Cytokine-induced neutrophil chemoattractant 1 (CINC-1) mediates the sympathetic component of inflammatory mechanical hypersensitivitiy in rats

The hyperalgesic effect of cytokine-induced neutrophil chemoattractant 1 (CINC-1/CXCL1) was measured in a model of mechanical hyperalgesia in rats. CINC-1 evoked a dose-dependent mechanical hypersensitivity, which was already significant 2 h after the cytokine injection, peaked 4 h after and decreased thereafter. The local pre-treatment of the rats with the beta-adrenoceptor antagonist, atenolol (25 microg paw-1), but not with the cyclooxygenase inhibitor indomethacin (100 microg paw-1), inhibited (86%) the CINC-1-induced hypersensitivity. Conversely, IL-1beta-evoked hypersensitivity was inhibited (76%) by local pre-treatment of the animals with indomethacin, but not by atenolol. Carrageenin- and TNF-alpha-evoked hypersensitivity were attenuated to about the same extent (50%) by antisera neutralising CINC-1 or IL-1beta. The association of both antisera abolished the hypersensitivity effect of carrageenin and TNF-alpha. In addition, carrageenin, LPS and TNF-alpha were shown to stimulate the production of immunoreactive CINC-1 in the skin of injected paws. These data suggest that CINC-1, released at sites of inflammation, mediates inflammatory hyperalgesia in rats via release of sympathomimetic amines.     

Antiproliferative effect of conjugated linoleic acid in caco-2 cells: involvement of PPARgamma and APC/beta-catenin pathways

Conjugated linoleic acid (CLA), a naturally occurring substance in food sources, occurs as mixtures of positional and geometrical isomers of octadecadienoate (18:2), and may inhibit colon tumorigenesis. It has been hypothesized that CLA can modulate cell proliferation and differentiation through the activation of peroxisome proliferator-activated receptors (PPARs), among which PPARgamma is involved in growth inhibition of transformed cells. The aim of the present study was to investigate whether the antiproliferative effects of CLA are mediated by its interaction with PPARgamma and APC/beta-catenin signalling pathway in human colon cancer cells. In CLA-treated caco-2 cells we found a remarkable increase in the expression of PPARgamma, which translocated into the nucleus, while PPARalpha and beta/delta protein levels were not affected. GW259662, a well known PPARgamma antagonist, blocked the increase in PPARgamma protein rate and abrogated some biological effects of CLA, as it restored the proliferative capability of the cells and ERK1/2 phosphorylation level. We demonstrated that CLA treatment determined the down-regulation of APC and c-myc proteins, but in this case the administration of the antagonist was not able to revert CLA effects. Furthermore, CLA induced a reorganization of E-cadherin and beta-catenin, as well as a redistribution of actin and tubulin filaments. Our data suggest that CLA may regulate PPARgamma expression by selectively acting as an agonist; however, the discrepancies in PPARgamma antagonist efficacy suggest the involvement of other pathways, independent of PPARgamma, in CLA antiproliferative activity.     

Vegetation Control Treatments to Favor Naturally Regenerated Betula alleghaniensis Saplings Following Seed-Tree Cut: Sapling Monitoring Two Years after Treatment

Control of competing vegetation is recommended to ensure successful Yellow birch (Betula alleghaniensis Britton) regeneration within juvenile stands that do not sustain high enough sapling densities of this species. Four contrasting vegetation control treatments were tested to determine their effect on the growth and vigor of eight‐year‐old B. alleghaniensis saplings regenerating after final cutting of a shelterwood seed cut. Vegetation control treatments were TC (total circular removal), PC (circular removal of codominant competing vegetation), TS (total semicircular removal on 180° section), and NC (no vegetation control). Two years after treatment application, diameter growth significantly improved in response to vegetation control treatments, whereas sapling height growth did not. This pattern of biomass allocation was directly related to sapling etiolation, which increased with decreasing severity of vegetation removal. As a result, application of vegetation control, especially TC and PC treatments, was valuable in reducing signs of stress in saplings. However, increasing the severity of vegetation removal also made saplings more conspicuous to herbivores, which increased browsing, especially in the TC and PC treatments. Browsing was sufficient in some plots of the TC and PC treatments to overcome the vigor and diameter growth enhancements observed when browsing was negligible. In contrast to the TC and PC treatments, the TS treatment kept browsing very low while largely removing competition. The results suggest that B. alleghaniensis saplings established after final cutting of a shelterwood seed cut do take advantage of vegetation control treatments, but the decision to apply these treatments must include consideration of local herbivore population densities.     

