The autophagy protein Def8 is altered in Alzheimer's disease and Aß42-expressing Drosophila brains.

Alzheimer's disease (AD) is the most common neurodegenerative disorder, characterized by protein accumulation in the brain as a main neuropathological hallmark. Among them, Aß42 peptides tend to aggregate and create oligomers and plaques. Macroautophagy, a form of autophagy characterized by a double-membrane vesicle, plays a crucial role in maintaining neuronal homeostasis by degrading protein aggregates and dysfunctional organelles as a quality control process. Recently, DEF8, a relatively uncharacterized protein, has been proposed as a participant in vesicular traffic and autophagy pathways. We have reported increased DEF8 levels in lymphocytes from mild cognitive impairment (MCI) and early-stage AD patients and a neuronal profile in a murine transgenic AD model. Here, we analyzed DEF8 localization and levels in the postmortem frontal cortex of AD patients, finding increased levels compared to healthy controls. To evaluate the potential function of DEF8 in the nervous system, we performed an in silico assessment of its expression and network profiles, followed by an in vivo evaluation of a neuronal Def8 deficient model using a Drosophila melanogaster model of AD based on Aß42 expression. Our findings show that DEF8 is an essential protein for maintaining cellular homeostasis in the nervous system, and it is upregulated under stress conditions generated by Aß42 aggregation. This study suggests DEF8 as a novel actor in the physiopathology of AD, and its exploration may lead to new treatment avenues.

Inner mitochondrial membrane structure and fusion dynamics are altered in senescent human iPSC-derived and primary rat cardiomyocytes.

Dysfunction of the aging heart is a major cause of death in the human population. Amongst other tasks, mitochondria are pivotal to supply the working heart with ATP. The mitochondrial inner membrane (IMM) ultrastructure is tailored to meet these demands and to provide nano-compartments for specific tasks. Thus, function and morphology are closely coupled. Senescent cardiomyocytes from the mouse heart display alterations of the inner mitochondrial membrane. To study the relation between inner mitochondrial membrane architecture, dynamics and function is hardly possible in living organisms. Here, we present two cardiomyocyte senescence cell models that allow in cellular studies of mitochondrial performance. We show that doxorubicin treatment transforms human iPSC-derived cardiomyocytes and rat neonatal cardiomyocytes in an aged phenotype. The treated cardiomyocytes display double-strand breaks in the nDNA, have ß-galactosidase activity, possess enlarged nuclei, and show p21 upregulation. Most importantly, they also display a compromised inner mitochondrial structure. This prompted us to test whether the dynamics of the inner membrane was also altered. We found that the exchange of IMM components after organelle fusion was faster in doxorubicin-treated cells than in control cells, with no change in mitochondrial fusion dynamics at the meso-scale. Such altered IMM morphology and dynamics may have important implications for local OXPHOS protein organization, exchange of damaged components, and eventually the mitochondrial bioenergetics function of the aged cardiomyocyte.

Neuronal Rubicon Represses Extracellular APP/Amyloid ß Deposition in Alzheimer's Disease.

Alzheimer's disease (AD) is the most prevalent age-associated neurodegenerative disease. A decrease in autophagy during aging contributes to brain disorders by accumulating potentially toxic substrates in neurons. Rubicon is a well-established inhibitor of autophagy in all cells. However, Rubicon participates in different pathways depending on cell type, and little information is currently available on neuronal Rubicon's role in the AD context. Here, we investigated the cell-specific expression of Rubicon in postmortem brain samples from AD patients and 5xFAD mice and its impact on amyloid ß burden in vivo and neuroblastoma cells. Further, we assessed Rubicon levels in human-induced pluripotent stem cells (hiPSCs), derived from early-to-moderate AD and in postmortem samples from severe AD patients. We found increased Rubicon levels in AD-hiPSCs and postmortem samples and a notable Rubicon localization in neurons. In AD transgenic mice lacking Rubicon, we observed intensified amyloid ß burden in the hippocampus and decreased Pacer and p62 levels. In APP-expressing neuroblastoma cells, increased APP/amyloid ß secretion in the medium was found when Rubicon was absent, which was not observed in cells depleted of Atg5, essential for autophagy, or Rab27a, required for exosome secretion. Our results propose an uncharacterized role of Rubicon on APP/amyloid ß homeostasis, in which neuronal Rubicon is a repressor of APP/amyloid ß secretion, defining a new way to target AD and other similar diseases therapeutically.

