Defective erythroid maturation in gelsolin mutant mice

BackgroundDuring late differentiation, erythroid cells undergo profound changes involving actin filament remodeling. One of the proteins controlling actin dynamics is gelsolin, a calcium-activated actin filament severing and capping protein. Gelsolin-null (Gsn^−/−) mice generated in a C57BL/6 background are viable and fertile.¹ResultsIn the context of a BALB/c background, the Gsn^−/− mutation causes embryonic death. Gsn^−/− embryos show defective erythroid maturation with persistence of circulating nucleated cells. The few Gsn^−/− mice reaching adulthood fail to recover from phenylhydrazine-induced acute anemia, revealing an impaired response to stress erythropoiesis. In in vitro differentiation assays, E13.5 fetal liver Gsn^−/− cells failed to undergo terminal maturation, a defect partially rescued by Cytochalasin D, and mimicked by administration of Jasplakinolide to the wild-type control samples.ConclusionsIn BALB/c mice, gelsolin deficiency alters the equilibrium between erythrocyte actin polymerization and depolymerization, causing impaired terminal maturation. We suggest a non-redundant role for gelsolin in terminal erythroid differentiation, possibly contributing to the Gsn^−/− mice lethality observed in mid-gestation.     

Warfarin control in patients transitioning to warfarin after non-vitamin K oral anticoagulant (NOAC) therapy

Warfarin has long been the most widely prescribed oral anticoagulant. Introduction of non-vitamin K oral anticoagulants (NOACs) has provided anticoagulant options but also presented the potential challenge of transitioning between agents. Changes from NOACs to warfarin are particularly problematic with delays to therapeutic effect and limited real-world data regarding the impact on warfarin control. The aim of this study was to investigate the frequency of switching anticoagulants and the effect on warfarin control. Retrospective data was collected for patients at a warfarin program in Queensland Australia who had exited the program for NOACs plus those who had reverted to warfarin. Data included documented reasons for change and International Normalised Ratio (INR) results with time in therapeutic range (TTR) calculated as a measure of warfarin control. Over 5 years, a total of 3036 patients ceased warfarin to commence a NOAC but 142 (4.7%) reverted to warfarin. Majority of patients (60.6%) reverted to warfarin within 6 months of trialling NOACs with a median of 6 days to therapeutic INR. There was no significant difference in warfarin control before changing to NOACs and after reverting to warfarin (mean TTR 75%) but significantly more frequent testing and lower doses were required to achieve this control. Transitions from warfarin to NOACs results in almost a week to therapeutic effect and warfarin therapy may be further complicated by a need for increased frequency of testing. Further studies are required to refine transition strategies particularly from warfarin to NOAC and minimise potential risks to patients.     

Phagocytosis and chemotaxis of macrophages from normal and diabetic mice

Summary Experiments were designed to study the chemotaxis and phagocytic functions of mouse peritoneal exudate macrophages from normal and chronic diabetic mice. There was no difference in the total number of white blood cells (WBC's), or the total number of any one type of leukocyte (i.e., macrophage, neutrophil) migrating into the casein-induced peritoneal exudate of normal and diabetic mice.In vitro phagocytosis of heat-killedCandida albicans by peritoneal induced macrophages was also similar in both of these groups of mice. No correlation between the percent phagocytosis and blood glucose level within each group was observed. Therefore, the attenuated humoral and cellular immune response which has been previously observed in diabetes cannot be attributed to defects in chemotaxis or phagocytosis as studied using mouse peritoneal macrophages.     

Protection against an intranasal challenge by vaccines formulated with native and recombinant preparations of the Chlamydia trachomatis major outer membrane protein

To compare the ability of a native and a recombinant preparation of the major outer membrane protein of Chlamydia trachomatis mouse pneumonitis (MoPn; Ct-nMOMP and Ct-rMOMP) to protect against an intranasal (i.n.) challenge, BALB/c mice were vaccinated by the intramuscular (i.m.) and subcutaneous (s.c.) routes using CpG-1826 and Montanide ISA 720 as adjuvants. Animals inoculated i.n. with live elementary bodies (EB) of Chlamydia served as a positive control. Negative control groups were immunized with either Neisseria gonorrhoeae recombinant porin B (Ng-rPorB) or with minimal essential medium (MEM-0). Mice immunized with Ct-rMOMP, Ct-nMOMP and EB developed a strong immune response as shown by high levels of Chlamydia specific antibodies in serum and a strong T-cell lymphoproliferative response. Following the i.n. challenge with 10⁴ inclusion forming units (IFU) of C. trachomatis, mice immunized with Ct-nMOMP or Ct-rMOMP lost significantly less weight than the negative control animals immunized with Ng-rPorB or MEM-0 (P < 0.05). However, mice vaccinated with the Ct-nMOMP lost less weight than those immunized with the Ct-rMOMP (P < 0.05). Mice were euthanized at 10 days following the challenge, their lungs weighed and the number of IFU of Chlamydia determined. Based on the lung weight and number of IFU recovered, significant protection was observed in the groups of mice immunized with both Ct-nMOMP and the Ct-rMOMP (P < 0.05). Nevertheless, significantly better protection was achieved with the Ct-nMOMP in comparison with the Ct-rMOMP (P < 0.05). In conclusion, vaccination with a preparation of the nMOMP elicited a more robust protection than immunization with rMOMP, suggesting that the conformational structure of MOMP is critical for inducing strong protection.     

