Synthesis, crystal structure and characterization of alkali metal hydroxoantimonates

Several alkali metal hydroxoantimonates, K₂[Sb(O)(OH)₅], Na[Sb(OH)₆], Cs[Sb(OH)₆] and Cs₂[Sb₂(μ-O)₂(OH)₈] were isolated from aqueous solutions and characterized by single crystal and powder X-ray diffraction studies and by FTIR and thermal analysis. Crystal structures involving [Sb(O)(OH)₅]²⁻ were never anticipated before, and this is also the first disclosure of a dinuclear antimonate [Sb₂(μ-O)₂(OH)₈]²⁻. Aqueous antimonate solutions of different pH were studied by high resolution electrospray mass spectrometry showing pH indifferent spectra and predominance of the mono and dinuclear antimonate species at pH 4-10.     

Pyramided rice lines harbouring Allium sativum (asal) and Galanthus nivalis (gna) lectin genes impart enhanced resistance against major sap-sucking pests

We have developed transgene pyramided rice lines, endowed with enhanced resistance to major sap-sucking insects, through sexual crosses made between two stable transgenic rice lines containing Allium sativum (asal) and Galanthus nivalis (gna) lectin genes. Presence and expression of asal and gna genes in pyramided lines were confirmed by PCR and western blot analyses. Segregation analysis of F₂ progenies disclosed digenic (9:3:3:1) inheritance of the transgenes. Homozygous F₃ plants carrying asal and gna genes were identified employing genetic and molecular methods besides insect bioassays. Pyramided lines, infested with brown planthopper (BPH), green leafhopper (GLH) and whitebacked planthopper (WBPH), proved more effective in reducing insect survival, fecundity, feeding ability besides delayed development of insects as compared to the parental transgenics. Under infested conditions, pyramided lines were found superior to the parental transgenics in their seed yield potential. This study represents first report on pyramiding of two lectin genes into rice exhibiting enhanced resistance against major sucking pests. The pyramided lines appear promising and might serve as a novel genetic resource in rice breeding aimed at durable and broad based resistance against hoppers.     

Functional and Molecular Characterization of Hyposensitive Underactive Bladder Tissue and Urine in Streptozotocin-Induced Diabetic Rat

Background

The functional and molecular alterations of nerve growth factor (NGF) and Prostaglandin E2 (PGE2) and its receptors were studied in bladder and urine in streptozotocin (STZ)-induced diabetic rats.

Methodology/Principal Findings

Diabetes mellitus was induced with a single dose of 45 mg/kg STZ Intraperitoneally (i.p) in female Sprague-Dawley rats. Continuous cystometrogram were performed on control rats and STZ treated rats at week 4 or 12 under urethane anesthesia. Bladder was then harvested for histology, expression of EP receptors and NGF by western blotting, PGE2 levels by ELISA, and detection of apoptosis by TUNEL staining. In addition, 4-hr urine was collected from all groups for urine levels of PGE2, and NGF assay. DM induced progressive increase of bladder weight, urine production, intercontraction interval (ICI) and residual urine in a time dependent fashion. Upregulation of Prostaglandin E receptor (EP)1 and EP3 receptors and downregulation of NGF expression, increase in urine NGF and decrease levels of urine PGE2 at week 12 was observed. The decrease in ICI by intravesical instillation of PGE2 was by 51% in control rats and 31.4% in DM group at week 12.

Conclusions/Significance

DM induced hyposensitive underactive bladder which is characterized by increased inflammatory reaction, apoptosis, urine NGF levels, upregulation of EP1 and EP3 receptors and decreased bladder NGF and urine PGE2. The data suggest that EP3 receptor are potential targets in the treatment of diabetes induced underactive bladder.     

