Characterization of a migrative subpopulation of adult human nasoseptal chondrocytes with progenitor cell features and their potential for in vivo cartilage regeneration strategies.

BACKGROUND: Progenitor cells display interesting features for tissue repair and reconstruction. In the last years, such cells have been identified in different cartilage types. In this study, we isolated a migrative subpopulation of adult human nasoseptal chondrocytes with progenitor cell features by outgrowth from human nasal septum cartilage. These putative progenitor cells were comparatively characterized with mesenchymal stem cells (MSC) and human nasal septum chondrocytes with respect to their cellular characteristics as well as surface marker profile using flow cytometric analyses. Differentiation capacity was evaluated on protein and gene expression levels. RESULTS: The migrative subpopulation differentiated into osteogenic and chondrogenic lineages with distinct differences to chondrocytes and MSC. Cells of the migrative subpopulation showed an intermediate surface marker profile positioned between MSC and chondrocytes. Significant differences were found for CD9, CD29, CD44, CD90, CD105 and CD106. The cells possessed a high migratory ability in a Boyden chamber assay and responded to chemotactic stimulation. To evaluate their potential use in tissue engineering applications, a decellularized septal cartilage matrix was either seeded with cells from the migrative subpopulation or chondrocytes. Matrix production was demonstrated immunohistochemically and verified on gene expression level. Along with secretion of matrix metalloproteinases, cells of the migrative subpopulation migrated faster into the collagen matrix than chondrocytes, while synthesis of cartilage specific matrix was comparable. CONCLUSIONS: Cells of the migrative subpopulation, due to their migratory characteristics, are a potential cell source for in vivo regeneration of nasal cartilage. The in vivo mobilization of nasal cartilage progenitor cells is envisioned to be the basis for in situ tissue engineering procedures, aiming at the use of unseeded biomaterials which are able to recruit local progenitor cells for cartilage regeneration.

Effect of intracerebroventricular injection of TiO2 nanoparticles on complex behaviour in the rat.

There are no data available on the behavioural effects of centrally administered nanoparticles in freely moving intact mammals. Consequently, in the current study male Sprague-Dawley rats were trained to respond under an alternating-lever cyclic-ratio (ALCR) schedule of food reinforcement. Under this schedule, ascending and descending sequences of fixed-ratio (FR) lever press requirements for food reinforcement were presented over six cycles, with each discrete FR component completed on the alternate lever to the previous component. The final version of the schedule was comprised of an ascending followed by a descending sequence of the ratio values 2, 6, 12, 20, 30, 42 and 56, repeated over six cycles. When the rats were able to complete this version of the ALCR schedule in 40 min, each was implanted with a permanently indwelling ICV cannula aimed at the lateral ventricle of the brain, and allowed to recover for 7 days. On the first day of the experiment, all rats were injected with either titanium dioxide (TiO2, 9 nm, stabilised with gallic acid, 10 microl volume, 2 mg/ml) nanoparticles, or 10 microl saline (control). Two-hours after the ICV injections, the behaviour of all rats was measured using the ALCR schedule, and their behaviour was also measured (no ICV injection) for the next 7 days. Under the ALCR schedule, the number of lever-switching errors and incorrect lever perseverations significantly increased in the TiO2 group (p < 0.05). Other parameters of the ALCR schedule (RRRs and PRPs), which indicate the induction of malaise or general motor retardation, were not altered following ICV TiO2 injection. The findings of the current study indicate that central administration of TiO2 nanoparticles induced behavioural deterioration in freely moving intact animals, that the induced behavioural deterioration was a result of central rather than peripheral outcomes, and that this effect was chronic rather than acute.

Ecotoxicity and uptake of polymer coated gold nanoparticles.

Bioconjugated gold nanoparticles (Au NPs) are a promising tool for pharmaceutical applications. However, the ecotoxicity of these types of NPs has hardly been studied. We investigated the ecotoxicity and uptake of 4-5 nm Au NPs to which two types of polymer coatings were attached. One coating was an amphiphilic polymer only and the other an amphiphilic coating to which 10 kDa polyethylene glycol chains were attached. In both 72 h algal growth inhibition tests with the alga Pseudokirchneriella subcapitata and in 24 h resazurin cytotoxicity tests with the rainbow trout gill cell line RTGill-W1, the pegylated Au NPs were found less toxic compared to the amphiphilic coated particles. No uptake or direct interaction between particles and algal cells was observed. However, uptake/adsorption in fish gill cells reached up to >10(6) particles/cell after 1 h and particles were eliminated for ≥96% after 24 h depuration. Both particle types were found within membrane enclosed vesicles in the cytoplasm of RTgill-W1 cells.

Focussed ion beam milling at grazing incidence angles.

A combined scanning electron microscope and focussed ion beam instrument is suitable for micro- and nanopatterning, cross-sectioning and subsequent imaging, of specimens at room temperature as well as under cryo conditions. In order to reveal internal details, samples are conventionally milled with the ion beam positioned perpendicular to the sample surface. Using this approach certain limitations are frequently encountered, e.g. accumulation of redeposited material, shadowing effects, image distortion and a limited imaging area. Here we show an approach in which samples are pre-trimmed using a microtome to obtain a sample block face that is parallel to the ion beam. This new grazing incidence geometry eliminates the need for removal of bulk material with the ion beam and enables immediate fine polishing of a pre-selected area of interest. Many of the limitations previously described are avoided and in addition milling time is reduced, whilst creating larger cross-sectional areas. Another advantage is that electron imaging can be accomplished by tilting the sample surface perpendicular to the electron beam, providing a geometrically undistorted image. The proposed approach is suitable for materials that can be microtomed, both in ambient and cryogenic conditions, and proves to be of particular benefit for biological and food samples.