Serum levels of insulin-like growth factor-1 and insulin-like growth factor-1 binding proteins after radical prostatectomy

PURPOSE: Elevated serum levels of insulin-like growth factor-1 (IGF-1) have been consistently shown to be a risk factor for prostate cancer. Alterations in serum IGF-1 binding proteins 1 to 3 have also been associated with prostate cancer risk. A potentially important complication in these studies is that prostate tissue, perhaps especially malignant prostate tissue, may secrete IGF-1 and its binding proteins into serum. In fact, it is possible that altered levels of these proteins observed in subjects at risk for prostate cancer are the result of prostate cancer rather than related to its cause. MATERIALS AND METHODS: The contribution of prostate cancer to serum levels of IGF-1 and IGF-1 binding proteins was determined by analyzing serum samples from 86 patients with prostate cancer 2 weeks before and 8 weeks after radical prostatectomy. Preoperative and postoperative values for IGF-1 and its 3 major binding proteins were analyzed using univariate and multivariate analysis models. RESULTS: On univariate analysis significant increases and not decreases in IGF-1, IGF binding protein-1 and 3 were observed after prostatectomy. On multivariate analysis a significant post-prostatectomy increase was observed for IGF-1 binding proteins 1 and 3 but the increase in IGF-1 was not significant. CONCLUSIONS: Increased levels of IGF-1 and IGF-1 binding proteins were unexpected after prostatectomy. This result makes it extremely unlikely that secretion from the prostate, even if it contains cancer, affects serum levels of these proteins. The implication of these findings is that endocrine production of IGF-1 is a factor in prostate cancer risk. Therefore, strategies to lower serum IGF-1 may be potentially useful.     

Synthesis and characterization of a new mebendazole salt: mebendazole hydrochloride

Mebendazole hydrochloride [(5-benzoyl-1H-benzimidazole-2-yl)-carbamic acid methyl ester hydrochloride, MBZ.HCl], a new stable salt of mebendazole (MBZ), has been synthesized and characterized. It can easily be obtained from recrystallization of forms A, B, or C of MBZ in diverse solvents with the addition of hydrochloric acid solution. Crystallographic data reveals that the particular conformation adopted by the carbamic group contributes to the stability of the network. The crystal packing is stabilized by the presence of three N-H...Cl intermolecular interactions that form chains along the b axis. The XRD analyses of the three crystalline habits found in the crystallization process (square-based pyramids, pseudohexagonal plates, and prismatic) show equivalent diffraction patterns. The vibrational behavior is consistent with crystal structure. The most important functional groups show shifts to lower or higher frequencies in relation to the MBZ polymorphs. The thermal study on MBZ.HCl indicates that the compound is stable up to 160 degrees C approximately. Decomposition occurs in four steps. In the first step the HCl group is eliminated, and after that the remaining MBZ polymorph A decomposes in three steps, as happens with polymorphs B and C. (     

Hand-operated and rotary ProTaper instruments: a comparison of working time and number of rotations in simulated root canals

The aim of this study was to compare the effective shaping time and number of rotations required by an endodontist working with hand and rotary ProTaper instruments to completely shape simulated root canals. Eighty Endo Training Blocks (curved canal shape) were used. Manual preflaring was performed with K-Flexofiles #08-10-12-15-17 and #20 Nitiflex at a working length of 18 mm. Specimens were then randomly assigned to 2 different groups (n = 40); group 1 was shaped by using hand ProTaper and group 2 with ProTaper rotary. The number of rotations made in the canal and the effective time required to achieve complete canal shaping were recorded for each instrument. Differences between groups were analyzed with the nonparametric Mann-Whitney U test (P < .05). Hand ProTaper required significantly fewer rotations (P < .001) than rotary ProTaper, whereas the effective working time to fully shape the simulated canal was significantly higher (P < .001) with hand ProTaper.     

Inclusion Fluorescent-Antibody Test as a Screening Assay for Detection of Antibodies to Chlamydia pneumoniae

A study was conducted to determine the ability of the inclusion immunofluorescence assay (inclusion IFA) to act as a screening test to detect samples with antibodies to Chlamydia pneumoniae; microimmunofluorescence (MIF) was used as the “gold standard.” In addition, the inclusion IFA was compared using HEp-2 cells infected with either C. pneumoniae CM-1 or Chlamydia trachomatis serovar E. A total of 331 serum samples representing a range of MIF titers were evaluated. The sensitivities of the inclusion IFA for detecting samples with C. pneumoniae MIF titers of ≥16 were 96.9 and 74.8% with C. pneumoniae- and C. trachomatis-infected cells, respectively. For samples with an elevated C. pneumoniae MIF titer of ≥512, the sensitivities of the C. pneumoniae- and C. trachomatis-based inclusion IFA were 97.0 and 8.8%, respectively. These results suggest that the inclusion IFA is not a genus-specific test, as evidenced by the failure of the C. trachomatis-infected cells to detect a significant number of samples with C. pneumoniae antibodies. Samples that had elevated C. pneumoniae inclusion IFA and MIF titers but that were found negative (titer, <16) by the C. trachomatis inclusion IFA were further tested by an in vitro neutralization assay for functional antibodies that might not have been detected by the serological assays. The in vitro neutralization results corroborated the serological results in that all seven sera tested had a neutralization titer for C. pneumoniae (range, 20 to 225), while all but one failed to have any effect on the infectivity of C. trachomatis serovar E. While the C. pneumoniae inclusion IFA had a high sensitivity for detecting chlamydial antibodies, depending on whether it was used as a screening test for detecting samples with low (≥16) or elevated (≥512) MIF titers, its specificity ranged from 53.4 to 77.1%. In conclusion, the inclusion IFA with C. pneumoniae-infected cells was best suited as a sensitive screening test for identifying specimens with elevated MIF titers (those associated with a possible acute infection with C. pneumoniae).