Transgenic plants expressing human glutamic acid decarboxylase (GAD65), a major autoantigen in insulin-dependent diabetes mellitus

Parenteral and oral administration of autoantigens can induce immune tolerance in autoimmune diseases. Prophylactic therapy based on oral administration of human autoantigens is not, however, feasible when sufficient quantities of candidate autoantigens are not available. Transgenic plants that express high levels of recombinant proteins would allow large quantities of autoantigens to be produced at relatively low costs. In addition, transgenic food would provide a simple and direct method of delivering autoantigens. The production and the characterization of transgenic tobacco and carrot plants expressing human GAD65, a major autoantigen in human insulin-dependent diabetes mellitus (IDDM), is reported. Immunogold labeling and electron microscopy of transgenic tobacco tissue shows the selective targeting of human GAD65 to chloroplast tylacoids and mitochondria. In planta expressed GAD65 has a correct immunoreactivity with IDDM-associated autoantibodies and retains enzymatic activity, a finding that suggests a correct protein folding. In transgenic tobacco and carrot the expression levels of human GAD65 varies between 0.01% and 0.04% of total soluble proteins. Transgenic edible plant organs are now available to study the feasibility of inducing immune tolerance in IDDM animals by oral administration of GAD65.     

Soft Tissue Artefacts of the Human Back: Comparison of the Sagittal Curvature of the Spine Measured Using Skin Markers and an Open Upright MRI

Soft tissue artefact affects the determination of skeletal kinematics. Thus, it is important to know the accuracy and limitations of kinematic parameters determined and modelled based on skin marker data. Here, the curvature angles, as well as the rotations of the lumbar and thoracic segments, of seven healthy subjects were determined in the sagittal plane using a skin marker set and compared to measurements taken in an open upright MRI scanner in order to understand the influence of soft tissue artefact at the back. The mean STA in the flexed compared to the extended positions were 10.2±6.1 mm (lumbar)/9.3±4.2 mm (thoracic) and 10.7±4.8 mm (lumbar)/9.2±4.9 mm (thoracic) respectively. A linear regression of the lumbar and thoracic curvatures between the marker-based measurements and MRI-based measurements resulted in coefficients of determination, R², of 0.552 and 0.385 respectively. Skin marker measurements therefore allow for the assessment of changes in the lumbar and thoracic curvature angles, but the absolute values suffer from uncertainty. Nevertheless, this marker set appears to be suitable for quantifying lumbar and thoracic spinal changes between quasi-static whole body postural changes.     

Elemental profile of cerebrospinal fluid in patients with Parkinson''s disease

To ascertain the potential role of chemical elements (namely, Al, Ba, Be, Bi, Ca, Cd, Co, Cr, Cu, Fe, Hg, Li, Mg, Mn, Mo, Ni, Pb, Sb, Si, Sn, Sr, Tl, V, W, Zn and Zr) as markers in the Parkinson's disease (PD), the elemental concentration of cerebrospinal fluid (CSF) of 42 patients with PD and 20 age-matched controls was assessed. Analyses were performed by Inductively Coupled Plasma Atomic Emission Spectrometry (ICP-AES) and Sector Field Inductively Coupled Plasma Mass Spectrometry (SF-ICP-MS). Significantly lower levels of Co, Cr, Fe, Pb, Si and Sn were observed in the CSF of PD patients compared with those in controls, with a percentage of depletion up to 50% for Cr and Pb. No such variations were detected for all the other elements. Results suggested that Pb, Cr, Fe were the most suitable elements to distinguish between normality and PD. Different cut-off concentrations for these elements could be tentatively proposed as a predictive tool for the PD condition.     

Polymorphism analysis of the CTLA-4 gene in paracoccidioidomycosis patients

The CTLA-4 protein is expressed in activated T cells and plays an essential role in the immune response through its regulatory effect on T cell activation. Polymorphisms of the CTLA-4 gene have been correlated with autoimmune, neoplastic and infectious illnesses. This work aimed to verify possible associations between single nucleotide polymorphisms (SNPs) in CTLA-4, -318C/T in the promoter and +49A/G in exon 1 and paracoccidioidomycosis (PCM) caused by Paracoccidioides brasiliensis. For this purpose, 66 chronic form PCM patients and 76 healthy controls had their allele, genotype and haplotype frequencies determined. The genetic admixture structure of the patients and controls was evaluated to eliminate ancestral bias. The comparison of frequencies indicated no significant differences between patients and controls that could link the SNPs to PCM. Groups were admixture matched with no difference observed in population ancestry inference, indicating that the absence of association between CTLA-4 polymorphisms and PCM could not be attributed to ancestral bias. This study showed that there was no association between the CTLA-4 SNPs -318 and +49 and the resistance or susceptibility to PCM.     