DEF8 and Autophagy-Associated Genes Are Altered in Mild Cognitive Impairment, Probable Alzheimer's Disease Patients, and a Transgenic Model of the Disease.

BACKGROUND: Disturbances in the autophagy/endolysosomal systems are proposed as early signatures of Alzheimer's disease (AD). However, few studies are available concerning autophagy gene expression in AD patients. OBJECTIVE: To explore the differential expression of classical genes involved in the autophagy pathway, among them a less characterized one, DEF8 (Differentially expressed in FDCP 8), initially considered a Rubicon family member, in peripheralblood mononuclear cells (PBMCs) from individuals with mild cognitive impairment (MCI) and probable AD (pAD) and correlate the results with the expression of DEF8 in the brain of 5xFAD mice. METHOD: By real-time PCR and flow cytometry, we evaluated autophagy genes levels in PBMCs from MCI and pAD patients. We evaluated DEF8 levels and its localization in brain samples of the 5xFAD mice by real-time PCR, western blot, and immunofluorescence. RESULTS: Transcriptional levels of DEF8 were significantly reduced in PBMCs of MCI and pAD patients compared with healthy donors, correlating with the MoCA and MoCA-MIS cognitive tests scores. DEF8 protein levels were increased in lymphocytes from MCI but not pAD, compared to controls. In the case of brain samples from 5xFAD mice, we observed a reduced mRNA expression and augmented protein levels in 5xFAD compared to age-matched wild-type mice. DEF8 presented a neuronal localization. CONCLUSION: DEF8, a protein proposed to act at the final step of the autophagy/endolysosomal pathway, is differentially expressed in PBMCs of MCI and pAD and neurons of 5xFAD mice. These results suggest a potential role for DEF8 in the pathophysiology of AD.

New Molecular and Organelle Alterations Linked to Down Syndrome Heart Disease.

Down syndrome (DS) is a genetic disorder caused by a trisomy of the human chromosome 21 (Hsa21). Overexpression of Hsa21 genes that encode proteins and non-coding RNAs (ncRNAs) can disrupt several cellular functions and biological processes, especially in the heart. Congenital heart defects (CHDs) are present in 45-50% of individuals with DS. Here, we describe the genetic background of this condition (Hsa21 and non-Hsa21 genes), including the role of ncRNAs, and the relevance of these new players in the study of the pathophysiology of DS heart diseases. Additionally, we discuss several distinct pathways in cardiomyocytes which help maintain a functional heart, but that might trigger hypertrophy and oxidative stress when altered. Moreover, we highlight the importance of investigating how mitochondrial and lysosomal dysfunction could eventually contribute to understanding impaired heart function and development in subjects with the Hsa21 trisomy. Altogether, this review focuses on the newest insights about the gene expression, molecular pathways, and organelle alterations involved in the cardiac phenotype of DS.

Down syndrome and Alzheimer's disease: common molecular traits beyond the amyloid precursor protein.

Alzheimer's disease (AD) is the most prevalent type of dementia. Down syndrome (DS) is the leading genetic risk factor for Early-Onset AD, prematurely presenting the classic pathological features of the brain with AD. Augmented gene dosage, including the APP gene, could partially cause this predisposition. Recent works have revealed that alterations in chromosome location due to the extra Chromosome 21, as well as epigenetic modifications, could promote changes in gene expression other than those from Chromosome 21. As a result, similar pathological features and cellular dysfunctions in DS and AD, including impaired autophagy, lysosomal activity, and mitochondrial dysfunction, could be controlled beyond APP overexpression. In this review, we highlight some recent data regarding the origin of the shared features between DS and AD and explore the mechanisms concerning cognitive deficiencies in DS associated with dementia, which could shed some light into the search for new therapeutic targets for AD treatment.

Diversity of culturable bacteria recovered from Pico Bolívar's glacial and subglacial environments, at 4950 m, in Venezuelan tropical Andes.

