Ablation of Promyelocytic Leukemia Protein (PML) Re-patterns Energy Balance and Protects Mice from Obesity Induced by a Western Diet

The promyelocytic leukemia protein is a well known tumor suppressor, but its role in metabolism is largely unknown. Mice with a deletion in the gene for PML (KO mice) exhibit altered gene expression in liver, adipose tissue, and skeletal muscle, an accelerated rate of fatty acid metabolism, abnormal glucose metabolism, constitutive AMP-activating kinase (AMPK) activation, and insulin resistance in skeletal muscle. Last, an increased rate of energy expenditure protects PML KO mice from the effects of obesity induced by a Western diet. Collectively, our study uncovers a previously unappreciated role of PML in the regulation of metabolism and energy balance in mice.     

VEGF gene mRNA expression in patients with coronary artery disease

To assess the expression of vascular endothelium growth factor (VEGF) mRNA in unstimulated peripheral blood mononuclear cells of patients with and without coronary artery disease (CAD). We also studied whether the functional VEGF -2,578C/A polymorphism may influence the level of VEGF mRNA expression in individuals undergoing coronary angiography because chest pain. We assessed 50 consecutive patients with angiographically confirmed CAD (CAD+). Also, 50 consecutive individuals with normal coronary studies were included in the study for comparison. VEGF mRNA expression was examined using quantitative real-time PCR and genotyping for VEGF -2,578C/A was performed using ARMS-PCR technique. VEGF mRNA expression was significantly decreased in CAD+ patients when compared to CAD- individuals (p = 0.01). The frequency of VEGF -2578 allele C and genotype CC was increased in CAD+ patients. In this regard, homozygosity for the CC genotype was more commonly observed in CAD+ (30 %) than in those without CAD disease (18 %). However, the difference was slightly out of the range of significance (p = 0.1). In addition, a trend for reduction in the expression of VEGF mRNA was observed when patients carrying the VEGF -2,578AA genotype were compared with those VEGF -2,578AC heterozygous or those homozygous for the VEGF -2,578CC genotype. VEGF gene expression is decreased in individuals with CAD+ disease. The VEGF -2,578C/A polymorphism may influences the expression of VEGF.     

Involvement of vesicle-cytoskeleton interaction in AQP5 trafficking in AQP5-gene-transfected HSG cells

We examined the regulatory role of a reduction/oxidation (redox) control protein, thioredoxin (TRX), in tumor necrosis factor-alpha (TNF-alpha)-induced p38 MAP kinase activation and p38 MAP kinase-mediated cytokine expression utilizing TRX-transfected murine L929 cells (TRX14). The results showed that TNF-alpha-induced p38 MAP kinase activation and interleukin-6 (IL-6) production by TRX 14 were less than those by the parental L cells and the control transfected L cells (Neo-1). SB 203580 as the specific inhibitor for p38 MAP kinase activity inhibited TNF-alpha-induced IL-6 production by the parental L cells, indicating that TNF-alpha-activated p38 MAP kinase regulates IL-6 production by the cell lines used in this study. These results showed that overexpression of TRX negatively regulates p38 MAP kinase activation and p38 MAP kinase-mediated IL-6 production by TNF-alpha-stimulated cells, indicating that TRX is critical for p38 MAP kinase activation which regulates cytokine expression.     

Buckyballs conjugated with nucleic acid sequences identifies microorganisms in live cell assays

Background

Rapid identification of bacteria can play an important role at the point of care, evaluating the health of the ecosystem, and discovering spatiotemporal distributions of a bacterial community. We introduce a method for rapid identification of bacteria in live cell assays based on cargo delivery of a nucleic acid sequence and demonstrate how a mixed culture can be differentiated using a simple microfluidic system.

Methods

C60 Buckyballs are functionalized with nucleic acid sequences and a fluorescent reporter to show that a diversity of microorganisms can be detected and identified in live cell assays. The nucleic acid complexes include an RNA detector, targeting a species-specific sequence in the 16S rRNA, and a complementary DNA with an attached fluorescent reporter. As a result, each bacterium can be detected and visualized at a specific emission frequency through fluorescence microscopy.

Results

The C60 probe complexes can detect and identify a diversity of microorganisms that include gram-position and negative bacteria, yeast, and fungi. More specifically, nucleic-acid probes are designed to identify mixed cultures of Bacillus subtilis and Streptococcus sanguinis, or Bacillus subtilis and Pseudomonas aeruginosa. The efficiency, cross talk, and accuracy for the C60 probe complexes are reported. Finally, to demonstrate that mixed cultures can be separated, a microfluidic system is designed that connects a single source-well to multiple sinks wells, where chemo-attractants are placed in the sink wells. The microfluidic system allows for differentiating a mixed culture.

