Wiskott-Aldrich Syndrome (WAS) and Dedicator of Cytokinesis 8- (DOCK8) Deficiency

Both Wiskott-Aldrich syndrome (WAS) and dedicator of cytokinesis 8 (DOCK8) deficiency are primary immunodeficiency diseases caused by mutations in genes that result in defective organization of the cytoskeleton in hematopoietic tissues. They share some overlapping features such as a combined immunodeficiency, eczema and a predisposition to autoimmunity and malignancy, but also have some unique features that make them relatively easy to diagnose by clinical means. Both diseases can be cured by HSCT in a large proportion of patients. In WAS it is sometimes difficult to establish an indication for HSCT due to the large variability of disease severity, while HSCT is probably indicated in all patients affected by DOCK8 deficiency. There is considerably more published HSCT experience for WAS than for DOCK8 deficiency, but many open questions remain, which will be discussed in this review

In situ three-dimensional reconstruction of mouse heart sympathetic innervation by two-photon excitation fluorescence imaging

Background Sympathetic nerve wiring in the mammalian heart has remained largely unexplored. Resolving the wiring diagram of the cardiac sympathetic network would help establish the structural underpinnings of neurocardiac coupling. New Method We used two-photon excitation fluorescence microscopy, combined with a computer-assisted 3-D tracking algorithm, to map the local sympathetic circuits in living hearts from adult transgenic mice expressing enhanced green fluorescent protein (EGFP) in peripheral adrenergic neurons. Results Quantitative co-localization analyses confirmed that the intramyocardial EGFP distribution recapitulated the anatomy of the sympathetic arbor. In the left ventricular subepicardium of the uninjured heart, the sympathetic network was composed of multiple subarbors, exhibiting variable branching and looping topology. Axonal branches did not overlap with each other within their respective parental subarbor nor with neurites of annexed subarbors. The sympathetic network in the border zone of a 2-week-old myocardial infarction was characterized by substantive rewiring, which included spatially heterogeneous loss and gain of sympathetic fibers and formation of multiple, predominately nested, axon loops of widely variable circumference and geometry. Comparison with Existing Methods In contrast to mechanical tissue sectioning methods that may involve deformation of tissue and uncertainty in registration across sections, our approach preserves continuity of structure, which allows tracing of neurites over distances, and thus enables derivation of the three-dimensional and topological morphology of cardiac sympathetic nerves. Conclusions Our assay should be of general utility to unravel the mechanisms governing sympathetic axon spacing during development and disease

Novel methods for spatial prediction of soil functions within landscapes (SP0531)

Previous studies showed that soil patterns could be predicted in agriculturally managed landscapes by modelling and extrapolating from extensive existing but related integrated datasets. Based on these results we proposed to develop and apply predictive models of the relationships between environmental data and known soil patterns to predict capacity for key soil functions within diverse landscapes for which there is little detailed underpinning soil information available. Objectives were: To develop a high-level framework in which the non-specialist user-community could explore questions. To generate digital soil maps for three selected catchments at a target resolution of 1:50000 to provide the base information for soil function prediction. To use a modelling approach to predict the performance of key soil functions in catchments undergoing change but where only sparse or low resolution soil survey data are available. To use a modelling approach to assess the impact of different management scenarios and/or environmental conditions on the delivery of multiple soil functions within a catchment. To create a detailed outline of the requirements for ground-truthing to test the predicted model outputs at a catchment scale. To contribute to the development of a high-level framework for decision makers

KERUING and APITONG: A Timely Review of the Perfect ChoiceWood Species for Crossarms and Brace Arms, including Data on Copper Naphthenate Treatments

Apitong or the wood species more commonly known as Keruing is an important tropical hardwood known for its high strength to weight ratio and its reasonable durability and preservative treatability. This wood has been exported from managed permanent forest estates in Malaysia into the United States for over 4 decades for its widespread use in wooden utility industry brace arms and crossarms, yet currently it is not listed as an approved species by either ANSI or ASTM. Because it has not had any significant failures in its use patterns now for almost four decades, members of the AWPA Utility Users Task Group have asked for a comprehensive and Critical review of the species and its durability or treatability properties, since they have been using it and specifying it in their systems under product acception for years. This paper is meant to be a comprehensive review of both public domain data and privately sponsored studies to yield additional product information, for those using, specifying and treating this tropical, sustainable species from Malaysia

Stable isotope metabolomics of pulmonary artery smooth muscle and endothelial cells in pulmonary hypertension and with TGF-beta treatment.

