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Autologous cancellous bone graft is the gold standard in large bone defect repair. However, studies using autologous bone grafting in rats are rare. To determine the feasibility of autologous cancellous bone graft harvest from different anatomical donor sites (humerus, ilium, femur, tibia, and tail vertebrae) in rats and compare their suitability as donor sites, a total of 13 freshly euthanized rats were used to describe the surgical technique, determine the cancellous bone volume and microstructure, and compare the cancellous bone collected quantitatively and qualitatively. It was feasible to harvest cancellous bone grafts from all five anatomical sites with the humerus and tail being more surgically challenging. The microstructural analysis using micro‐computed tomography showed a significantly lower bone volume fraction, bone mineral density, and trabecular thickness of the humerus and iliac crest compared to the femur, tibia, and tail vertebrae. The harvested weight and volume did not differ between the donor sites. All donor sites apart from the femur yielded primary osteogenic cells confirmed by the presence of alkaline phosphatase and Alizarin Red S stain. Bone samples from the iliac crest showed the most consistent outgrowth of osteoprogenitor cells. In conclusion, the tibia and iliac crest may be the most favorable donor sites considering the surgical approach. However, due to the differences in microstructure of the cancellous bone and the consistency of outgrowth of osteoprogenitor cells, the donor sites may have different healing properties, that need further investigation in an in vivo study. [ABSTRACT FROM AUTHOR]

Bone tissue engineering is a rapidly developing field with potential for the regeneration of craniomaxillofacial (CMF) bones, with 3D printing being a suitable fabrication tool for patient‐specific implants. The CMF region includes a variety of different bones with distinct functions. The clinical implementation of tissue engineering concepts is currently poor, likely due to multiple reasons including the complexity of the CMF anatomy and biology, and the limited relevance of the currently used preclinical models. The 'recapitulation of a human disease' is a core requisite of preclinical animal models, but this aspect is often neglected, with a vast majority of studies failing to identify the specific clinical indication they are targeting and/or the rationale for choosing one animal model over another. Currently, there are no suitable guidelines that propose the most appropriate animal model to address a specific CMF pathology and no standards are established to test the efficacy of biomaterials or tissue engineered constructs in the CMF field. This review reports the current clinical scenario of CMF reconstruction, then discusses the numerous limitations of currently used preclinical animal models employed for validating 3D‐printed tissue engineered constructs and the need to reduce animal work that does not address a specific clinical question. We will highlight critical research aspects to consider, to pave a clinically driven path for the development of new tissue engineered materials for CMF reconstruction. [ABSTRACT FROM AUTHOR]

Seasonal forecasting of tropical cyclones is a topic of considerable interest to the public, government and private sectors. To improve understanding of the dynamics controlling the predictability of tropical cyclone (TC) activity, and improve the accuracy of forecasts, multiple studies have related TC activity to empirically‐defined indices including the El Niño‐Southern Oscillation, the Atlantic Multidecadal Oscillation, and the North Atlantic Oscillation. These indices were not developed to forecast TC activity but rather summarize other aspects of atmosphere‐ocean variability. In this study we use a statistical approach, based on redundancy analysis, to define two indices related to overall activity and steering of TCs. We focus on North Atlantic TCs that reached tropical storm strength (≥34 kt) between August and October 1948–2016. TC occurrences are binned using an equal area grid that covers the North Atlantic. The redundancy indices are linear combinations of mean sea level pressure for the same season. Cross validation is used to guard against over fitting in the definition of the indices. This approach provides two physically interpretable redundancy indices related to North Atlantic TC activity. The leading redundancy index is used to successfully reconstruct the total number of TCs and the accumulated cyclone energy, over the extended period 1878–2014 using seasonal mean sea level pressures from the National Centers for Environmental Prediction 20th‐Century Reanalysis version 2c. Extensions of the approach for seasonal forecasting are discussed. Key Points: Relationship between North Atlantic tropical cyclone activity and concurrent mean sea level pressure is studied with redundancy analysisPrimary redundancy index is physically interpreted in terms of overall tropical cyclone activity with ∼50% variance explained from 1948 to 2016Secondary redundancy index is physically interpreted in terms of steering of storm tracks toward the coast of North America [ABSTRACT FROM AUTHOR]

