Systemic BCG immunization induces persistent lung mucosal multifunctional CD4 T(EM) cells which expand following virulent mycobacterial challenge

To more closely understand the mechanisms of how BCG vaccination confers immunity would help to rationally design improved tuberculosis vaccines that are urgently required. Given the established central role of CD4 T cells in BCG induced immunity, we sought to characterise the generation of memory CD4 T cell responses to BCG vaccination and M. bovis infection in a murine challenge model. We demonstrate that a single systemic BCG vaccination induces distinct systemic and mucosal populations of T effector memory (T_EM) cells in vaccinated mice. These CD4⁺CD44^hiCD62L^loCD27⁻ T cells concomitantly produce IFN-γ and TNF-α, or IFN-γ, IL-2 and TNF-α and have a higher cytokine median fluorescence intensity MFI or ‘quality of response’ than single cytokine producing cells. These cells are maintained for long periods (>16 months) in BCG protected mice, maintaining a vaccine–specific functionality. Following virulent mycobacterial challenge, these cells underwent significant expansion in the lungs and are, therefore, strongly associated with protection against M. bovis challenge. Our data demonstrate that a persistent mucosal population of T_EM cells can be induced by parenteral immunization, a feature only previously associated with mucosal immunization routes; and that these multifunctional T_EM cells are strongly associated with protection. We propose that these cells mediate protective immunity, and that vaccines designed to increase the number of relevant antigen-specific T_EM in the lung may represent a new generation of TB vaccines.     

Zonal rate model for axial and radial flow membrane chromatography. Part I: Knowledge transfer across operating conditions and scales

The zonal rate model (ZRM) has previously been applied for analyzing the performance of axial flow membrane chromatography capsules by independently determining the impacts of flow and binding related non‐idealities on measured breakthrough curves. In the present study, the ZRM is extended to radial flow configurations, which are commonly used at larger scales. The axial flow XT5 capsule and the radial flow XT140 capsule from Pall are rigorously analyzed under binding and non‐binding conditions with bovine serum albumin (BSA) as test molecule. The binding data of this molecule is much better reproduced by the spreading model, which hypothesizes different binding orientations, than by the well‐known Langmuir model. Moreover, a revised cleaning protocol with NaCl instead of NaOH and minimizing the storage time has been identified as most critical for quantitatively reproducing the measured breakthrough curves. The internal geometry of both capsules is visualized by magnetic resonance imaging (MRI). The flow in the external hold‐up volumes of the XT140 capsule was found to be more homogeneous as in the previously studied XT5 capsule. An attempt for model‐based scale‐up was apparently impeded by irregular pleat structures in the used XT140 capsule, which might lead to local variations in the linear velocity through the membrane stack. However, the presented approach is universal and can be applied to different capsules. The ZRM is shown to potentially help save valuable material and time, as the experiments required for model calibration are much cheaper than the predicted large‐scale experiment at binding conditions. Biotechnol. Bioeng. 2013; 110: 1129–1141. © 2012 Wiley Periodicals, Inc.     

Increasing the expression of programmed death ligand 2 (PD-L2) but not 4-1BB ligand in colorectal cancer cells

Molecular Biology Reports - Immune checkpoint (ICP) molecules modulate the immune response by either inducing or preventing T cell activation. Over-expression of some ICPs on malignant cells has...     

Hydrolytic mechanism of OXA-58 enzyme, a carbapenem-hydrolyzing class D β-lactamase from Acinetobacter baumannii

Carbapenem-hydrolyzing class D β-lactamases (CHDLs) represent an emerging antibiotic resistance mechanism encountered among the most opportunistic Gram-negative bacterial pathogens. We report here the substrate kinetics and mechanistic characterization of a prominent CHDL, the OXA-58 enzyme, from Acinetobacter baumannii. OXA-58 uses a carbamylated lysine to activate the nucleophilic serine used for β-lactam hydrolysis. The deacylating water molecule approaches the acyl-enzyme species, anchored at this serine (Ser-83), from the α-face. Our data show that OXA-58 retains the catalytic machinery found in class D β-lactamases, of which OXA-10 is representative. Comparison of the homology model of OXA-58 and the recently solved crystal structures of OXA-24 and OXA-48 with the OXA-10 crystal structure suggests that these CHDLs have evolved the ability to hydrolyze imipenem, an important carbapenem in clinical use, by subtle structural changes in the active site. These changes may contribute to tighter binding of imipenem to the active site and removal of steric hindrances from the path of the deacylating water molecule.     

Genetic mapping of the <Emphasis Type="Italic">Leptosphaeria maculans</Emphasis> avirulence gene corresponding to the <Emphasis Type="Italic">LepR1</Emphasis> resistance gene of <Emphasis Type="Italic">Brassica napus</Emphasis>