Even though tropical glaciers are retreating rapidly and many will disappear in the next few years, their microbial diversity remains to be studied in depth. In this paper we report on the biodiversity of the culturable fraction of bacteria colonizing Pico Bolívar's glacier ice and subglacial meltwaters, at ∼4950 m in the Venezuelan Andean Mountains. Microbial cells of diverse morphologies and exhibiting uncompromised membranes were present at densities ranging from 1.5 × 104 to 4.7 × 104 cells/mL in glacier ice and from 4.1 × 105 to 9.6 × 105 cells/mL in subglacial meltwater. Of 89 pure isolates recovered from the samples, the majority were eurypsychrophilic or stenopsychrophilic, according to their temperature range of growth. Following analysis of their 16S rDNA nucleotidic sequence, 54 pure isolates were assigned to 23 phylotypes distributed within 4 different phyla or classes: Beta- and Gammaproteobacteria, Actinobacteria, and Bacteroidetes. Actinobacteria dominated the culturable fraction of glacier ice samples, whereas Proteobacteria were dominant in subglacial meltwater samples. Chloramphenicol and ampicillin resistance was exhibited by 73.07% and 65.38%, respectively, of the subglacial isolates, and nearly 35% of them were multiresistant. Considering the fast rate at which tropical glaciers are melting, this study confirms the urgent need to study the microbial communities immured in such environments.

Bioprospecting glacial ice for plant growth promoting bacteria.

Glaciers harbor a wide diversity of microorganisms, metabolically versatile, highly tolerant to multiple environmental stresses and potentially useful for biotechnological purposes. Among these, we hypothesized the presence of bacteria able to exhibit well-known plant growth promoting traits (PGP). These kinds of bacteria have been employed for the development of commercial biofertilizers; unfortunately, these biotechnological products have proven ineffective in colder climates, like the ones prevailing in mountainous ecosystems. In the present work, we prospected glacial ice collected from two small tropical glaciers, located above 4.900 m in the Venezuelan Andes, for cold-active PGP bacteria. The initial screening strategy allowed us to detect the best inorganic-P solubilizers at low temperatures, from a sub-sample of 50 bacterial isolates. Solubilization of tricalcium phosphate, aluminum- and iron-phosphate, occurred in liquid cultures at low temperatures and was dependent on medium acidification by gluconic acid production, when bacteria were supplied with an appropriate source of carbon. Besides, the isolates were psychrophilic and in some cases exhibited a broad range of growth-temperatures, from 4 °C to 30 °C. Additional PGP abilities, including phytohormone- and HCN production, siderophore excretion and inhibition of phytopathogens, were confirmed in vitro. Nucleotidic sequence analysis of 16S rRNA genes allowed us to place the isolates within the Pseudomonas genus. Our results support the possible use of these strains to develop cold-active biofertilizers to be used in mountainous agriculture.

Bacteria recovered from a high-altitude, tropical glacier in Venezuelan Andes.

Glacial-ice microorganisms are intensively studied world-wide for a number of reasons, including their psychrophilic lifestyle, their usefulness in biotechnology procedures and their relationship with the search of life outside our planet. However, because of the difficulties for accessing and working at altitudes of >5.000 m above sea level, tropical glaciers have received much less attention than their arctic and antarctic counterparts. In the present work we isolated and characterized a total of forty-five pure isolates originating from direct plating of melted ice collected at the base of a rapidly-retreating, small glacier located at around 4.900 m.a.s.l. in Mount Humboldt (Sierra Nevada National Park, Mérida State, Venezuela). Initial examination of melted ice showed the presence of abundant- (>106 cells ml⁻¹), morphologically diverse- and active bacterial cells, many of which were very small ("dwarf cells"). The majority of the isolates were psychrophilic or psychrotolerant and many produced and excreted cold-active extracellular enzymes (proteases and amylases). The antibiotic tests showed an elevated percentage of isolates resistant to high doses (100 µg/ml) of different antibiotics including ampicillin, penicillin, nalidixic acid, streptomycin, chloramphenicol, kanamycin and tetracycline. Multiresistance was also observed, with 22.22 % of the strains simultaneously resistant up to five of the antibiotics tested. Metal resistance against Ni⁺⁺, Zn⁺⁺ and Cu⁺⁺ was also detected. In accordance with these results, plasmids of low and high molecular weight were detected in 47 % of the isolates. Twenty-two partial 16S rDNA sequences analyzed allowed grouping the isolates within five different phyla/classes: Alpha-, Beta- and Gamma-proteobacteria, Actinobacteria and Flavobacteria. This is the first report concerning South American Andean glacial ice microorganisms.