Altered metabolism in pulmonary artery smooth muscle cells (PASMCs) and endothelial cells (PAECs) contributes to the pathology of pulmonary hypertension (PH), but changes in substrate uptake and how substrates are utilized have not been fully characterized. We hypothesized stable isotope metabolomics would identify increased glucose, glutamine and fatty acid uptake and utilization in human PASMCs and PAECs from PH versus control specimens, and that TGF-β treatment would phenocopy these metabolic changes. We used 13C-labeled glucose, glutamine or a long-chain fatty acid mixture added to cell culture media, and mass spectrometry-based metabolomics to detect and quantify 13C-labeled metabolites. We found PH PASMCs had increased glucose uptake and utilization by glycolysis and the pentose shunt, but no changes in glutamine or fatty acid uptake or utilization. Diseased PAECs had increased proximate glycolysis pathway intermediates, less pentose shunt flux, increased anaplerosis from glutamine, and decreased fatty acid β-oxidation. TGF-β treatment increased glycolysis in PASMCs, but did not recapitulate the PAEC disease phenotype. In TGF-β-treated PASMCs, glucose, glutamine and fatty acids all contributed carbons to the TCA cycle. In conclusion, PASMCs and PAECs collected from PH subjects have significant changes in metabolite uptake and utilization, partially recapitulated by TGF-β treatment

Regulation of vascular smooth muscle cell expression and function of matrix metalloproteinases is mediated by estrogen and progesterone exposure

ObjectivePostmenopausal women receiving hormone replacement therapy (HRT) have been reported to have more adverse outcomes after vascular reconstructions, including increased intimal hyperplasia development and bypass graft failure. HRT may be affecting the pathway contributing to intimal hyperplasia. An important component of this pathway involves matrix metalloproteinases (MMPs), implicated in vascular remodeling due to their ability to degrade components of the extracellular matrix. We hypothesize that estrogen (Est) and progesterone (Prog) upregulate the MMP pathway in vascular smooth muscle cells (VSMCs) thereby increasing MMP activity and function.Methods and ResultsVSMCs were incubated with Est (5 ng/mL), Prog (50 ng/mL), Est + Prog combination (Est/Prog), and/or doxycycline (40 μg/mL; Doxy). Using reverse transcriptase polymerase chain reaction (RT-PCR) analysis we have previously shown membrane type 1-MMP (MT1-MMP) messenger ribonucleic acid (mRNA) levels are significantly increased by Est. Here, Western blot analyses indicated MT1-MMP and MMP-2 protein levels, not tissue inhibitor of MMP-2 (TIMP-2), were increased in response to Est and Est/Prog (P < .05 vs control). In-gel zymography revealed that Est and Est/Prog resulted in increased MMP-2 activity (hormone groups, P < .05 vs control) with no significant difference among the hormone groups. VSMC migration was increased by 45 ± 14% in response to Est (P < .05 vs control), as measured using a modified Boyden chamber assay. Doxycycline significantly inhibited basal and Est/Prog-stimulated increases in MMP-2 activity (P < .05 vs control; P < .05 vs hormone groups), and partially blocked basal and hormonally stimulated migration (P < .05 vs control and Est).ConclusionEstrogen and progesterone affects the MMP pathway by increasing MMP-2 enzymatic activity, possibly via the upregulation of MT1-MMP expression without a corresponding increase in TIMP expression. This increased collagenase activity increases VSMC motility and their ability to migrate through a collagen type IV lattice. Est/Prog upregulation of MT1-MMP may contribute to the adverse effect of HRT on vascular interventions.Clinical RelevancePostmenopausal women receiving HRT have more adverse outcomes after vascular reconstructions, including intimal hyperplasia, restenosis, and decreased graft patency. MMPs play a major role in vascular remodeling due to their degradation of components of the basement membrane separating vascular cell layers. Specifically, MMP-2 has a strong affinity for collagen type IV degradation, and MT1-MMP is a transmembrane protein known to activate MMP-2 by proteolytic cleavage. Here we provide strong evidence for MT1-MMP's role in increased MMP-2 activity and increased cellular migration in VSMCs exposed to estrogen and progesterone. Manipulations of the MMP pathway specifically targeting MT1-MMP expression at the time of vascular interventions may improve outcomes in females receiving HRT

