families, some fishery targets (e.g., Carcharhinidae and Lutjanidae) responded positively to multiple restrictions on fishing gears (i.e., where gears other than hook and line were not permitted). Remoteness also positively affected the response of community-level fish biomass and many fish families. Our findings provide strong support for the role of fishing restrictions in building recovery of fish biomass and indicate important interactions among fishing-gear types that affect biomass of a diverse set of reef fish families.

Conservation Genetics of the Scalloped Hammerhead Shark in the Pacific Coast of Colombia.

Previous investigations of the population genetics of the scalloped hammerhead sharks (Sphyrna lewini) in the Eastern Tropical Pacific have lacked information about nursery areas. Such areas are key to promoting conservation initiatives that can protect young sharks from threats such as overfishing. Here, we investigated the genetic diversity, phylogeography, and connectivity of S. lewini found in 3 areas of Colombia's Pacific coast: around Malpelo Island and in 2 National Natural Parks on the Colombian Pacific mainland (Sanquianga and Ensenada de Utría). We analyzed mtDNA control region (CR) sequences and genotyped 15 microsatellite loci in 137 samples of adults and juveniles. The mtDNA analyses showed haplotypes shared between the Colombian Pacific individuals sampled in this investigation and other areas in the Eastern Tropical Pacific, the Indo-Pacific, and with sequences previously reported in Colombia (Buenaventura Port), as well as 4 unique haplotypes. Population assignment and paternity analyses detected 3 parent-offspring pairs between Malpelo and Sanquianga and 1 between Malpelo and Utría. These results indicate high genetic connectivity between Malpelo Island and the Colombian Pacific coast, suggesting that these 2 areas are nurseries for S. lewini. This is, to our knowledge, the first evidence of nursery areas identified for the scalloped hammerhead shark anywhere in the world. Additional conservation planning may be required to protect these nursery habitats of this endangered shark species.

Global human footprint on the linkage between biodiversity and ecosystem functioning in reef fishes.

Difficulties in scaling up theoretical and experimental results have raised controversy over the consequences of biodiversity loss for the functioning of natural ecosystems. Using a global survey of reef fish assemblages, we show that in contrast to previous theoretical and experimental studies, ecosystem functioning (as measured by standing biomass) scales in a non-saturating manner with biodiversity (as measured by species and functional richness) in this ecosystem. Our field study also shows a significant and negative interaction between human population density and biodiversity on ecosystem functioning (i.e., for the same human density there were larger reductions in standing biomass at more diverse reefs). Human effects were found to be related to fishing, coastal development, and land use stressors, and currently affect over 75% of the world's coral reefs. Our results indicate that the consequences of biodiversity loss in coral reefs have been considerably underestimated based on existing knowledge and that reef fish assemblages, particularly the most diverse, are greatly vulnerable to the expansion and intensity of anthropogenic stressors in coastal areas.

WNT Activation and TGFß-Smad Inhibition Potentiate Stemness of Mammalian Auditory Neuroprogenitors for High-Throughput Generation of Functional Auditory Neurons In Vitro.

