Chromatin compaction and cell death by high molecular weight FGF-2 depend on its nuclear localization, intracrine ERK activation, and engagement of mitochondria

Fibroblast growth factor 2 (FGF-2) is produced as CUG-initiated, 22-34 kDa or AUG-initiated 18 kDa isoforms (hi- and lo-FGF-2, respectively), with potentially distinct functions. We report that expression of hi-FGF-2 in HEK293 cells elicited chromatin compaction preceding cell death with apoptotic features. Nuclear localization of the intact protein was required as expression of a non-nuclear hi-FGF-2 mutant failed to elicit chromatin compaction. Equally ineffective, despite nuclear localization, was the over-expression of the 18 kDa core sequence (lo-FGF-2). Chromatin compaction by hi-FGF-2 was accompanied by increased cytosolic cytochrome C, and was attenuated either by over-expression of Bcl-2 or by a peptide inhibitor of the pro-apoptotic protein Bax. In addition hi-FGF-2 elicited sustained activation of total and nuclear extracellular signal regulated kinase (ERK1/2) by an intracrine route, as it was not prevented by neutralizing anti-FGF-2 antibodies. Inhibition of the ERK1/2 activating pathway by dominant negative upstream activating kinase, or by PD 98059, prevented chromatin compaction by hi-FGF-2. ERK1/2 activation was not affected by the Bax-inhibiting peptide suggesting that it occurred upstream of mitochondrial involvement. We conclude that the hi-FGF-2-induced chromatin compaction and cell death requires its nuclear localization, intracrine ERK1/2 activation and mitochondrial engagement.     

Species-diagnostic single-nucleotide polymorphism and sequence-tagged site markers for the parasitic wasp genus Nasonia (Hymenoptera: Pteromalidae)

Wasps of the genus Nasonia are important biological control agents of house flies and related filth flies, which are major vectors of human pathogens. Species of Nasonia (Hymenoptera: Pteromalidae) are not easily differentiated from one another by morphological characters, and molecular markers for their reliable identification have been missing so far. Here, we report eight single-nucleotide polymorphism and three sequence-tagged site markers derived from expressed sequenced tag libraries for the two closely related and regionally sympatric species N. giraulti and N. vitripennis. We studied variation of these markers in natural populations of the two species, and we mapped them in the Nasonia genome. The markers are species-diagnostic and evenly spread over all five chromosomes. They are ideal for rapid species identification and hybrid recognition, and they can be used to map economically relevant quantitative trait loci in the Nasonia genome.     

SIRT1 deacetylase protects against neurodegeneration in models for Alzheimer's disease and amyotrophic lateral sclerosis

A progressive loss of neurons with age underlies a variety of debilitating neurological disorders, including Alzheimer's disease (AD) and amyotrophic lateral sclerosis (ALS), yet few effective treatments are currently available. The SIR2 gene promotes longevity in a variety of organisms and may underlie the health benefits of caloric restriction, a diet that delays aging and neurodegeneration in mammals. Here, we report that a human homologue of SIR2, SIRT1, is upregulated in mouse models for AD, ALS and in primary neurons challenged with neurotoxic insults. In cell-based models for AD/tauopathies and ALS, SIRT1 and resveratrol, a SIRT1-activating molecule, both promote neuronal survival. In the inducible p25 transgenic mouse, a model of AD and tauopathies, resveratrol reduced neurodegeneration in the hippocampus, prevented learning impairment, and decreased the acetylation of the known SIRT1 substrates PGC-1alpha and p53. Furthermore, injection of SIRT1 lentivirus in the hippocampus of p25 transgenic mice conferred significant protection against neurodegeneration. Thus, SIRT1 constitutes a unique molecular link between aging and human neurodegenerative disorders and provides a promising avenue for therapeutic intervention.     

Dynamic changes in nitric oxide and mitochondrial oxidative stress with site-dependent differential tissue response during anoxic preconditioning in rat heart

In this study, dynamic changes in nitric oxide (NO) and mitochondrial superoxide (O(2)(*-)) were examined during anoxic preconditioning (AP) in rat heart model. AP and anoxia-reoxygenation (A/R) were performed on isolated hearts and single cardiomyocytes. The cellular insult in the form of infarct size and DNA damage were localized and correlated with NO synthases (endothelial and inducible) expression levels. The results showed that endocardium was the most affected region in AP groups, whereas the larger area of infarct was confined to mid- and epicardium in the A/R group. Interestingly, a high-level expression of immunofluorescent NO synthases was restricted to viable areas in the AP. In contrast to the gradual increase in O(2)(*-) level that occurred in the AP group, a sudden massive increase in its level was demonstrated at the onset of reoxygenation in the A/R group. The observed increase in NO production during reoxygenation in the AP group was attenuated by inducible NO synthase inhibitor. The study revealed, on a real-time basis, the role played by preconditioning for modulating NO and O(2)(*-) levels on behalf of cell survival. The results afford a better understanding of cardiac-adapting mechanism during AP and the role of inducible NO synthase in this important phenomenon.     

Synthesis and antibacterial activity of novel fluoroquinolones containing substituted piperidines

The design and synthesis of new fluoroquinolone antibacterial agents having substituted piperidine rings at the C-7 position are described. Most of the new compounds demonstrated high in vitro antibacterial activity. Several of them exhibited significant activities against gram-positive organisms, which were more potent than those of gemifloxacin, Linezolid, and vancomycin.     

