Conserved Extracellular Cysteines Differentially Regulate the Potentiation Produced by Zn2+ in Rat P2X4 Receptors

One feature of the amino acid sequence of P2X receptors identified from mammalian species, Xenopus laevis and zebrafish is the conservation of ten cysteines in the extracellular loop. Little information is available about the role of these conserved ectodomain cysteines in the function of P2X receptors. Here, we investigated the possibility that ten conserved cysteine residues in the extracellular loop of the rat P2X4 receptor may regulate zinc potentiation of the receptor using a series of individual cysteine to alanine point mutations and functional characterization of recombinant receptors expressed in Xenopus oocytes. For the C116A, C132A, C159A, C165A, C217A and C227A mutants, 10 µM zinc did not significantly affect the current activated by an EC40 concentration of ATP. By contrast, 5 µM zinc shifted the ATP concentration-response curve to the right in a parallel manner for both the C261A and C270A mutants and the magnitudes of those shifts were similar to that of the wildtype receptor. Interestingly, for the C126A and C149A mutants, 5 µM zinc potentiated ATP-activated current, but increased the maximal response to ATP by 90% and 81% respectively, without significantly changing the EC50 value of ATP. Thus, these results suggest that cysteines and disulfide bonds between cysteines are differentially involved in the potentiation of the rat P2X4 receptor by zinc

Role of TRPM8 in dorsal root ganglion in nerve injury-induced chronic pain

<p>Abstract</p> <p>Background</p> <p>Chronic neuropathic pain is an intractable pain with few effective treatments. Moderate cold stimulation can relieve pain, and this may be a novel train of thought for exploring new methods of analgesia. Transient receptor potential melastatin 8 (TRPM8) ion channel has been proposed to be an important molecular sensor for cold. Here we investigate the role of TRPM8 in the mechanism of chronic neuropathic pain using a rat model of chronic constriction injury (CCI) to the sciatic nerve.</p> <p>Results</p> <p>Mechanical allodynia, cold and thermal hyperalgesia of CCI rats began on the 4th day following surgery and maintained at the peak during the period from the 10th to 14th day after operation. The level of TRPM8 protein in L5 dorsal root ganglion (DRG) ipsilateral to nerve injury was significantly increased on the 4th day after CCI, and reached the peak on the 10th day, and remained elevated on the 14th day following CCI. This time course of the alteration of TRPM8 expression was consistent with that of CCI-induced hyperalgesic response of the operated hind paw. Besides, activation of cold receptor TRPM8 of CCI rats by intrathecal application of menthol resulted in the inhibition of mechanical allodynia and thermal hyperalgesia and the enhancement of cold hyperalgesia. In contrast, downregulation of TRPM8 protein in ipsilateral L5 DRG of CCI rats by intrathecal TRPM8 antisense oligonucleotide attenuated cold hyperalgesia, but it had no effect on CCI-induced mechanical allodynia and thermal hyperalgesia.</p> <p>Conclusions</p> <p>TRPM8 may play different roles in mechanical allodynia, cold and thermal hyperalgesia that develop after nerve injury, and it is a very promising research direction for the development of new therapies for chronic neuroapthic pain.</p

ProGMap: an integrated annotation resource for protein orthology

Current protein sequence databases employ different classification schemes that often provide conflicting annotations, especially for poorly characterized proteins. ProGMap (Protein Group Mappings, http://www.bioinformatics.nl/progmap) is a web-tool designed to help researchers and database annotators to assess the coherence of protein groups defined in various databases and thereby facilitate the annotation of newly sequenced proteins. ProGMap is based on a non-redundant dataset of over 6.6 million protein sequences which is mapped to 240 000 protein group descriptions collected from UniProt, RefSeq, Ensembl, COG, KOG, OrthoMCL-DB, HomoloGene, TRIBES and PIRSF. ProGMap combines the underlying classification schemes via a network of links constructed by a fast and fully automated mapping approach originally developed for document classification. The web interface enables queries to be made using sequence identifiers, gene symbols, protein functions or amino acid and nucleotide sequences. For the latter query type BLAST similarity search and QuickMatch identity search services have been incorporated, for finding sequences similar (or identical) to a query sequence. ProGMap is meant to help users of high throughput methodologies who deal with partially annotated genomic data

Decrease in CD4+CD25+FoxP3+ Treg cells after pulmonary resection in the treatment of cavity multidrug-resistant tuberculosis

SummaryObjectivesImmune regulatory mechanisms may limit the immunopathologic condition of infection with Mycobacterium tuberculosis and suppress cellular immune responses in the host. We investigated the CD4+CD25+FoxP3+ circulating regulatory T cells (Treg) in patients with cavity multidrug-resistant tuberculosis (MDR-TB) before and after surgery.MethodsWe compared the proportion of Treg cells in 13 patients with cavity MDR-TB pre- and postoperatively and in 10 healthy control subjects by flow cytometry using three specific markers in peripheral blood lymphocytes: cell-surface CD4 and CD25 expression and intracellular FoxP3 expression.ResultsThe proportion of CD4+CD25high and CD4+CD25+FoxP3+ Treg was significantly higher in patients with cavity MDR-TB and at 1-month postoperatively than in healthy controls (p<0.001). The proportion of CD4+ and CD4+CD25− cells was significantly lower in patients with cavity MDR-TB than in controls (p<0.001). Pre- and postoperative proportions of CD4+CD25high and CD4+CD25+FoxP3+ Treg cells showed a positive correlation (r=0.878, p<0.001).ConclusionCirculating Treg cells are increased in proportion in patients with cavity MDR-TB and decreased after surgery. Infection with M. tuberculosis may induce Treg cell-surface molecular changes with increased numbers of cells

Clinical observation of intense pulsed light therapy for meibomian gland dysfunction

AIM: To investigate the effect of intense pulsed light on the meibomian gland dysfunction(MGD), and to observe whether the intense pulsed light can improve the symptoms and objective indexes of the patients with meibomian gland dysfunction. METHODS: A retrospective noncomparative interventional case series was conducted. Totally 21 MGD patients(42 eyes)were selected from November 2016 to February 2017 in Renmin Hospital of Wuhan University, the results of the following tests is recorded: OSDI score, corneal fluorescein staining, tear break-up time, ocular surface analyzer, scanning confocal microscopy, all patients respectively were received intense pulsed light treatment 3 times at 3wk intervals, and followed up after 1mo. Paired sample t test was used to analyze the difference in outcome. RESULTS: In the 21 cases(42 eyes), the OSDI score decreased, the mean value before treatment was 30.18±4.07, and the mean value after treatment was 24.87±4.32. The first tear film break-up time(first, BUT)increased, the mean value before treatment was 5.37±0.82s, the mean value after treatment was 7.12±0.74s. The mean value of meibomian gland secretion scores before the treatment was 1.57±0.52,the mean value after treatment was 1.22±0.52. The mean value of corneal fluorescence staining before treatment was 0.82±0.41, and the mean value after treatment was 0.51±0.53, the difference was statistically significant(t=11.2, 2.68, 3.31, 2.78, 2.61; PCONCLUSION: Intense pulsed light can effectively improve the subjective symptoms of patients with MGD and alleviate the obstruction of meibomian glands. It is an important method for MGD treatment