Hearing loss affects over 460 million people worldwide and is a major socioeconomic burden. Both genetic and environmental factors (i.e., noise overexposure, ototoxic drug treatment and ageing), promote the irreversible degeneration of cochlear hair cells and associated auditory neurons, leading to sensorineural hearing loss. In contrast to birds, fish and amphibians, the mammalian inner ear is virtually unable to regenerate due to the limited stemness of auditory progenitors, and no causal treatment is able to prevent or reverse hearing loss. As of today, a main limitation for the development of otoprotective or otoregenerative therapies is the lack of efficient preclinical models compatible with high-throughput screening of drug candidates. Currently, the research field mainly relies on primary organotypic inner ear cultures, resulting in high variability, low throughput, high associated costs and ethical concerns. We previously identified and characterized the phoenix auditory neuroprogenitors (ANPGs) as highly proliferative progenitor cells isolated from the A/J mouse cochlea. In the present study, we aim at identifying the signaling pathways responsible for the intrinsic high stemness of phoenix ANPGs. A transcriptomic comparison of traditionally low-stemness ANPGs, isolated from C57Bl/6 and A/J mice at early passages, and high-stemness phoenix ANPGs was performed, allowing the identification of several differentially expressed pathways. Based on differentially regulated pathways, we developed a reprogramming protocol to induce high stemness in presenescent ANPGs (i.e., from C57Bl6 mouse). The pharmacological combination of the WNT agonist (CHIR99021) and TGFß/Smad inhibitors (LDN193189 and SB431542) resulted in a dramatic increase in presenescent neurosphere growth, and the possibility to expand ANPGs is virtually limitless. As with the phoenix ANPGs, stemness-induced ANPGs could be frozen and thawed, enabling distribution to other laboratories. Importantly, even after 20 passages, stemness-induced ANPGs retained their ability to differentiate into electrophysiologically mature type I auditory neurons. Both stemness-induced and phoenix ANPGs resolve a main bottleneck in the field, allowing efficient, high-throughput, low-cost and 3R-compatible in vitro screening of otoprotective and otoregenerative drug candidates. This study may also add new perspectives to the field of inner ear regeneration.

Development and in vivo validation of small interfering RNAs targeting NOX3 to prevent sensorineural hearing loss.

The reactive oxygen species (ROS)-generating enzyme NOX3 has recently been implicated in the pathophysiology of several acquired forms of sensorineural hearing loss, including cisplatin-, noise- and age-related hearing loss. NOX3 is highly and specifically expressed in the inner ear and therefore represents an attractive target for specific intervention aiming at otoprotection. Despite the strong rationale to inhibit NOX3, there is currently no specific pharmacological inhibitor available. Molecular therapy may represent a powerful alternative. In this study, we developed and tested a collection of small interfering (si) RNA constructs to establish a proof of concept of NOX3 inhibition through local delivery in the mouse inner ear. The inhibitory potential of 10 different siRNA constructs was first assessed in three different cells lines expressing the NOX3 complex. Efficacy of the most promising siRNA construct to knock-down NOX3 was then further assessed in vivo, comparing middle ear delivery and direct intracochlear delivery through the posterior semi-circular canal. While hearing was completely preserved through the intervention, a significant downregulation of NOX3 expression in the mouse inner ear and particularly in the spiral ganglion area at clinically relevant levels (>60%) was observed 48 h after treatment. In contrast to successful intracochlear delivery, middle ear administration of siRNA failed to significantly inhibit Nox3 mRNA expression. In conclusion, intracochlear delivery of NOX3-siRNAs induces a robust temporal NOX3 downregulation, which could be of relevance to prevent predictable acute insults such as cisplatin chemotherapy-mediated ototoxicity and other forms of acquired hearing loss, including post-prevention of noise-induced hearing loss immediately after trauma. Successful translation of our concept into an eventual clinical use in humans will depend on the development of atraumatic and efficient delivery routes into the cochlea without a risk to induce hearing loss through the intervention.

NADPH Oxidase 3 Deficiency Protects From Noise-Induced Sensorineural Hearing Loss.

The reactive oxygen species (ROS)-generating NADPH oxidase NOX3 isoform is highly and specifically expressed in the inner ear. NOX3 is needed for normal vestibular development but NOX-derived ROS have also been implicated in the pathophysiology of sensorineural hearing loss. The role of NOX-derived ROS in noise-induced hearing loss, however, remains unclear and was addressed with the present study. Two different mouse strains, deficient in NOX3 or its critical subunit p22phox, were subjected to a single noise exposure of 2 h using an 8-16 kHz band noise at an intensity of 116-120 decibel sound pressure level. In the hours following noise exposure, there was a significant increase in cochlear mRNA expression of NOX3 in wild type animals. By using RNAscope in situ hybridization, NOX3 expression was primarily found in the Rosenthal canal area, colocalizing with auditory neurons. One day after the noise trauma, we observed a high frequency hearing loss in both knock-out mice, as well as their wild type littermates. At day seven after noise trauma however, NOX3 and p22phox knockout mice showed a significantly improved hearing recovery and a marked preservation of neurosensory cochlear structures compared to their wild type littermates. Based on these findings, an active role of NOX3 in the pathophysiology of noise-induced hearing loss can be demonstrated, in line with recent evidence obtained in other forms of acquired hearing loss. The present data demonstrates that the absence of functional NOX3 enhances the hearing recovery phase following noise trauma. This opens an interesting clinical window for pharmacological or molecular intervention aiming at post prevention of noise-induced hearing loss.

