Autonomic dysfunction and plasticity in micturition reflexes in human α-synuclein mice

Although often overshadowed by the motor dysfunction associated with Parkinson's disease (PD), autonomic dysfunction including urinary bladder and bowel dysfunctions are often associated with PD and may precede motoric changes; such autonomic dysfunction may permit early detection and intervention. Lower urinary tract symptoms are common in PD patients and result in significant morbidity. This studies focus on nonmotor symptoms in PD using a transgenic mouse model with overexpression of human α-synuclein (hSNCA), the peptide found in high concentrations in Lewy body neuronal inclusions, the histopathologic hallmark of PD. We examined changes in the physiological, molecular, chemical, and electrical properties of neuronal pathways controlling urinary bladder function in transgenic mice. The results of these studies reveal that autonomic dysfunction (i.e., urinary bladder) can precede motor dysfunction. In addition, mice with hSNCA overexpression in relevant neuronal populations is associated with alterations in expression of neurotransmitter/neuromodulatory molecules (PACAP, VIP, substance P, and neuronal NOS) within neuronal pathways regulating bladder function as well as with increased NGF expression in the urinary bladder. Changes in the electrical and synaptic properties of neurons in the major pelvic ganglia that provide postganglionic innervation to urogenital tissues were not changed as determined with intracellular recording. The urinary bladder dysfunction observed in transgenic mice likely reflects changes in peripheral (i.e., afferent) and/or central micturition pathways or changes in the urinary bladder. SYN-OE mice provide an opportunity to examine early events underlying the molecular and cellular plasticity of autonomic nervous system pathways underlying synucleinopathies.     

Regulation of secretory granule pH in insulin-secreting cells

Luminal acidification is important for the maturation of secretory granules, yet little is known regarding the regulation of pH within them. A pH-sensitive green fluorescent protein (EGFP) was targeted to secretory granules in RIN1046-38 insulinoma cells by using a construct in which the EGFP gene was preceded by the nucleotide sequence for human growth hormone. Stimulatory levels of glucose doubled EGFP secretion from cell cultures, and potentiators of glucose-induced insulin secretion enhanced EGFP release. Thus this targeted EGFP is useful for population measurements of secretion. However, less than ~4% of total cell EGFP was released after 1.5 h of stimulation. Consequently, when analyzed in single cells, fluorescence of the targeted EGFP acts as an indicator of pH within secretory granules. Glucose elicited a decrease in granule pH, whereas inhibitors of the V-type H(+)-ATPase increased pH and blocked the glucose effect. Granule pH also was modified by effectors of the protein kinase A pathway, with activation eliciting granule alkalinization, suggesting that potentiation of peptide release by cAMP may involve regulated changes in secretory granule pH.     

quick-to-court, a Drosophila mutant with elevated levels of sexual behavior, is defective in a predicted coiled-coil protein

Remarkably little is known about the molecular mechanisms that drive sexual behavior. We have identified a new gene, quick-to-court (qtc), whose mutations cause males to show high levels of male-male courtship. qtc males also show a novel phenotype: when placed in the presence of a virgin female, they begin courtship abnormally quickly. qtc mutations are striking in their specificity, in that many aspects of male sexual behavior are normal. We have cloned the qtc gene and found that it encodes a predicted coiled-coil protein and is expressed in the olfactory organs, central nervous system, and male reproductive tract.     

Nectar to improve parasitoid fitness in biological control: Does the sucrose:hexose ratio matter?

The consumption of saccharide-rich foods such as floral nectar is crucial for the survival of many adult parasitoid wasps. The importance to parasitoids of nectar quality, with regards to its sucrose:hexose ratio, was investigated. Nectar, an aqueous solution of sugars, amino acids and other compounds, differs between plant species. Nectar composition is dominated by sucrose, glucose and fructose. Previous studies have shown that the ratio of sucrose to hexose (glucose+fructose) sugars can explain nectar associations in a range of flower visiting arthropods. It has been suggested that this ratio may be important in terms of parasitoid fitness. Analysis of floral nectar from fourteen plant species confirmed that the sucrose/hexose ratio significantly differed between species. An opportunity to select floral resources based on this measure of nectar quality arose and highlighted the potential to utilize native flowering plant species in place of the seven most commonly deployed, which are usually not native to the countries in which they are used.Results presented in this paper indicate, however, that the sucrose/hexose ratio is not a significant factor explaining parasitoid longevity. The hymenopteran parasitoids Diadegma semiclausum (Ichneumonidae) and Dolichogenidea tasmanica (Braconidae) were fed 40% w/w sugar solutions, differing in their sugar ratios. Solutions were classified as either sucrose-dominant (ratio >0.99), sucrose-rich (ratio 0.5–0.99), hexose-rich (ratio 0.1–0.499) or hexose-dominant (ratio <0.1). No significant differences in parasitoid longevity were found between the different treatments for either species. This suggests there is not an optimal sucrose/hexose ratio for parasitoid wasps, although a greater number of parasitoid species should ideally be tested to confirm if this is true for the wider parasitoid taxonomic groups.     

