Previous reports suggested that ex vivo cultured primary nasal epithelial cells from allergic patients differ from those from non-allergic individuals by genuinely reduced barrier function. By contrast, we found that primary nasal epithelial cells from allergic and non-allergic individuals showed comparable barrier function and secretion of cytokines.
Cytokines ; Epithelial Cells ; Humans ; Immunoglobulin E ; Rhinitis, Allergic

The effects of storage temperatures, repeated freeze-thaw cycles, or delays in separating plasma or serum from blood samples are largely unknown for heat shock protein 27 (HSP27). We evaluated (1) the imprecision of the HSP27 assay used in this study; (2) the in vitro stability of HSP27 in blood samples stored at 4℃ for up to 6 hr with immediate and delayed serum/plasma separation from cells; and (3) the in vitro stability of HSP27 in blood samples stored at -80℃ after repeated freeze-thaw cycles. The ELISA to detect HSP27 in this study showed a within-run CV of <9% and a total CV of <15%. After 4-6 hr of storage at 4℃, HSP27 concentrations remained stable when using serum tubes irrespective of sample handling, but HSP27 concentrations decreased by 25-45% when using EDTA plasma tubes. Compared with baseline HSP27, one freeze-thaw cycle had no effect on serum concentrations. However, plasma concentrations increased by 3.1-fold after one freeze-thaw cycle and by 7.3-fold after five freeze-thaw cycles. In conclusion, serum is an appropriate biological sample type for use in epidemiological and large-scale clinical studies.
*Enzyme-Linked Immunosorbent Assay ; Freezing ; HSP27 Heat-Shock Proteins/*blood ; Humans ; Protein Stability ; Reproducibility of Results ; Specimen Handling ; Temperature ; Time Factors

BACKGROUND AND OBJECTIVES: Mesenchymal stem cells (MSCs) become hypertrophic in long term despite chondrogenic differentiation following the pathway of growth plate chondrocytes. This terminal differentiation leads to phenotypically unstable cartilage and was mirrored in vitro by addition of hypertrophy inducing medium. We investigated how intrinsic TGF-β signaling is altered in pro-hypertrophic conditions. METHODS AND RESULTS: Human bone marrow derived MSC were chondrogenically differentiated in 3D culture. At day 14 medium conditions were changed to 1. pro-hypertrophic by addition of T3 and withdrawal of TGF-β and dexamethasone 2. pro-hypertrophic by addition of BMP 4 and withdrawal of TGF-β and dexamethasone and 3. kept in prochondrogenic medium conditions. All groups were treated with and without TGFβ-type-1-receptor inhibitor SB431542 from day 14 on. Aggregates were harvested for histo- and immunohistological analysis at d14 and d28, for gene expression analysis (rt-PCR) on d1, d3, d7, d14, d17, d21 and d28 and for Western blot analysis on d21 and d28. Induction of hypertrophy was achieved in the pro-hypertrophic groups while expression of TGFβ-type-1- and 2-receptor and Sox 9 were significantly downregulated compared to pro-chondrogenic conditions. Western blotting showed reduced phosphorylation of Smad 2 and 3 in hypertrophic samples, reduced TGF-β-1 receptor proteins and reduced SOX 9. Addition of SB431542 did not initiate hypertrophy under pro-chondrogenic conditions, but was capable of enhancing hypertrophy when applied simultaneously with BMP-4. CONCLUSIONS: Our results suggest that the enhancement of hypertrophy in this model is a result of both activation of pro-hypertrophic BMP signaling and reduction of anti-hypertrophic TGFβ signaling.
Blotting, Western ; Bone Marrow ; Cartilage ; Chondrocytes ; Chondrogenesis ; Dexamethasone ; Down-Regulation ; Gene Expression ; Growth Plate ; Humans ; Hypertrophy ; In Vitro Techniques ; Mesenchymal Stromal Cells ; Phosphorylation