Tracking widespread climate-driven change on temperate and tropical reefs.

Warming seas, marine heatwaves, and habitat degradation are increasingly widespread phenomena affecting marine biodiversity, yet our understanding of their broader impacts is largely derived from collective insights from independent localized studies. Insufficient systematic broadscale monitoring limits our understanding of the true extent of these impacts and our capacity to track these at scales relevant to national policies and international agreements. Using an extensive time series of co-located reef fish community structure and habitat data spanning 12 years and the entire Australian continent, we found that reef fish community responses to changing temperatures and habitats are dynamic and widespread but regionally patchy. Shifts in composition and abundance of the fish community often occurred within 2 years of environmental or habitat change, although the relative importance of these two mechanisms of climate impact tended to differ between tropical and temperate zones. The clearest of these changes on temperate and subtropical reefs were temperature related, with responses measured by the reef fish thermal index indicating reshuffling according to the thermal affinities of species present. On low latitude coral reefs, the community generalization index indicated shifting dominance of habitat generalist fishes through time, concurrent with changing coral cover. Our results emphasize the importance of maintaining local ecological detail when scaling up datasets to inform national policies and global biodiversity targets. Scaled-up ecological monitoring is needed to discriminate among increasingly diverse drivers of large-scale biodiversity change and better connect presently disjointed systems of biodiversity observation, indicator research, and governance.

Local Cisplatin Delivery in Mouse Reliably Models Sensorineural Ototoxicity Without Systemic Adverse Effects.

Cisplatin is a lifesaving chemotherapeutic drug with marked ototoxic adverse effects. Cisplatin-induced hearing loss affects a significant part of cancer-surviving patients and is an unmet clinical need with important socioeconomic consequences. Unfortunately, in current preclinical animal models of cisplatin ototoxicity, which are mainly based on systemic delivery, important morbidity is observed, leading to premature death. This methodology not only raises obvious animal welfare concerns but also increases the number of animals used in ototoxicity studies to compensate for dropouts related to early death. To overcome these important limitations, we developed a local delivery model based on the application of a cisplatin solution directly into the otic bulla through a retroauricular approach. The local delivery model reliably induced significant hearing loss with a mean threshold shift ranging from 10 to 30 dB, strongly affecting the high frequencies (22 and 32 kHz). Importantly, mice did not show visible stress or distress indicators and no significant morbidity in comparison with a traditional systemic delivery control group of mice injected intraperitoneally with 10 mg/kg cisplatin, where significant weight loss >10% in all treated animals (without any recovery) led to premature abortion of experiments on day 3. Mass spectrometry confirmed the absence of relevant systemic uptake after local delivery, with platinum accumulation restricted to the cochlea, whereas important platinum concentrations were detected in the liver and kidney of the systemic cisplatin group. A clear correlation between the cochlear platinum concentration and the auditory threshold shift was observed. Immunohistochemistry revealed statistically significant loss of outer hair cells in the basal and apical turns of the cochlea and an important and statistically significant loss of auditory neurons and synapses in all cochlear regions. In conclusion, local cisplatin delivery induces robust hearing loss with minimal morbidity, thereby offering a reliable rodent model for human cisplatin ototoxicity, reducing the number of animals required and showing improved animal welfare compared with traditional systemic models.

Redox activation of excitatory pathways in auditory neurons as mechanism of age-related hearing loss.