A significant proportion of orthopedic devices are implanted in osteoporotic patients, but it is currently unclear how estrogen deficiency and/or exposure to antiresorptive bisphosphonates (BPs) influence orthopedic device‐related infection (ODRI), or response to therapy. The aim of this study is to characterize the bone changes resulting from Staphylococcus epidermidis infection in a rodent ODRI model and to determine if ovariectomy (OVX) or BP treatment influences the infection or the success of antibiotic therapy. A sterile or S. epidermidis‐contaminated screw was implanted into the proximal tibia of skeletally mature female Wistar rats (n = 6–9 per group). Bone changes were monitored over 28 days using in vivo micro‐computed tomography scanning. OVX was performed 12 weeks before screw implantation. The BP zoledronic acid (ZOL) was administered 4 days before screw insertion. A combination antibiotic regimen (rifampin plus cefazolin) was administered from Days 7–21. In skeletally healthy animals, S. epidermidis induced marked changes in bone, with peak osteolysis occurring at Day 9 and woven bone deposition and periosteal mineralization from Day 14 onwards. Antibiotic therapy cleared the infection in the majority of animals (2/9 infected) but did not affect bone responses. OVX did not affect the pattern of infection‐induced changes in bone, nor bacterial load, but reduced antibiotic efficacy (5/9 infected). ZOL treatment did not protect from osteolysis in OVX animals, or further affect antibiotic efficacy (5/9 infected) but did significantly increase the bacterial load. This study suggests that both BPs and OVX can influence host responses to bone infections involving S. epidermidis. [ABSTRACT FROM AUTHOR]

Bone is a hard yet dynamic tissue with remarkable healing capacities. Research to date has greatly advanced the understanding of how bone heals and has led to marked success in the treatment of bone injuries. Nevertheless, the effective treatment of nonunions and large bone defects continues to present a challenge for orthopedic surgeons. Biomaterials provide researchers with a powerful instrument to potentially guide effective bone tissue regeneration in challenging healing environments. However, the most appropriate biomaterial for bone tissue engineering continues to be an area of intense debate. Indeed, the mechanical properties of real bone can be reproduced in vitro but the development of a functional bone substitute goes beyond the sole mechanical properties. The faithful reproduction of bone as a functional organ requires the combination of different cell types and temporal regulation of the molecular signaling involved during the different stages of bone formation and regeneration. This is not at all a trivial task. Herein, critical aspects of bone healing/regeneration including mechanical loading, inflammation, vascularization, and innervation are described. The success and the challenges behind the development of biomaterials, and the technologies used to functionalize them, in order to support the underlying cellular mechanisms are also highlighted. [ABSTRACT FROM AUTHOR]

Complex composite materials are carefully engineered on the microscale in order to deliver a specific macro-level performance. Understanding the chemical composition on the microscale is therefore crucial to developing next generation composites and to quality monitoring of current generation composites. Scanning Electron Microscopes equipped with Energy Dispersive Spectroscopy x-ray analysers are well-positioned for this type of micro-scale analysis. The results, however, are qualitative and prone to analytical user bias. A new technique is presented here that generates quantitative elemental phase maps with no user bias and, in fact, no user input except for the raw data. This technique is used here to analyse and compare the underlying chemistries of a semi-metallic and a ceramic brake pad and the macro level performance of the two brake pads. [ABSTRACT FROM AUTHOR]

Along the Atlantic coast of Nova Scotia sea urchin disease outbreaks are statistically linked to North Atlantic hurricanes and warm sea temperatures. The amoebic pathogen Paramoeba invadens, which causes these disease outbreaks, is unable to withstand typical minimum sea temperatures along this coast, suggesting that it is reintroduced during periods of peak temperatures. Here, we examine hypotheses for mechanisms of introduction or persistence of P. invadens using data on sea urchin disease outbreaks from a 5-yr field experiment (2010-2014), in combination with high-temporal-resolution oceanographic and meteorologic data. Disease outbreaks were observed in 4 yr, with the onset of mass mortality (≥50% morbidity or mortality) ranging from mid August to mid October. Physical data suggest that P. invadens originates in warm offshore surface waters that are transported to the coast during strong storms. Disease outbreaks were associated with passing hurricanes in 2 yr (2010 and 2011). In 2012, a disease outbreak occurred in the absence of a storm and following a strong positive anomaly in winter sea temperature, suggesting survival of the pathogen from 2011. In 2014, a disease outbreak occurred in association with a strong storm (nor'easter) that was not a hurricane. Our findings indicate that changing environmental conditions of increasing minimum sea temperatures and increasing intensity of storms may be altering the dynamics of this disease. These inferences remain equivocal, however, indicating the need for high-resolution dynamical modeling of the physical environment and rapid identification of P. invadens in the environment using genetic tools. [ABSTRACT FROM AUTHOR]