Serum levels of cytoplasmic melanoma-associated antigen at diagnosis may predict clinical relapse in neuroblastoma patients

The high molecular weight melanoma-associated antigen (HMW-MAA) and the cytoplasmic melanoma-associated antigen (cyt-MAA/LGALS3BP) are expressed in melanoma. Their serum levels are increased in melanoma patients and correlate with clinical outcome. We investigated whether these molecules can serve as prognostic markers for neuroblastoma (NB) patients. Expression of cyt-MAA and HMW-MAA was evaluated by flow cytometry in NB cell lines, patients' neuroblasts ((FI)-NB), and short-term cultures of these latter cells (cNB). LGALS3BP gene expression was evaluated by RT-qPCR on (FI)-NB, cNB, and primary tumor specimens. Soluble HMW-MAA and cyt-MAA were tested by ELISA. Cyt-MAA and HMW-MAA were expressed in NB cell lines, cNB, and (FI)-NB samples. LGALS3BP gene expression was higher in primary tumors and cNB than in (FI)-NB samples. Soluble cyt-MAA, but not HMW-MAA, was detected in NB cell lines and cNBs supernatants. NB patients' serum levels of both antigens were higher than those of the healthy children. High cyt-MAA serum levels at diagnosis associated with higher incidence of relapse, independently from other known risk factors. In conclusion, both HMW-MAA and cyt-MAA antigens, and LGALS3BP gene, were expressed by NB cell lines and patients' neuroblasts, and both antigens' serum levels were increased in NB patients. Elevated serum levels of cyt-MAA at diagnosis correlated with relapse, supporting that cyt-MAA may serve as early serological biomarker to individuate patients at higher risk of relapse that may require a more careful follow-up, after being validated in a larger cohort of patients at different time-points during follow-up. Given its immunogenicity, cyt-MAA may also be a potential target for NB immunotherapy.     

Using Skin Markers for Spinal Curvature Quantification in Main Thoracic Adolescent Idiopathic Scoliosis: An Explorative Radiographic Study

Background and Purpose

Although the relevance of understanding spinal kinematics during functional activities in patients with complex spinal deformities is undisputed among researchers and clinicians, evidence using skin marker-based motion capture systems is still limited to a handful of studies, mostly conducted on healthy subjects and using non-validated marker configurations. The current study therefore aimed to explore the validity of a previously developed enhanced trunk marker set for the static measurement of spinal curvature angles in patients with main thoracic adolescent idiopathic scoliosis. In addition, the impact of inaccurate marker placement on curvature angle calculation was investigated.

Methods

Ten patients (Cobb angle: 44.4±17.7 degrees) were equipped with radio-opaque markers on selected spinous processes and underwent a standard biplanar radiographic examination. Subsequently, radio-opaque markers were replaced with retro-reflective markers and the patients were measured statically using a Vicon motion capture system. Thoracolumbar / lumbar and thoracic curvature angles in the sagittal and frontal planes were calculated based on the centers of area of the vertebral bodies and radio-opaque markers as well as the three-dimensional position of the retro-reflective markers. To investigate curvature angle estimation accuracy, linear regression analyses among the respective parameters were used. The impact of inaccurate marker placement was explored using linear regression analyses among the radio-opaque marker- and spinous process-derived curvature angles.

Results and Discussion

The results demonstrate that curvatures angles in the sagittal plane can be measured with reasonable accuracy, whereas in the frontal plane, angles were systematically underestimated, mainly due to the positional and structural deformities of the scoliotic vertebrae. Inaccuracy of marker placement had a greater impact on thoracolumbar / lumbar than thoracic curvature angles. It is suggested that spinal curvature measurements are included in marker-based clinical gait analysis protocols in order to enable a deeper understanding of the biomechanical behavior of the healthy and pathological spine in dynamic situations as well as to comprehensively evaluate treatment effects.