Age-related hearing (ARHL) loss affects a large part of the human population with a major impact on our aging societies. Yet, underlying mechanisms are not understood, and no validated therapy or prevention exists. NADPH oxidases (NOX), are important sources of reactive oxygen species (ROS) in the cochlea and might therefore be involved in the pathogenesis of ARHL. Here we investigate ARHL in a mouse model. Wild type mice showed early loss of hearing and cochlear integrity, while animals deficient in the NOX subunit p22phox remained unaffected up to six months. Genes of the excitatory pathway were down-regulated in p22phox-deficient auditory neurons. Our results demonstrate that NOX activity leads to upregulation of genes of the excitatory pathway, to excitotoxic cochlear damage, and ultimately to ARHL. In the absence of functional NOXs, aging mice conserve hearing and cochlear morphology. Our study offers new insights into pathomechanisms and future therapeutic targets of ARHL.

Intrinsically Self-renewing Neuroprogenitors From the A/J Mouse Spiral Ganglion as Virtually Unlimited Source of Mature Auditory Neurons.

Nearly 460 million individuals are affected by sensorineural hearing loss (SNHL), one of the most common human sensory disorders. In mammals, hearing loss is permanent due to the lack of efficient regenerative capacity of the sensory epithelia and spiral ganglion neurons (SGN). Sphere-forming progenitor cells can be isolated from the mammalian inner ear and give rise to inner ear specific cell types in vitro. However, the self-renewing capacities of auditory progenitor cells from the sensory and neuronal compartment are limited to few passages, even after adding powerful growth factor cocktails. Here, we provide phenotypical and functional characterization of a new pool of auditory progenitors as sustainable source for sphere-derived auditory neurons. The so-called phoenix auditory neuroprogenitors, isolated from the A/J mouse spiral ganglion, exhibit robust intrinsic self-renewal properties beyond 40 passages. At any passage or freezing-thawing cycle, phoenix spheres can be efficiently differentiated into mature spiral ganglion cells by withdrawing growth factors. The differentiated cells express both neuronal and glial cell phenotypic markers and exhibit similar functional properties as mouse spiral ganglion primary explants and human sphere-derived spiral ganglion cells. In contrast to other rodent models aiming at sustained production of auditory neurons, no genetic transformation of the progenitors is needed. Phoenix spheres therefore represent an interesting starting point to further investigate self-renewal in the mammalian inner ear, which is still far from any clinical application. In the meantime, phoenix spheres already offer an unlimited source of mammalian auditory neurons for high-throughput screens while substantially reducing the numbers of animals needed.

Silencing DJ-1 reveals its contribution in paraquat-induced autophagy.

The role of autophagy as a survival strategy of cells constitutes an emerging topic in the study of the pathogenesis of several diseases with autophagic changes being described in a number of age-related neurodegenerative disorders, including Parkinson's disease (PD). Although the etiology of PD is still unknown, both environmental (for example, paraquat exposure) and genetic factors have been investigated as putative causes of the disease. In the latter case, mutations or changes in the protein DJ-1 have been reported to be associated with autosomal recessive, early-onset parkinsonism. In this paper we established a model system to study the involvement of the DJ-1 protein in paraquat-induced autophagy. When human neuroblastoma SH-SY5Y cells were transfected with DJ-1-specific small interfering RNAs and exposed to paraquat, we observed (i) sensitization additive with paraquat-induced apoptotic cell death, (ii) inhibition of the cytoplasmic accumulation of autophagic vacuoles as well as the recruitment of LC3 fusion protein to the vacuoles, (iii) exacerbation of apoptotic cell death in the presence of the autophagy inhibitor 3-methyladenine, and (iv) an increase in mammalian target of rapamycin phosphorylation. Taken together, these findings suggest an active role for DJ-1 in the autophagic response produced by paraquat, providing evidence for the role of PD-related proteins in the autophagic degradation pathway, a factor that should be considered in the design of potential therapies for the treatment of the disease.

Hearing regeneration and regenerative medicine: present and future approaches.