Mediastinal lesions occur in a wide variety of clinical conditions. Metastatic granulosa cell tumour (GCT) in the mediastinum is a rare occurrence. We report a case of a woman who had a metastatic (GCT) in her mediastinum 40 years after treatment of the initial neoplasm. Surgical resection of the mediastinal mass revealed a low-grade epithelioid neoplasm with coffee bean-shaped nuclei and immunohistochemical stains that were consistent with metastatic GCT. [ABSTRACT FROM AUTHOR]

Gamma delta (γδ) T cells contribute to both innate and acquired immune responses during infection. In this pilot study, we measured the in vitro responses of γδT cell populations from patients with sepsis compared to cells from healthy subjects. We also measured production of interferon (IFN)γ. Mononuclear cells were isolated from 10 healthy control subjects and 20 patients with sepsis. Cells were cultured for 7 days with interleukin (IL)-2 plus the bisphosphonate zoledronic acid which results in indirect cell activation. Flow cytometry was used to characterise the γδT cells and enzyme immunoassay was used to measure IFNγ production. The median [range] proportion of γδT cells in healthy controls after activation was 19.2% [2.0-55.9%], compared to only 0.61% [0.1-3.6%] ( P < 0.0001) in patients with sepsis. However, IFNγ levels in culture supernatants were similar in both the patients and healthy subjects. We therefore characterised the cells further by CD27 and CD45RA expression in a additional group of patients and found that the population of γδT cells was mainly CD27 negative which characterised these cells as non-proliferating effector cells. Our results suggest predominance of a non-proliferative effector subset of γδT cells in patients with sepsis, which retain functional activity and may contribute towards the host response to inflammation and infection. [ABSTRACT FROM AUTHOR]

A homodimeric fusion DNA vaccine targeting idiotype (Id) to antigen-presenting cells (APC) induced robust tumor protection in a mouse model of multiple myeloma (MM). Similar Id vaccine molecules were generated for four patients with MM with three main objectives: (i) do the vaccine molecules induce bona fide anti-Id immune responses in mice? (ii) does targeting of the vaccine molecules to APC enhance immune responses? (iii) can anti-Id antibodies, generated as by-product in vaccinated mice, be used to establish sensitive assays for complete remission (CR) prior to patient vaccination? Chimeric vaccine molecules targeting patient Id to mouse major histocompatibility complex (MHC) class II molecules were genetically constructed for four patients with MM. DNA vaccination of mice with chimeric vaccines targeting patient Id to mouse MHC class II molecules elicited antibodies specific for the patient's myeloma protein. Targeting MHC class II greatly enhanced anti-Id responses. Mouse anti-Id antibodies were used to establish myeloma protein-specific enzyme-linked immunosorbent assays (ELISAs) that were between 75 and 1500 times more sensitive than conventional serum protein electrophoresis and immunofixation. These results pave the way for testing targeted DNA Id vaccines in patients in CR. Id- and patient-specific ELISA could be established affording evaluation of CR depth beyond current serological methods. [ABSTRACT FROM AUTHOR]

Nitrogen-containing bisphosphonates indirectly activate Vγ9Vδ2 T cells through inhibition of farnesyl pyrophosphate synthase and intracellular accumulation of isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP), but the cells responsible for Vγ9Vδ2 T cell activation through IPP/DMAPP accumulation are unknown. Treatment of human peripheral blood mononuclear cells (PBMCs) with a pharmacologically relevant concentration of zoledronic acid induced accumulation of IPP/DMAPP selectively in monocytes, which correlated with efficient drug uptake by these cells. Furthermore, zoledronic acid-pulsed monocytes triggered activation of γδ T cells in a cell contact-dependent manner. These observations identify monocytes as the cell type directly affected by bisphosphonates responsible for Vγ9Vδ2 T cell activation. [ABSTRACT FROM AUTHOR]

BACKGROUND: The heavy-chain V4-34 germline gene segment is mandatory for pathologic cold-reacting autoantibodies with anti-I/i specificity (cold agglutinins) and is also preferentially used by monoclonal immunoglobulin M alloantibodies against D and other Rh antigens. The use of the V4-34 segment by monoclonal anti-D has previously been shown to also confer anti-I/i reactivity (cold agglutinin activity), which has implications for the use of such antibodies for Rh blood typing. V4-34 framework 1 (FR1) sequence is believed to be critical for cold agglutinin activity of cold agglutinins. STUDY DESIGN AND METHODS: The aim of this investigation was to use site-directed mutagenesis of a recombinant V4-34-encoded anti-D to determine the contribution of V4-34 FR1 sequence to anti-D activity and whether mutational modifications in the FR1 region could separately alter anti-D and anti-i activities. RESULTS: The results show that amino acid changes in V4-34 FR1 at W7, A23, and Y25 have a profound effect on anti-D activity as well as on anti-i activity. It was not possible to substantially reduce or remove anti-i activity without reducing anti-D activity to a comparable extent. CONCLUSIONS: The same nonpolar hydrophobic amino acids in FR1 are critical for maintaining both anti-D and anti-i activity. It is proposed that these residues influence the conformation of the antigen-binding site. [ABSTRACT FROM AUTHOR]