Trading in your spindles for blebs: the amoeboid tumor cell phenotype in prostate cancer

Prostate cancer (PCa) remains a principal cause of mortality in developed countries. Because no clinical interventions overcome resistance to androgen ablation therapy, management of castration resistance and metastatic disease remains largely untreatable. Metastasis is a multistep process in which tumor cells lose cell-cell contacts, egress from the primary tumor, intravasate, survive shear stress within the vasculature and extravasate into tissues to colonize ectopic sites. Tumor cells reestablish migratory behaviors employed during nonneoplastic processes such as embryonic development, leukocyte trafficking and wound healing. While mesenchymal motility is an established paradigm of dissemination, an alternate, ‘amoeboid’ phenotype is increasingly appreciated as relevant to human cancer. Here we discuss characteristics and pathways underlying the phenotype, and highlight our findings that the cytoskeletal regulator DIAPH3 governs the mesenchymal-amoeboid transition. We also describe our identification of a new class of tumor-derived microvesicles, large oncosomes, produced by amoeboid cells and with potential clinical utility in prostate and other cancers

Contemporary outcomes of vertebral artery injury

ObjectiveVertebral artery injury (VAI) associated with cervical trauma is being increasingly recognized with more aggressive screening. Disparate results from previous literature have led to uncertainty of the significance, natural history, and optimal therapy for VAI.MethodsTo understand the natural history and treatment outcomes from our experience, we performed a retrospective, single-center review from a level I trauma center for the previous 10 years of all VAI. Injuries were identified from search of an administrative trauma database, a resident-run working database, and all radiology dictations for the same period. All VAI were classified according to segmental involvement, Denver grading scale, and laterality. Analysis of associated injuries, demographics, neurologic outcome, mortality, length of stay, treatment plan, and follow-up imaging was also performed.ResultsFifty-one patients with VAI were identified from 2001 to 2011 from a total of 36,942 trauma admissions (0.13% incidence). Associated injuries were significant with an average New Injury Severity Score of 29.6. Penetrating trauma occurred in 14%. Cervical spine fracture was present in 88% with VAI. Diagnosis was obtained with computed tomographic angiography (CTA) in 95%. Screening was prompted by injury pattern or high-risk mechanism in all cases. Injuries classified according to the Denver grading scale were grade I = 24%, grade II = 35%, grade III = 4%, grade IV = 35%, and grade V = 2%. Distribution across segments included V1 = 18%, V2 = 67%, V3 = 31%, and V4 = 6%. Only one posterior circulation stroke was attributable to VAI. Overall mortality was 8%, with each mortality being associated with significant other organ injuries. Treatment rendered for VAI was antiplatelet therapy (50%), observation (31%), warfarin (17%), and stent (2%). There were no significant differences between treatment groups on any variable with the exception of body mass index (P = .047). Follow-up was obtained for 13% (n = 6) of survivors. The CTA demonstrated injury stability in four patients and resolution in two patients. Accuracy of the administrative trauma database was 53% compared with 96% for the resident-run working database.ConclusionsNeurologic sequelae attributable to VAI were rare. Grade of VAI or vertebral artery segment did not correlate with morbidity. We did not observe any differences in short-term outcomes between systemic anticoagulation and antiplatelet therapy. Of those patients seen at follow-up, injury resolution or stability was documented by CTA. A conservative approach with either observation or antithrombotic therapy is suggested. If the natural history of VAI includes a very low stroke rate, then therapies with a lower therapeutic index, such as systemic anticoagulation, in the severely injured trauma patient are not supported. Our search strategy urges awareness of the limitations of administrative databases for retrospective vascular study