More than 5% of the world population lives with a hearing impairment. The main factors responsible for hearing degeneration are ototoxic drugs, aging, continued exposure to excessive noise and infections. The pool of adult stem cells in the inner ear drops dramatically after birth, and therefore an endogenous cellular source for regeneration is absent. Hearing loss can emerge after the degeneration of different cochlear components, so there are multiple targets to be reached, such as hair cells (HCs), spiral ganglion neurons (SGNs), supporting cells (SCs) and ribbon synapses. Important discoveries in the hearing regeneration field have been reported regarding stem cell transplantation, migration and survival; genetic systems for cell fate monitoring; and stem cell differentiation to HCs, SGNs and SCs using adult stem cells, embryonic stem cells and induced pluripotent stem cells. Moreover, some molecular mediators that affect the establishment of functional synapses have been identified. In this review, we will focus on reporting the state of the art in the regenerative medicine field for hearing recovery. Stem cell research has enabled remarkable advances in regeneration, particularly in neuronal cells and synapses. Despite the progress achieved, there are certain issues that need a deeper development to improve the results already obtained, or to develop new approaches aiming for the clinical application.

Movements of scalloped hammerhead sharks (Sphyrna lewini) at Cocos Island, Costa Rica and between oceanic islands in the Eastern Tropical Pacific.

Many species of sharks form aggregations around oceanic islands, yet their levels of residency and their site specificity around these islands may vary. In some cases, the waters around oceanic islands have been designated as marine protected areas, yet the conservation value for threatened shark species will depend greatly on how much time they spend within these protected waters. Eighty-four scalloped hammerhead sharks (Sphyrna lewini Griffith & Smith), were tagged with acoustic transmitters at Cocos Island between 2005-2013. The average residence index, expressed as a proportion of days present in our receiver array at the island over the entire monitoring period, was 0.52±0.31, implying that overall the sharks are strongly associated with the island. Residency was significantly greater at Alcyone, a shallow seamount located 3.6 km offshore from the main island, than at the other sites. Timing of presence at the receiver locations was mostly during daytime hours. Although only a single individual from Cocos was detected on a region-wide array, nine hammerheads tagged at Galapagos and Malpelo travelled to Cocos. The hammerheads tagged at Cocos were more resident than those visiting from elsewhere, suggesting that the Galapagos and Malpelo populations may use Cocos as a navigational waypoint or stopover during seasonal migrations to the coastal Central and South America. Our study demonstrates the importance of oceanic islands for this species, and shows that they may form a network of hotspots in the Eastern Tropical Pacific.

Identification of genes associated with paraquat-induced toxicity in SH-SY5Y cells by PCR array focused on apoptotic pathways.

Paraquat (PQ) (1,1-dimethyl-4,4'-bipyridinium dichloride), a widely used herbicide, has been suggested as a potential etiologic factor for the development of Parkinson's disease (PD). In this sense, understanding of the molecular mechanism underlying PQ-induced toxicity to neural cells is important for optimal use as well as for the development of new drugs. To gain insights into PQ-induced neurotoxicity, polymerase chain reaction (PCR) array analysis focused on a panel of apoptosis-related genes was performed using neuroblastoma SH-SY5Y cells. Up to 65 apoptosis-related genes were monitored. Our analysis of apoptotic process through microarray technology showed that in PQ-induced neuroblastoma SH-SY5Y cells, there is a different expression of BIK, CASP3, CASP7, CRADD, DAPK, FAS, and other related genes, in comparison to unstimulated cells. Evaluation of genes regulated differentially is essential for the development of therapeutic approaches in multifactorial diseases as PD. Our data provide a useful basis for screening candidate targets for early diagnosis and further intervention in PQ-mediated toxicity of neural cells.

Inhibition of paraquat-induced autophagy accelerates the apoptotic cell death in neuroblastoma SH-SY5Y cells.