The ubiquitin proteasome system acutely regulates presynaptic protein turnover and synaptic efficacy

BACKGROUND: The ubiquitin proteasome system (UPS) mediates regulated protein degradation and provides a mechanism for closely controlling protein abundance in spatially restricted domains within cells. We hypothesized that the UPS may acutely determine the local concentration of key regulatory proteins at neuronal synapses as a means for locally modulating synaptic efficacy and the strength of neurotransmission communication. RESULTS: We investigated this hypothesis at the Drosophila neuromuscular synapse by using an array of genetic and pharmacological tools. This study demonstrates that UPS components are present in presynaptic boutons and that the UPS functions locally in the presynaptic compartment to rapidly eliminate a conditional transgenic reporter of proteasome activity. We assayed a panel of synaptic proteins to determine whether the UPS acutely regulates the local abundance of native synaptic targets. Both acute pharmacological inhibition of the proteasome (<1 hr) and targeted genetic perturbation of proteasome function in the presynaptic neuron cause the specific accumulation of the essential synaptic vesicle-priming protein DUNC-13. Most importantly, acute pharmacological inhibition of the proteasome (<1 hr) causes a rapid strengthening of neurotransmission (an approximately 50% increase in evoked amplitude) because of increased presynaptic efficacy. The proteasome-dependent regulation of presynaptic protein abundance, both of the exogenous reporter and native DUNC-13, and the modulation of presynaptic neurotransmitter release occur on an intermediate, rapid (tens of minutes) timescale. CONCLUSIONS: Taken together, these studies demonstrate that the UPS functions locally within synaptic boutons to acutely control levels of presynaptic protein and that the rate of UPS-dependent protein degradation is a primary determinant of neurotransmission strength.     

Role of Bruton’s tyrosine kinase in macrophage apoptosis

Macrophages and polymorphonuclear cells (PMNs) rapidly respond to microbial and immune inflammatory stimuli and die during these responses. We have shown earlier that many macrophage and PMN functions are compromised in x-linked immunodeficient (Xid) mice with functional deficiency in Bruton’s tyrosine kinase (Btk). We now report that Btk-deficient macrophages show enhanced susceptibility to apoptotic death on exposure to the microbial and immune inflammatory signals bacterial lipopolysaccharide (LPS) and interferon-gamma (IFNγ) in vitro. In vivo in mixed bone marrow (BM) chimeras Btk deficiency leads primarily to loss of peripheral macrophage numbers without affecting BM development, suggesting a role of inflammation-induced apoptosis in regulating macrophage life span. Surprisingly, Btk deficiency does not affect macrophage apoptosis induced by DNA damage or CD95 engagement. Reactive nitrogen and oxygen species also do not contribute to inflammation-induced apoptosis, but apoptotic process involves loss of mitochondrial potential, shows increased activation of caspase 9 and enhanced loss of Bcl-xL. The lack of pro-survival signaling through the Btk-phosphotidylinositol 3-kinase-Akt pathway, and persistent MEK signaling, lead to enhanced death in Btk-deficient macrophages only downstream of inflammatory triggers. These data underline the complex role of Btk in the regulation of macrophage survival and function.     

Oxidation of Akt2 kinase promotes cell migration and regulates G1-S transition in the cell cycle

Phosphorylation has long been recognized as the key mediator of protein signaling. New modes of signaling regulation are emerging with the development of specific chemical probes and application of high-throughput mass spectrometry technologies. Using biotin-tagged chemical probes for protein oxidation, mass spectrometry and functional assays, our group has recently reported isoform-specific oxidation of Akt2 in response to PDGF signaling. The studies included here investigate the functional consequence of oxidation on Akt2-mediated cell migration and cell cycle. Akt2-KO MEFs transduced with WT and Cys124Ser Akt2 were used as the model system for these studies. The implications of these findings on disease pathology are discussed.Key words: oxidation, ROS, cell migration, cell cycle, wound healing, Akt2, starvation     

Improved responsiveness and reduced sample size requirements of PROMIS physical function scales with item response theory

Introduction  

The Health Assessment Questionnaire Disability Index (HAQ) and the SF-36 PF-10, among other instruments, yield sensitive and valid Disability (Physical Function) endpoints. Modern techniques, such as Item Response Theory (IRT), now enable development of more precise instruments using improved items. The NIH Patient Reported Outcomes Measurement Information System (PROMIS) is charged with developing improved IRT-based tools. We compared the ability to detect change in physical function using original (Legacy) instruments with Item-Improved and PROMIS IRT-based instruments.     

