Role of IFN-gamma in regulating T2 immunity and the development of alternatively activated macrophages during allergic bronchopulmonary mycosis

Pulmonary Cryptococcus neoformans infection of C57BL/6 mice is an established model of a chronic pulmonary fungal infection accompanied by an "allergic" response (T2) to the infection, i.e., a model of an allergic bronchopulmonary mycosis. Our objective was to determine whether IFN-gamma plays a role in regulating the pulmonary T2 immune response in C. neoformans-infected C57BL/6 mice. Long-term pulmonary fungistasis was lost in IFN-gamma knockout (KO) mice, resulting in an increased pulmonary burden of fungi at wk 3. IFN-gamma was required for the early influx of leukocytes into the lungs but was not required later in the infection. By wk 3, eosinophil and macrophage numbers were elevated in the absence of IFN-gamma. The inducible NO synthase to arginase ratio was lower in the lungs of IFN-gamma KO mice and the macrophages had increased numbers of intracellular cryptococci and YM1 crystals, indicative of alternatively activated macrophages in these mice. There was evidence of pulmonary fibrosis in both wild-type and IFN-gamma KO mice by 5 wk postinfection. IFN-gamma production was not required for the development of T2 cytokine (IL-4, IL-5, IL-13) producing cells in the lungs and lung-associated lymph nodes or induction of an IgE response. At a number of time points, T2 cytokine production was enhanced in IFN-gamma KO mice. Thus, in the absence of IFN-gamma, C57BL/6 mice develop an augmented allergic response to C. neoformans, including enhanced generation of alternatively activated macrophages, which is accompanied by a switch from a chronic to a progressive pulmonary cryptococcal infection.     

<Emphasis Type="Italic">C.el</Emphasis> Phosphatome: A Catalogue of Actual and Pseudo Phosphatases Based on In-Silico Studies in <Emphasis Type="Italic">Caenorhabditis elegans</Emphasis>

Phosphatases are well known to carry out important functions via counter activity of kinases and they serve as mechanism for dephosphorylating the monophosphate esters from the phosphorylated serine, threonine, tyrosine and histidine residues. The biological relevance of phosphatases could be explored further employing newer technologies and models. Caenorhabditis elegans is a powerful genetic model system that bears significant homology with humans, hence providing with a precious tool towards studying important signalling pathways. We carried out the present study to catalogue the C. elegans protein phosphatome, referred here as ‘C.el phosphatome’ and annotated the corresponding dataset. We further classified these phosphatases based on presence of catalytic conserved motif; GDxHG, GDxVDRG, GNHE, RxxD, DGxxG, DG, GxxDN for Ser/Thr phosphatases, HC(x)5 R for tyrosine phosphatases and DxDxT/V for aspartate based phosphatases. Bioinformatics tool DAVID was employed to decipher the biological relevance of phosphatases. Our findings show Ser/Thr phosphatases (114), Tyr phosphatases (121) and Asp phosphatases (0) in C. elegans genome based on the hallmark sequence identification. Amongst them, 34 and 57 Ser/Thr and Tyr phosphatases respectively contain the catalytic motif. This catalogue offers a precious tool for further studies towards understanding important biological processes and disease conditions.     

Hydrolysis of di-substituted hydantoins,by an enzyme preparation from lentil (Lens esculenta) seeds,for the synthesis of α,α-dialkylated amino acids with linear and cyclic substituents

Hydrolysis of 5,5-di-substituted hydantoins with linear and cyclic side chains, to produce the corresponding ,-dialkylated amino acids and 1-aminocycloalkane-1-carboxylic acids, using an enzyme preparation from lentil (Lens esculenta) seeds is reported for the first time. This method could be developed on a large scale and should be of interest to the chemical and pharmaceutical industries.R. Rai and V. Taneja are with the Department of Biochemistry, Faculty of Science, Banaras Hindu University, Varanasi-221 005, India; R. Balaji Rao is with the Department of Chemistry, Faculty of Science, Banaras Hindu University, Varanasi-221 005, India.     

Characterization of three HLA-DR beta genes isolated from an HLA-DR 3/4 insulin-dependent diabetic patient

Three HLA-DR genes were isolated from a Swedish HLA-DR3/4 insulin-dependent diabetes mellitus (IDDM) patient and characterized by restriction endonuclease mapping and nucleotide sequence analysis. Two out of the three DNA sequences differed from those of published DR-chain sequences. A DR-gene probe prepared from exon 4 and flanking sequences was used in a Southern blot analysis of blood donors' DNA and DNA from HLA-DR3/4 IDDM patients and HLA-DR-matched healthy control subjects. This probe differentiated HLA-DR3/4 IDDM patients and HLA-DR-matched controls in the Scandinavian population but not in the North American Caucasoid population.     