The implantation of bioengineered scaffolds into lesion-induced gaps of the spinal cord is a promising strategy for promoting functional tissue repair because it can be combined with other intervention strategies. Our previous investigations showed that functional improvement following the implantation of a longitudinally microstructured collagen scaffold into unilateral mid-cervical spinal cord resection injuries of adult Lewis rats was associated with only poor axon regeneration within the scaffold. In an attempt to improve graft-host integration as well as functional recovery, scaffolds were seeded with highly enriched populations of syngeneic, olfactory bulb-derived ensheathing cells (OECs) prior to implantation into the same lesion model. Regenerating neurofilament-positive axons closely followed the trajectory of the donor OECs, as well as that of the migrating host cells within the scaffold. However, there was only a trend for increased numbers of regenerating axons above that supported by non-seeded scaffolds or in the untreated lesions. Nonetheless, significant functional recovery in skilled forelimb motor function was observed following the implantation of both seeded and non-seeded scaffolds which could not be correlated to the extent of axon regeneration within the scaffold. Mechanisms other than simple bridging of axon regeneration across the lesion must be responsible for the improved motor function.
Adult ; Animals ; Axons* ; Collagen ; Forelimb ; Humans ; Rats ; Regeneration* ; Spinal Cord Injuries* ; Spinal Cord* ; Tissue Donors

Background: and Purpose Knowledge about different etiologies of non-traumatic intracerebral hemorrhage (ICH) and their outcomes is scarce. Methods: We assessed prevalence of pre-specified ICH etiologies and their association with outcomes in consecutive ICH patients enrolled in the prospective Swiss Stroke Registry (2014 to 2019). Results We included 2,650 patients (mean±standard deviation age 72±14 years, 46.5% female, median National Institutes of Health Stroke Scale 8 [interquartile range, 3 to 15]). Etiology was as follows: hypertension, 1,238 (46.7%); unknown, 566 (21.4%); antithrombotic therapy, 227 (8.6%); cerebral amyloid angiopathy (CAA), 217 (8.2%); macrovascular cause, 128 (4.8%); other determined etiology, 274 patients (10.3%). At 3 months, 880 patients (33.2%) were functionally independent and 664 had died (25.1%). ICH due to hypertension had a higher odds of functional independence (adjusted odds ratio [aOR], 1.33; 95% confidence interval [CI], 1.00 to 1.77; P=0.05) and lower mortality (aOR, 0.64; 95% CI, 0.47 to 0.86; P=0.003). ICH due to antithrombotic therapy had higher mortality (aOR, 1.62; 95% CI, 1.01 to 2.61; P=0.045). Within 3 months, 4.2% of patients had cerebrovascular events. The rate of ischemic stroke was higher than that of recurrent ICH in all etiologies but CAA and unknown etiology. CAA had high odds of recurrent ICH (aOR, 3.38; 95% CI, 1.48 to 7.69; P=0.004) while the odds was lower in ICH due to hypertension (aOR, 0.42; 95% CI, 0.19 to 0.93; P=0.031). Conclusions Although hypertension is the leading etiology of ICH, other etiologies are frequent. One-third of ICH patients are functionally independent at 3 months. Except for patients with presumed CAA, the risk of ischemic stroke within 3 months of ICH was higher than the risk of recurrent hemorrhage.

Objective: To determine the utility of a highly sensitive troponin assay when utilized in the emergency department. Methods The FAST-TRAC study prospectively enrolled >1,500 emergency department patients with suspected acute coronary syndrome within 6 hours of symptom onset and 2 hours of emergency department presentation. It has several unique features that are not found in the majority of studies evaluating troponin. These include a very early presenting population in whom prospective data collection of risk score parameters and the physician’s clinical impression of the probability of acute coronary syndrome before any troponin data were available. Furthermore, two gold standard diagnostic definitions were determined by a pair of cardiologists reviewing two separate data sets; one that included all local troponin testing results and a second that excluded troponin testing so that diagnosis was based solely on clinical grounds. By this method, a statistically valid head-to-head comparison of contemporary and high sensitivity troponin testing is obtainable. Finally, because of a significant delay in sample processing, a unique ability to define the molecular stability of various troponin assays is possible.Trial registration ClinicalTrials.gov Identifier NCT00880802