Neurogenesis response of middle-aged hippocampus to acute seizure activity

Acute Seizure (AS) activity in young adult age conspicuously modifies hippocampal neurogenesis. This is epitomized by both increased addition of new neurons to the granule cell layer (GCL) by neural stem/progenitor cells (NSCs) in the dentate subgranular zone (SGZ), and greatly enhanced numbers of newly born neurons located abnormally in the dentate hilus (DH). Interestingly, AS activity in old age does not induce such changes in hippocampal neurogenesis. However, the effect of AS activity on neurogenesis in the middle-aged hippocampus is yet to be elucidated. We examined hippocampal neurogenesis in middle-aged F344 rats after a continuous AS activity for >4 hrs, induced through graded intraperitoneal injections of the kainic acid. We labeled newly born cells via daily intraperitoneal injections of the 5'-bromodeoxyuridine (BrdU) for 12 days, commencing from the day of induction of AS activity. AS activity enhanced the addition of newly born BrdU+ cells by 5.6 fold and newly born neurons (expressing both BrdU and doublecortin [DCX]) by 2.2 fold to the SGZ-GCL. Measurement of the total number of DCX+ newly born neurons also revealed a similar trend. Furthermore, AS activity increased DCX+ newly born neurons located ectopically in the DH (2.7 fold increase and 17% of total newly born neurons). This rate of ectopic migration is however considerably less than what was observed earlier for the young adult hippocampus after similar AS activity. Thus, the plasticity of hippocampal neurogenesis to AS activity in middle age is closer to its response observed in the young adult age. However, the extent of abnormal migration of newly born neurons into the DH is less than that of the young adult hippocampus after similar AS activity. These results also point out a highly divergent response of neurogenesis to AS activity between middle age and old age.     

A metalloproteinase secreted by Streptococcus pneumoniae removes membrane mucin MUC16 from the epithelial glycocalyx barrier

The majority of bacterial infections occur across wet-surfaced mucosal epithelia, including those that cover the eye, respiratory tract, gastrointestinal tract and genitourinary tract. The apical surface of all these mucosal epithelia is covered by a heavily glycosylated glycocalyx, a major component of which are membrane-associated mucins (MAMs). MAMs form a barrier that serves as one of the first lines of defense against invading bacteria. While opportunistic bacteria rely on pre-existing defects or wounds to gain entry to epithelia, non opportunistic bacteria, especially the epidemic disease-causing ones, gain access to epithelial cells without evidence of predisposing injury. The molecular mechanisms employed by these non opportunistic pathogens to breach the MAM barrier remain unknown. To test the hypothesis that disease-causing non opportunistic bacteria gain access to the epithelium by removal of MAMs, corneal, conjunctival, and tracheobronchial epithelial cells, cultured to differentiate to express the MAMs, MUCs 1, 4, and 16, were exposed to a non encapsulated, non typeable strain of Streptococcus pneumoniae (SP168), which causes epidemic conjunctivitis. The ability of strain SP168 to induce MAM ectodomain release from epithelia was compared to that of other strains of S. pneumoniae, as well as the opportunistic pathogen Staphylococcus aureus. The experiments reported herein demonstrate that the epidemic disease-causing S. pneumoniae species secretes a metalloproteinase, ZmpC, which selectively induces ectodomain shedding of the MAM MUC16. Furthermore, ZmpC-induced removal of MUC16 from the epithelium leads to loss of the glycocalyx barrier function and enhanced internalization of the bacterium. These data suggest that removal of MAMs by bacterial enzymes may be an important virulence mechanism employed by disease-causing non opportunistic bacteria to gain access to epithelial cells to cause infection.