AvrLepR1 of the fungal pathogen Leptosphaeria maculans is the avirulence gene that corresponds to Brassica LepR1, a plant gene controlling dominant, race-specific resistance to this pathogen. An in vitro cross between the virulent L. maculans isolate, 87-41, and the avirulent isolate, 99-56, was performed in order to map the AvrLepR1 gene. The disease reactions of the 94 of the resulting F₁ progenies were tested on the canola line ddm-12-6s-1, which carries LepR1. There were 44 avirulent progenies and 50 virulent progenies suggesting a 1:1 segregation ratio and that the avirulence of 99-56 on ddm-12-6s-1 is controlled by a single gene. Tetrad analysis also indicated a 1:1 segregation ratio. The AvrLepR1 gene was positioned on a genetic map of L. maculans relative to 259 sequence-related amplified polymorphism (SRAP) markers, two cloned avirulence genes (AvrLm1 and AvrLm4-7) and the mating type locus (MAT1). The genetic map consisted of 36 linkage groups, ranging in size from 13.1 to 163.7 cM, and spanned a total of 2,076.4 cM. The AvrLepR1 locus was mapped to linkage group 4, in the 13.1 cM interval flanked by the SRAP markers SBG49-110 and FT161-223. The AvrLm4-7 locus was also positioned on linkage group 4, close to but distinct from the AvrLepR1 locus, in the 5.4 cM interval flanked by FT161-223 and P1314-300. This work will make possible the further characterization and map-based cloning of AvrLepR1. A combination of genetic mapping and pathogenicity tests demonstrated that AvrLepR1 is different from each of the L. maculans avirulence genes that have been characterized previously.     

Serotonin regulates C. elegans fat and feeding through independent molecular mechanisms

We investigated serotonin signaling in C. elegans as a paradigm for neural regulation of energy balance and found that serotonergic regulation of fat is molecularly distinct from feeding regulation. Serotonergic feeding regulation is mediated by receptors whose functions are not required for fat regulation. Serotonergic fat regulation is dependent on a neurally expressed channel and a G protein-coupled receptor that initiate signaling cascades that ultimately promote lipid breakdown at peripheral sites of fat storage. In turn, intermediates of lipid metabolism generated in the periphery modulate feeding behavior. These findings suggest that, as in mammals, C. elegans feeding behavior is regulated by extrinsic and intrinsic cues. Moreover, obesity and thinness are not solely determined by feeding behavior. Rather, feeding behavior and fat metabolism are coordinated but independent responses of the nervous system to the perception of nutrient availability.     

Stem cells for the treatment of musculoskeletal pain

Musculoskeletal-related pain is one of the most disabling health conditions affecting more than one third of the adult population worldwide. Pain from various mechanisms and origins is currently underdiagnosed and undertreated. The complexity of molecular mechanisms correlating pain and the progression of musculoskeletal diseases is not yet fully understood. Molecular biomarkers for objective evaluation and treatment follow-up are needed as a step towards targeted treatment of pain as a symptom or as a disease. Stem cell therapy is already under investigation for the treatment of different types of musculoskeletal-related pain. Mesenchymal stem cell-based therapies are already being tested in various clinical trials that use musculoskeletal system-related pain as the primary or secondary endpoint. Genetically engineered stem cells, as well as induced pluripotent stem cells, offer promising novel perspectives for pain treatment. It is possible that a more focused approach and reassessment of therapeutic goals will contribute to the overall efficacy, as well as to the clinical acceptance of regenerative medicine therapies. This article briefly describes the principal types of musculoskeletal-related pain and reviews the stem cell-based therapies that have been specifically designed for its treatment.     

Reversible histochemical modifications of endoplasmic reticulum following arginine vasopressin stimulation of granular cells of toad bladder

The endoplasmic reticulum is generally absent from schematic representations of transport phenomena, although it shows a well-organized network in most transport epithelial cells. In order to examine the correlation between this organelle and cellular activity, bladders of Bufo marinus were studied under different experimental conditions and fixed by immersion in glutaraldehyde, followed by OsO₄ impregnation for 3 days. Normal granular and mitochondria-rich cells showed a rich cytoplasmic network of canaliculi, well-impregnated by osmium deposits. Following a 2 to 15-min stimulation (serosal bath) with arginine vasopressin, the V₂ receptor agonist dD-arginine-vasopressin or cyclic AMP (cAMP), the staining of endoplasmic reticulum in granular cells disappeared. After washing out of the hormone or the agonist, impregnation of the endoplasmic reticulum could be observed once again. Arginine vasopressin did not modify the impregnation of endoplasmic reticulum of either mitochondria-rich or basal cells. Our data indicate a correlation between the reactivity of endoplasmic reticulum to osmium, and a cAMP-dependent effect of arginine vasopressin through its V₂ receptors. Incubation of toad bladders carried out with agents interfering with cellular calcium (calcium ionophores, high or low bath calcium) or with calcium release from the endoplasmic reticulum (TMB-8, thapsigargin) suggested that an early step in the cAMP-dependent effect of arginine vasopressin must involve the release of intracellular calcium from the endoplasmic reticulum. However, calcium ATPases in this organelle do not seem to participate in the hormonal effect. The reversible loss of osmium impregnation induced by arginine vasopressin may represent protein changes in the endoplasmic reticulum accompanying a cAMP-dependent calcium release, from the organelle.     

Multiple ophthalmic anomalies and digital hypoplasia.

Multiple congenital eye and hand anomalies occurred in a young female born to normal but consanguineous parents. Both eyes were microphthalmic with severe corneal, iris lens pathology. Ultrasonography revealed multiple echos from the vitreous. The ocular findings are suggestive of retinal dysplasia. A skeletal dysplasia, presenting as distal phalangeal hypoplasia, was found in both hands. There was no history of intrauterine exposure to drugs. This appears to be a unique association of congenital malformations, without other systemic involvement. Diagnostic and genetic implications are discussed.