Gender Differences in Bed Rest: Preliminary Analysis of Vascular Function

Orthostatic intolerance is a recognized consequence of spaceflight. Numerous studies have shown that women are more susceptible to orthostatic intolerance following spaceflight as well as bed rest, the most commonly used ground-based analog for spaceflight. One of the possible mechanisms proposed to account for this is a difference in vascular responsiveness between genders. We hypothesized that women and men would have differing vascular responses to 90 days of 6-degree head down tilt bed rest. Additionally, we hypothesized that vessels in the upper and lower body would respond differently, as has been shown in the animal literature. Thirteen subjects were placed in bedrest for 90 days (8 men, 5 women) at the Flight Analogs Unit, UTMB. Direct arterial and venous measurements were made with ultrasound to evaluate changes in vascular structure and function. Arterial function was assessed, in the arm and leg, during a reactive hyperemia protocol and during sublingual nitroglycerin administration to gauge the contributions of endothelial dependent and independent dilator function respectively. Venous function was assessed in dorsal hand and foot veins during the administration of pharmaceuticals to assess constrictor and dilator function. Both gender and day effects are seen in arterial dilator function to reactive hyperemia, but none are seen with nitroglycerin. There are also differences in the wall thickness in the arm vs the leg during bed rest, which return toward pre-bed rest levels by day 90. More subjects are required, especially females as there is not sufficient power to properly analyze venous function. Day 90 data are most underpowered

Vascular smooth muscle cell apoptosis in aneurysmal, occlusive, and normal human aortas

AbstractPurpose: Apoptosis is a physiologic mechanism of cell death that regulates mass and architecture in many tissues. Apoptosis has been described as a feature in human vascular atherosclerosis and large vessel structural integrity. We examined the extent of vascular smooth muscle cell (VSMC) apoptosis in aneurysmal, occlusive, and normal human aortic tissue. Methods: Tissue samples of aneurysmal, occlusive, and normal human infrarenal aorta were evaluated. DNA fragmentation detection methods, immunohistochemistry, and DNA electrophoresis determined VSMC density, VSMC apoptosis, and apoptosis markers. Apoptotic cells and VSMC nuclei were counted with the use of computer-generated image analysis. Aortic subtypes were compared statistically by analysis of variance. Results: Seventeen aneurysmal, ten occlusive, and five normal human aortas were evaluated. By α1-actin immunostaining, VSMC density was least in aneurysmal aortas (271.8 ± 13.5 cells/high-power field [HPF]) compared with occlusive aorta (278.2 ± 39.4 cells/HPF) and normal aortas (291.0 ± 25.4 cells/HPF; P = not significant). Presence of apoptotic VSMCs was demonstrated by terminal deoxynucleotidyl transferase fragment end labeling and propidium iodide nuclear staining. VSMC apoptosis was greatest within aneurysmal aortas with 11.7 ± 1.5 cells/HPF compared with occlusive aortas with 3.3 ± 0.8 cells/HPF (P <.05) and normal aortas with 3.75 ± 4.6 cells/HPF (P <.05). Significant differences in apoptosis markers, p53 or bcl-2, could not be demonstrated by immunohistochemistry or DNA electrophoresis in aortic subtypes. Conclusion: Apoptosis of VSMCs is increased and VSMC density is decreased within the medial layer of aneurysmal aortic tissue. Structural degeneration of aortic tissue at the cellular level contributes to aneurysmal formation. (J Vasc Surg 2000;31:567-576.