Induction and Role of Indoleamine 2,3 Dioxygenase in Mouse Models of Influenza A Virus Infection

Influenza infection stimulates protective host immune responses but paradoxically enhances lung indoleamine 2,3 dioxygenase (IDO) activity, an enzyme that suppresses helper/effector T cells and activates Foxp3-lineage regulatory CD4 T cells (Tregs). Influenza A/PR/8/34 (PR8) infection stimulated rapid elevation of IDO activity in lungs and lung-draining mediastinal lymph nodes (msLNs). Mice lacking intact IDO1 genes (IDO1-KO mice) exhibited significantly lower morbidity after sub-lethal PR8 infection, and genetic or pharmacologic IDO ablation led to much faster recovery after virus clearance. More robust influenza-specific effector CD8 T cell responses manifested in lungs of PR8-infected IDO1-KO mice, though virus clearance rates were unaffected by IDO ablation. Similar outcomes manifested in mice infected with a less virulent influenza A strain (X31). IDO induction in X31-infected lungs was dependent on IFN type II (IFNγ) signaling and was restricted to non-hematopoietic cells, while redundant IFN type 1 or type II signaling induced IDO exclusively in hematopoietic cells from msLNs. Memory T cells generated in X31-primed IDO1-KO mice protected mice from subsequent challenge with lethal doses of PR8 (100×LD₅₀). However recall T cell responses were less robust in lung interstitial tissues, and classic dominance of TCR Vβ8.3 chain usage amongst memory CD8⁺ T cells specific for influenza nucleoprotein (NP₃₆₆) did not manifest in IDO1-KO mice. Thus, influenza induced IDO activity in lungs enhanced morbidity, slowed recovery, restrained effector T cell responses in lungs and shaped memory T cell repertoire generation, but did not attenuate virus clearance during primary influenza A infection.     

ARTEMIS stabilizes the genome and modulates proliferative responses in multipotent mesenchymal cells

Background  

Unrepaired DNA double-stranded breaks (DSBs) cause chromosomal rearrangements, loss of genetic information, neoplastic transformation or cell death. The nonhomologous end joining (NHEJ) pathway, catalyzing sequence-independent direct rejoining of DSBs, is a crucial mechanism for repairing both stochastically occurring and developmentally programmed DSBs. In lymphocytes, NHEJ is critical for both development and genome stability. NHEJ defects lead to severe combined immunodeficiency (SCID) and lymphoid cancer predisposition in both mice and humans. While NHEJ has been thoroughly investigated in lymphocytes, the importance of NHEJ in other cell types, especially with regard to tumor suppression, is less well documented. We previously reported evidence that the NHEJ pathway functions to suppress a range of nonlymphoid tumor types, including various classes of sarcomas, by unknown mechanisms.     

Reduced IgG anti-small nuclear ribonucleoprotein autoantibody production in systemic lupus erythematosus patients with positive IgM anti-cytomegalovirus antibodies

Introduction

In rheumatoid arthritis (RA), synovial fluid (SF) contains a large number of neutrophils that contribute to the inflammation and destruction of the joints. The SF also contains granulocyte-macrophage colony-stimulating factor (GM-CSF), which sustains viability of neutrophils and activates their functions. Using proteomic surveillance, we here tried to elucidate the effects of GM-CSF on neutrophils.

Methods

Neutrophils stimulated by GM-CSF were divided into four subcellular fractions: cytosol, membrane/organelle, nuclei, and cytoskeleton. Then, proteins were extracted from each fraction and digested by trypsin. The produced peptides were detected using matrix-assisted laser desorption ionisation-time-of-flight mass spectrometry (MALDI-TOF MS).

Results

We detected 33 peptide peaks whose expression was upregulated by more than 2.5-fold in GM-CSF stimulated neutrophils and identified 11 proteins out of the 33 peptides using MALDI-TOF/TOF MS analysis and protein database searches. One of the identified proteins was neutrophil gelatinase-associated lipocalin (NGAL). We confirmed that the level of NGAL in SF was significantly higher in patients with RA than in those with osteoarthritis. We next addressed possible roles of the increased NGAL in RA. We analysed proteome alteration of synoviocytes from patients with RA by treatment with NGAL in vitro. We found that, out of the detected protein spots (approximately 3,600 protein spots), the intensity of 21 protein spots increased by more than 1.5-fold and the intensity of 10 protein spots decreased by less than 1 to 1.5-fold as a result of the NGAL treatment. Among the 21 increased protein spots, we identified 9 proteins including transitional endoplasmic reticulum ATPase (TERA), cathepsin D, and transglutaminase 2 (TG2), which increased to 4.8-fold, 1.5-fold and 1.6-fold, respectively. Two-dimensional electrophoresis followed by western blot analysis confirmed the upregulation of TERA by the NGAL treatment and, moreover, the western blot analysis showed that the NGAL treatment changed the protein spots caused by post-translational modification of TERA. Furthermore, NGAL cancelled out the proliferative effects of fibroblast growth factor (FGF)-2 and epidermal growth factor (EGF) on chondrocytes from a patient with RA and proliferative effect of FGF-2 on chondrosarcoma cells.

Conclusions

Our results indicate that GM-CSF contributes to the pathogenesis of RA through upregulation of NGAL in neutrophils, followed by induction of TERA, cathepsin D and TG2 in synoviocytes. NGAL and the upregulated enzymes may therefore play an important role in RA.