Detection and localization of actin mRNA isoforms in chicken muscle cells by in situ hybridization using biotinated oligonucleotide probes

We have developed in situ hybridization methodology for nonisotopically labeled oligonucleotide probes to detect cellular mRNA with improved speed, convenience, and resolution over previous techniques. Previous work using isotopically labeled oligonucleotide probes characterized important parameters for in situ hybridization (Anal Biochem 166:389, 1987). Eleven oligonucleotide probes were made to coding and noncoding regions of chick beta-actin mRNA and one oligonucleotide probe to chick alpha-cardiac actin mRNA. All the probes were 3' end-labeled with bio-11-dUTP using terminal transferase, and the labeled probes were hybridized to chicken myoblast and myotube cultures. The hybridized probe was detected using a streptavidin-alkaline phosphatase conjugate. Our assay for the success of probe hybridization and detection was the demonstration of beta-actin mRNA highly localized in the lamellipodia of single cells (Lawrence and Singer, Cell 45:407, 1986) as well as the expression of alpha-cardiac actin mRNA and the repression of beta-actin mRNA in differentiating myoblasts and in myotubes. With the alpha-cardiac probe, we found that this mRNA was distributed all over the cytoplasm of myotubes and differentiated (bipolar) single cells and negative in undifferentiated single cells and at the ends of myotubes. When beta-actin probes were used, two of 11 probes were highly sensitive, and, in pooling them together, the localization of beta-actin mRNA in fibroblastic single cells was evident at the leading edge of the motile cells, the lamellipodium. beta-Actin mRNA was not detected in myotubes except at the ends where contact was made with substrate. This indicates that both beta and cardiac actin mRNA can coexist in the same myotube cytoplasm but at different locations.     

Metal phytoremediation potential of naturally growing plants on fly ash dumpsite of Patratu thermal power station,Jharkhand, India

Three naturally growing plants Ipomoea carnea, Lantana camara, and Solanum surattense were found in fly ash dumpsite of Patratu thermal power station, Jharkhand, India. They were assessed for their metal uptake potential. The fly ash was slightly alkaline with very less nitrogen and organic carbon but enriched with phosphorus and heavy metals. Lantana camara and Ipomoea carnea showed good translocation from root to shoot for most of the metals except Mn and Pb. The order of metal accumulation in stem of both the plants were Fe(205mg/kg)>Mn(65mg/kg)>Cu(22.35mg/kg)>Pb(6.6mg/kg)>Cr(3.05mg/kg)>Ni(1 mg/kg)>Cd(0.5 mg/kg) and Fe(741 mg/kg)>Mn(154.05 mg/kg)>Cu(20.75 mg/kg)>Pb(6.75 mg/kg)>Ni(4.0 mg/kg)>Cr(3.3mg/kg)>Cd(0.05mg/kg), respectively. But Solanum surattense accumulated most of the metals in roots. The order was in the following order, Mn (382.2mg/kg) >Fe (264.1mg/kg) > Cu (25.35mg/kg) >Pb (5.95 mg/kg) > Ni (1.9 mg/kg) > Cr (1.8mg/kg) > Cd (0.55 mg/kg). The order of Bioconcentration factor (BCF) in root and shoot followed almost the same order as, Mn>Fe>Ni>Pb>Cu>Cr≈ Cd in all the three species. ANOVA showed significant variation in metal accumulation by root and stem between the species. Finally, it can be concluded that Solanum surattense can be used as phytostabilizer and other two species as phytoextractor of metal for fly ash dumpsite reclamation.     

Expression of A152T human tau causes age‐dependent neuronal dysfunction and loss in transgenic mice

A152T‐variant human tau (hTau‐A152T) increases risk for tauopathies, including Alzheimer's disease. Comparing mice with regulatable expression of hTau‐A152T or wild‐type hTau (hTau‐WT), we find age‐dependent neuronal loss, cognitive impairments, and spontaneous nonconvulsive epileptiform activity primarily in hTau‐A152T mice. However, overexpression of either hTau species enhances neuronal responses to electrical stimulation of synaptic inputs and to an epileptogenic chemical. hTau‐A152T mice have higher hTau protein/mRNA ratios in brain, suggesting that A152T increases production or decreases clearance of hTau protein. Despite their functional abnormalities, aging hTau‐A152T mice show no evidence for accumulation of insoluble tau aggregates, suggesting that their dysfunctions are caused by soluble tau. In human amyloid precursor protein (hAPP) transgenic mice, co‐expression of hTau‐A152T enhances risk of early death and epileptic activity, suggesting copathogenic interactions between hTau‐A152T and amyloid‐β peptides or other hAPP metabolites. Thus, the A152T substitution may augment risk for neurodegenerative diseases by increasing hTau protein levels, promoting network hyperexcitability, and synergizing with the adverse effects of other pathogenic factors.