The Pivotal Role of Astrocytes in the Metabolism of Iron in the Brain

Iron is essential for the normal functioning of cells but since it is also capable of generating toxic reactive oxygen species, the metabolism of iron is tightly regulated. The present article advances the view that astrocytes are largely responsible for distributing iron in the brain. Capillary endothelial cells are separated from the neuropil by the endfeet of astrocytes, so astrocytes are ideally positioned to regulate the transport of iron to other brain cells and to protect them if iron breaches the blood-brain barrier. Astrocytes do not appear to have a high metabolic requirement for iron yet they possess transporters for transferrin, haemin and non-transferrin-bound iron. They store iron efficiently in ferritin and can export iron by a mechanism that involves ferroportin and ceruloplasmin. Since astrocytes are a common site of abnormal iron accumulation in ageing and neurodegenerative disorders, they may represent a new therapeutic target for the treatment of iron-mediated oxidative stress.     

Lysosomal trafficking functions of mucolipin-1 in murine macrophages

Background  

Mucolipidosis Type IV is currently characterized as a lysosomal storage disorder with defects that include corneal clouding, achlorhydria and psychomotor retardation. MCOLN1, the gene responsible for this disease, encodes the protein mucolipin-1 that belongs to the "Transient Receptor Potential" family of proteins and has been shown to function as a non-selective cation channel whose activity is modulated by pH. Two cell biological defects that have been described in MLIV fibroblasts are a hyperacidification of lysosomes and a delay in the exit of lipids from lysosomes.     

Whole genome analysis of Vietnamese G2P[4] rotavirus strains possessing the NSP2 gene sharing an ancestral sequence with Chinese sheep and goat rotavirus strains

Because imminent introduction into Vietnam of a vaccine against Rotavirus A is anticipated, baseline information on the whole genome of representative strains is needed to understand changes in circulating strains that may occur after vaccine introduction. In this study, the whole genomes of two G2P[4] strains detected in Nha Trang, Vietnam in 2008 were sequenced, this being the last period during which virtually no rotavirus vaccine was used in this country. The two strains were found to be > 99.9% identical in sequence and had a typical DS‐1 like G2‐P[4]‐I2‐R2‐C2‐M2‐A2‐N2‐T2‐E2‐H2 genotype constellation. Analysis of the Vietnamese strains with > 184 G2P[4] strains retrieved from GenBank/EMBL/DDBJ DNA databases placed the Vietnamese strains in one of the lineages commonly found among contemporary strains, with the exception of the NSP2 and NSP4 genes. The NSP2 genes were found to belong to a previously undescribed lineage that diverged from Chinese sheep and goat rotavirus strains, including a Chinese rotavirus vaccine strain LLR with 95% nucleotide identity; the time of their most recent common ancestor was 1975. The NSP4 genes were found to belong, together with Thai and USA strains, to an emergent lineage (VIII), adding further diversity to ever diversifying NSP4 lineages. Thus, there is a need to enhance surveillance of locally‐circulating strains from both children and animals at the whole genome level to address the effect of rotavirus vaccines on changing strain distribution.     

Seasonal variation in in vitro immune activity of milk leukocytes in elite and non-elite crossbred cows of Indian sub-tropical semi-arid climate

Immunity of mammary gland in terms of in vitro activity of milk leukocytes has been evaluated during hot-humid, summer, and winter season in elite (n = 10) and non-elite (n = 10) crossbred cows. Milk samples were collected from all the cows throughout the year at 15-day interval. Milk somatic cell counts (SCC) and differential leukocyte counts (DLC) were evaluated microscopically. Milk neutrophils, macrophages, and lymphocytes were isolated and cultured in vitro. In vitro PI of milk neutrophils and macrophages was evaluated by colorimetric NBT (nitro-blue tetrazolium) reductive assay. Mitogen-induced milk lymphocyte blastogenic response was measured by colorimetric MTT (thiazolyl blue tetrazolium bromide) assay. Milk SCC was found to be significantly (p < 0.01) higher in elite cows compared to non-elite cows irrespective of season. There was significant (p < 0.05) increase in milk SCC during hot-humid season compared to winter season in both the group of the cows. There was no significant difference between group and season in terms of DLC. In vitro phagocytic index of elite cows was significantly (p < 0.01) higher than non-elite cows. The phagocytic index was significantly (p < 0.01) decreased in summer and hot-humid season compared to winter season in both the group of animals. Macrophages isolated from elite cows having significantly (p < 0.01) lower phagocytic index than non-elite cows which significantly (p < 0.01) decreased during summer and hot-humid season compared to winter. In vitro milk lymphocyte proliferative response was significantly (p < 0.01) lower in elite cows. Activity of B-lymphocytes decreased significantly (p < 0.01) during summer and hot-humid season than winter, but activity of T-lymphocytes remains unaltered during different seasons. In conclusion, the mammary immunity in terms of in vitro activity of milk leukocytes is compromised during summer and hot-humid season in elite crossbred cows; therefore, better care and management should be taken in high-yielding cows during summer and hot-humid season to minimize intramammary infections.