Conclusions

The technology allows profiling of bacteria composition, at a very low cost, for field studies and point of care.

     

Neuraminidase-associated plasminogen recruitment enables systemic spread of natural avian Influenza viruses H3N1

Repeated outbreaks due to H3N1 low pathogenicity avian influenza viruses (LPAIV) in Belgium were associated with unusually high mortality in chicken in 2019. Those events caused considerable economic losses and prompted restriction measures normally implemented for eradicating high pathogenicity avian influenza viruses (HPAIV). Initial pathology investigations and infection studies suggested this virus to be able to replicate systemically, being very atypical for H3 LPAIV. Here, we investigate the pathogenesis of this H3N1 virus and propose a mechanism explaining its unusual systemic replication capability. By intravenous and intracerebral inoculation in chicken, we demonstrate systemic spread of this virus, extending to the central nervous system. Endoproteolytic viral hemagglutinin (HA) protein activation by either tissue-restricted serine peptidases or ubiquitous subtilisin-like proteases is the functional hallmark distinguishing (H5 or H7) LPAIV from HPAIV. However, luciferase reporter assays show that HA cleavage in case of the H3N1 strain in contrast to the HPAIV is not processed by intracellular proteases. Yet the H3N1 virus replicates efficiently in cell culture without trypsin, unlike LPAIVs. Moreover, this trypsin-independent virus replication is inhibited by 6-aminohexanoic acid, a plasmin inhibitor. Correspondingly, in silico analysis indicates that plasminogen is recruitable by the viral neuraminidase for proteolytic activation due to the loss of a strongly conserved N-glycosylation site at position 130. This mutation was shown responsible for plasminogen recruitment and neurovirulence of the mouse brain-passaged laboratory strain A/WSN/33 (H1N1). In conclusion, our findings provide good evidence in natural chicken strains for N1 neuraminidase-operated recruitment of plasminogen, enabling systemic replication leading to an unusual high pathogenicity phenotype. Such a gain of function in naturally occurring AIVs representing an established human influenza HA-subtype raises concerns over potential zoonotic threats.     

Evaluation of expression CXCL8 chemokine and its relationship with oocyte maturation and embryo quality in the intracytoplasmic sperm injection method

Molecular Biology Reports - The present study aimed to evaluate the expression of the chemokine CXCL8 in both mRNA and protein levels in the serum, follicular fluid (FF), and cumulus cells (CCs)...     

Biogenesis and topology of the secretory Na+-K+-2Cl- cotransporter (NKCC1) studied in intact mammalian cells

The "secretory" Na+-K+-2Cl- cotransporter (NKCC1) is a member of a small gene family with nine homologues in vertebrates. Of these, seven are known to be electroneutral chloride transporters. These transporters play a number of important physiological roles related to salt and water homeostasis and the control of intracellular chloride levels. Hydropathy analyses suggest that all of these transporters have a similar transmembrane topology consisting of relatively large intracellular N and C termini and a central hydrophobic domain containing 12 membrane-spanning segments (MSSs). In recent experiments from our laboratory [Gerelsaikhan, T., and Turner, R. J. (2000) J. Biol. Chem. 275, 40471-40477], we employed an in vitro translation system to confirm that each of the putative MSSs of NKCC1 was capable of membrane integration in a manner consistent with a 12 MSS model. Here, we extend that work to the study of the biogenesis of NKCC1 in intact cells. We employ a truncation mutant approach that allows us to monitor this process quantitatively as successive MSSs are synthesized. While the results presented here confirm the 12 MSS model, they also indicate that the integration of NKCC1 into the membrane does not occur via a simple cotranslational process. In particular, we demonstrate that two MSSs, the second and sixth, require the presence of downstream sequence to efficiently integrate into the membrane.     

Effect of exposure to extremely low electro-magnetic field during prenatal period on mice spleen

Total body weight of newborns, the volume of spleen, and the number of megakaryocytes decreased following the exposure to ELF-MF (6 x 10(-3) T and 50 Hz) at 1-5, 6-10, 11-15, and 16-20 days of pregnancy of mice. The complete period of gestation was sensitive to ELF-MF exposure; the initial days were more prone to exposure. The results suggest that the use of ELF-MF producing instruments should be limited during gestation.