Recombinant human transcobalamin (TC) was probed with 17 monoclonal antibodies (mAbs), using surface plasmon resonance measurements. These experiments identified five distinct epitope clusters on the surface of holo-TC. Western blot analysis of the CNBr cleavage fragments of TC allowed us to distribute the epitopes between two regions, which spanned either the second quarter of the TC sequence GQLA...TAAM(103–198) or the C-terminal peptide LEPA...LVSW(316–427). Proteolytic fragments of TC and the synthetic peptides were used to further specify the epitope map and define the functional domains of TC. Only one antibody showed some interference with cobalamin (Cbl) binding to TC, and the corresponding epitope was situated at the C-terminal stretch TQAS...QLLR(372–399). We explored the receptor-blocking effect of several mAbs and heparin to identify TC domains essential for the interaction between holo-TC and the receptor. The receptor-related epitopes were located within the TC sequence GQLA...HHSV(103–159). The putative heparin-binding site corresponded to a positively charged segment KRSN...RTVR(207–227), which also seemed to be necessary for receptor binding. We conclude that conformational changes in TC upon Cbl binding are accompanied by the convergence of multiple domains, and only the assembled conformation of the protein (i.e. holo-TC) has high affinity for the receptor. [ABSTRACT FROM AUTHOR]

Due to somatic recombination and hypermutation, Ig variable heavy (V) and light (V) regions contain unique immunogenic determinants, idiotopes (Id), which can stimulate T cells. To address the relevance of this in a human disease, monoclonal IgG (mAb)-secreting B cell clones were established from the cerebrospinal fluid (CSF) of two patients with multiple sclerosis (MS). HLA-DR-restricted CD4 T cell lines and clones from CSF of both patients specifically recognized autologous CSF mAb. The CSF T cell clones produced IFN-γ; some also produced TNF-α, IL-10 and IL-5. V and V on the monoclonal IgG derived from CSF B cells expressed amino acid replacements due to somatic mutations. A T cell epitope was mapped to a V framework region, where an amino acid replacement was critical for the T cell recognition. The finding of Id-specific T cells and Id-bearing B cells in the CSF indicates that they coexist within the diseased organ, and provide a basis for the study of Id-driven T-B cell collaboration in a human autoimmune disease. [ABSTRACT FROM AUTHOR]

Here, we have investigated if targeting of T cell epitopes to chemokine receptors results in improved CD4 T cell responses. Mouse monoclonal antibodies (mAb) with κL chains were targeted to various chemokine receptors expressed on human monocytes or immature dendritic cells (DC), and proliferation of cloned human, DR4-restricted CD4 T cells specific for mouse Cκ was measured. When using monocytes as antigen-presenting cells, mAb specific for CCR1, CCR2, CCR5, and CXCR4 were 100-10,000-fold more efficient at inducing T cell proliferationwhen compared to isotype-matched control mAb on a per molecule basis. Targeting of immature DC was less effective and was only seen with anti-CCR1 and anti-CXCR4 mAb. Anti-chemokine receptors mAb required to be processed by the conventional endosomal MHC class II presentation pathway. The mAb did not induce signaling through the chemokine receptors as they failed to induce mobilization of cytosolic Ca and actin polymerization. They also failed to induce APC maturation. The results strongly suggest that chemokine receptors channel antigen into the endocytic pathway for presentation on MHC class II molecules. Targeting T cell epitopes to chemokine receptors by recombinant antibody should be a useful vaccine strategy for the induction of strong CD4 T cell responses. [ABSTRACT FROM AUTHOR]