Autophagy is a degradative mechanism involved in the recycling and turnover of cytoplasmic constituents from eukaryotic cells. This phenomenon of autophagy has been observed in neurons from patients with Parkinson's disease (PD), suggesting a functional role for autophagy in neuronal cell death. On the other hand, it has been demonstrated that exposure to pesticides can be a risk factor in the incidence of PD. In this sense, paraquat (PQ) (1,1'-dimethyl-4,4'-bipyridinium dichloride), a widely used herbicide that is structurally similar to the known dopaminergic neurotoxicant MPP(+) (1-methyl-4-phenyl-pyridine), has been suggested as a potential etiologic factor for the development of PD. The current study shows, for the first time, that low concentrations of PQ induce several characteristics of autophagy in human neuroblastoma SH-SY5Y cells. In this way, PQ induced the accumulation of autophagic vacuoles (AVs) in the cytoplasm and the recruitment of a LC3-GFP fusion protein to AVs. Furthermore, the cells treated with PQ showed an increase of the long-lived protein degradation which is blocked in the presence of the autophagy inhibitor 3-methyladenine and regulated by the mammalian target of rapamycin (mTOR) signaling. Finally, the cells succumbed to cell death with hallmarks of apoptosis such as phosphatidylserine exposure, caspase activation, and chromatin condensation. While caspase inhibition retarded cell death, autophagy inhibition accelerated the apoptotic cell death induced by PQ. Altogether, these findings show the relationship between autophagy and apoptotic cell death in human neuroblastoma cells treated with PQ.

Arginase and polyamine synthesis are key factors in the regulation of experimental leishmaniasis in vivo.

Arginase 1, an enzyme induced by Th2 cytokines, is a hallmark of alternatively activated macrophages and is responsible for the hydrolysis of L-arginine into ornithine, the building block for the production of polyamines. Upregulation of arginase 1 has been observed in a variety of diseases, but the mechanisms by which arginase contributes to pathology are not well understood. We reveal here a unique role for arginase 1 in the pathogenesis of nonhealing leishmaniasis, a prototype Th2 disease, and demonstrate that the activity of this enzyme promotes pathology and uncontrolled growth of Leishmania parasites in vivo. Inhibition of arginase activity during the course of infection has a clear therapeutic effect, as evidenced by markedly reduced pathology and efficient control of parasite replication. Despite the clear amelioration of the disease, this treatment does not alter the Th2 response. To address the underlying mechanisms, the arginase-induced L-arginine catabolism was investigated and the results demonstrate that arginase regulates parasite growth directly by affecting the polyamine synthesis in macrophages.

Low concentrations of paraquat induces early activation of extracellular signal-regulated kinase 1/2, protein kinase B, and c-Jun N-terminal kinase 1/2 pathways: role of c-Jun N-terminal kinase in paraquat-induced cell death.

Paraquat is a herbicide with a potential risk to induce parkinsonism due to its demonstrated neurotoxicity and its strong structural similarity to 1-methyl-4-phenylpyridinium (MPP(+)), a well-known neurotoxin which causes a clinical syndrome similar to Parkinson's disease (PD). However, at present very little is known about the signaling pathways activated by paraquat in any cell system. In this study, we have investigated the effect of paraquat on extracellular signal-regulated kinases 1 and 2 (ERK1/2), c-Jun N-terminal kinase (JNK), and protein kinase B (PKB) activation in E18 cells. Low concentrations of paraquat stimulated very early increases in ERK1/2, JNK1/2, and PKB phosphorylation. The phosphatidylinositol 3-kinase (PI-3K) inhibitors wortmannin and LY 294002 (2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one) inhibited early paraquat-induced increases in PKB phosphorylation. Furthermore, early paraquat-mediated increases in ERK1/2 activation were sensitive to the mitogen-activated protein kinase kinase 1 (MEK1) inhibitor PD 98059 (2'-amino-3'-methoxyflavone), whereas JNK1/2 responses were blocked by the JNK1/2 inhibitor SP 600125 (anthra[1-9-cd]pyrazol-6(2H)-one). Pretreatment with wortmannin, LY 294002, or PD 98059 had no effect on paraquat cell death in E18 cells. In contrast, SP 600125 significantly decreased paraquat-induced cell death in E18 cells. In conclusion, we have shown that low concentrations of paraquat stimulate robust very early increases in ERK1/2, JNK1/2, and PKB phosphorylation in E18 cells. Furthermore, the data presented clearly suggest that inhibition of the JNK1/2 pathway protects E18 cells from paraquat-induced cell death and support the fact that inhibition of early activation of JNK1/2 can constitute a potential strategy in PD treatment.