1. The effect of macrophytes on the spatial distribution of littoral rotifers was examined in Lake Rotomanuka, New Zealand (37°55′S, 175°19′E). Total rotifer abundances and those of abundant species, were compared between three macrophyte species, Myriophyllum propinquum, Eleocharis sphacelata and Egeria densa, and spatially across a littoral transect in relation to these species. 2. The abundances of many species, for example Euchlanis dilatata, Lecane closterocerca and L. lunaris, differed significantly between macrophyte species. More planktonic forms, Ascomorpha saltans, Keratella cochlearis and Synchaeta oblonga, however, showed no significant preference for macrophyte species. 3. Differences in rotifer abundances were evident even when different species of macrophyte grew in close proximity to one another, indicating that variations in physical and chemical conditions, which occur in the littoral of Lake Rotomanuka, could be largely discounted for much of the variation between macrophyte species. 4. Variation in rotifers between macrophytes was probably the result of a number of factors, including differences in macrophyte morphology, macrophyte age, epiphytic algal growths and the differential effects of predation by invertebrates and fish between macrophytes. 5. Variability of rotifer abundances spatially across the ecotone was less marked than between macrophyte species. The species of macrophyte occurring, and therefore the community composition and distribution of macrophyte species in the littoral, appears to be a major influence in the spatial structuring of rotifer communities in the littoral region of lakes. [ABSTRACT FROM AUTHOR]

7B4, B6 and D12 are murine monoclonal antibodies (MoAb) that bind to some human immunoglobulin heavy chain products of the closely related V3–30, V3–30.3 and V3–33 genes from the VH3 family. B6 and D12 have additional reactivities with some immunoglobulins (Ig) encoded by the V3–11 and V3–7 genes; D12 also reacts with some V3–43 gene Ig. We show here, by site-directed mutagensis, that the lysine at position 57 in the complementarity-determining region 2 (CDR-2) of the V3–30 gene product is crucial for epitope recognition by all three anti-VH3 MoAbs. Further analysis of the amino-acid sequences of a large panel of Ig reactive, or nonreactive, with MoAb 7B4 indicates that the determinant recognized by 7B4 is dependent on the presence of the tetrapeptide sequence NKYY between positions 56 and 59 in the CDR-2. Comparing the efficiency of 7B4 reactivity with VH3 gene-encoded human Ig indicates that amino-acid position 4 in the frame region 1 (FR-1) may also influence the binding of 7B4 to Ig encoded by three very closely related germline genes, V3–30, V3–30.3 and V3–33. NKYY is also found on the gp120 V3 region of human immunodeficiency virus (HIV)-2, SIV and HTLV-4. We also report that other tetrapeptide sequences found on the 56–59 motif of heavy chain variable regions encoded by germline genes are expressed on the solvent exposed V2 region of gp120 of HIV-1 isolates. The possible significance of these observations is discussed. [ABSTRACT FROM AUTHOR]

Presents several abstracts on autoantibodies and autoimmunity. 'The Role of IL-12 in Autoimmune Diabetes,' by L. Adorini; 'The Role of HLA Genes in Coelic Disease,' by L.M. Sollid; 'B Cell Subsets and their Relation to Autoreactivity and Autoimmunity,' by M. Ferrarini.

The removal of ammonium-nitrogen (NH4-N) from groundwaterflowing through a grey willow (Salix cinerea) and native raupo (Typha orientalis) lake edge wetland was Investigated over 10 mo. Monthly groundwater sampling showed that more than 95% of incoming NH4-N was removed by the wetland. Laboratory assays found that the potential for NH4-N removal by nitrification and subsequentdenitrification was significantly higher in the organic surface sediments than in deeper, sandy, root zone sediments. In a laboratory 15N-tracer experiment, Salix cinerea and Typha orientalis readily assimilated NH4-N from the root zone and were found to enhance the natural diffusion of NH4-N up through the organic sediment layer and into the overlying water. Estimated rates ofremoval of NH4-N by coupled nitrification- denitrification and plant assimilation were 8 to 9 mg of N m- 2 d-1 and 9 to 46 mg of N m- 2 d-1, respectively. Assimilated N, returned to the overlying water-sediment surface via liner fall may be subject to sequential mineralization, nitrification, and denitrification, resulting in gaseous N loss. Sixty-five percent of the 15N in Salix leaf litter was transformed in such a way. The results of this study indicate that plants play a central role in the NH4-N processing capacity of wetlands. The relevance of our findings to constructed wetland design is discussed. [ABSTRACT FROM AUTHOR]

Nitrogen-containing bisphosphonates indirectly activate Vgamma9Vdelta2 T cells through inhibition of farnesyl pyrophosphate synthase and intracellular accumulation of isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP), but the cells responsible for Vgamma9Vdelta2 T cell activation through IPP/DMAPP accumulation are unknown. Treatment of human peripheral blood mononuclear cells (PBMCs) with a pharmacologically relevant concentration of zoledronic acid induced accumulation of IPP/DMAPP selectively in monocytes, which correlated with efficient drug uptake by these cells. Furthermore, zoledronic acid-pulsed monocytes triggered activation of gammadelta T cells in a cell contact-dependent manner. These observations identify monocytes as the cell type directly affected by bisphosphonates responsible for Vgamma9Vdelta2 T cell activation.