High speed laser tomography system.

A high speed laser tomography system was developed capable of acquiring three-dimensional (3D) images of optically thin clouds of moving micron-sized particles. It operates by parallel-shifting an illuminating laser sheet with a pair of galvanometer-driven mirrors and synchronously recording two-dimensional (2D) images of thin slices of the imaged volume. The maximum scanning speed achieved was 120,000 slices/s, sequences of 24 volume scans (up to 256 slices each) have been obtained. The 2D slices were stacked to form 3D images of the volume, then the positions of the particles were identified and followed in the consecutive scans. The system was used to image a complex plasma with particles moving at speeds up to cm/s.

Prognostic value of N-terminal pro-B-type natriuretic peptide for conservatively and surgically treated patients with aortic valve stenosis.

OBJECTIVE: To evaluate the prognostic value of N-terminal pro-B-type natriuretic peptide (NT-proBNP) in patients with aortic stenosis being treated conservatively or undergoing aortic valve replacement (AVR). METHODS: 159 patients were followed up for a median of 902 days. 102 patients underwent AVR and 57 were treated conservatively. NT-proBNP at baseline was raised in association with the degree of severity and of functional status. RESULTS: During follow up 21 patients (13%) died of cardiac causes or required rehospitalisation for decompensated heart failure. NT-proBNP at baseline was higher in patients with an adverse outcome than in event-free survivors (median 623 (interquartile range 204-1854) pg/ml v 1054 (687-2960) pg/ml, p = 0.028). This difference was even more obvious in conservatively treated patients (331 (129-881) pg/ml v 1102 (796-2960) pg/ml, p = 0.002). Baseline NT-proBNP independently predicted an adverse outcome in the entire study group and in particular in conservatively treated patients (area under the curve (AUC) = 0.65, p = 0.028 and AUC = 0.82, p = 0.002, respectively) but not in patients undergoing AVR (AUC = 0.544). At a cut-off value of 640 pg/ml, baseline NT-proBNP was discriminative for an adverse outcome. CONCLUSION: NT-proBNP concentration is related to severity of aortic stenosis and provides independent prognostic information for an adverse outcome. However, this predictive value is limited to conservatively treated patients. Thus, the data suggest that assessing NT-proBNP may have incremental value for selecting the optimal timing of valve replacement.

Release pattern of N-terminal pro B-type natriuretic peptide (NT-proBNP) in acute coronary syndromes.

BACKGROUND: Recently it has been found that BNP and NT-proBNP provide independent prognostic information in patients with acute coronary syndromes (ACS). However, little data are available on the time course of NT-proBNP levels in relation to onset of symptoms. METHODS AND RESULTS: We included 765 patients (236 females, aged 64 +/- 11 years) with an ACS (STEMI 42%, NSTEMI 41%, UAP 17%), who were referred for coronary angiography. NT-proBNP was assessed on admission and the next day. NT-proBNP values were related to the time duration from onset of symptoms until blood drawing with lowest values within 3 h and highest values 24-36 h after onset of symptoms (147 (64-436) pg/ml and 1099 (293-3795) pg/ml, respectively, p < 0.001). Highest values for NT-proBNP on admission were found in patients with NSTEMI compared to patients with STEMI and UAP (912 (310-2258) pg/ml) vs 262 (85-1282) pg/ml) vs 182 (74- 410) pg/ml; p < 0.001), but no difference was present between STEMI and NSTEMI the day after admission (1325 (532-2974) pg/ ml vs 1169 (555-3413) pg/ml; p = 0.676). In contrast NT-proBNP values remained unchanged in UAP (182 (74-410) pg/ml) vs 171 (53-474) pg/ml). CONCLUSION: The time interval from onset of symptoms to first blood collection is an important determinant for NT-proBNP values on admission in patients with an ACS and needs to be considered in clinical practice.

Left ventricular adaptation after atrial septal defect closure assessed by increased concentrations of N-terminal pro-brain natriuretic peptide and cardiac magnetic resonance imaging in adult patients.

OBJECTIVE: To determine in an observational study whether N-terminal pro-brain natriuretic peptide (NT-proBNP) is raised in patients with an atrial septal defect (ASD) and whether concentrations change after interventional closure. METHODS: 12 patients (6 men, mean (SD) age 44.4 (18.6) years) with a moderate sized ASD type II (23.3 (4.5) mm, pulmonary to systemic flow ratio 2.1 (0.68)) were investigated. In all patients a magnetic resonance imaging (MRI) study was performed and NT-proBNP was assessed at baseline and early (9 (13) days) and late (138 (64) days) after intervention. RESULTS: Concentrations of NT-proBNP were found to be within the normal range at baseline (median 87 pg/ml, interquartile range 65-181 pg/ml) but increased early after the interventional closure (315 pg/ml, 133-384 pg/ml, p = 0.005 versus baseline). The increase of NT-proBNP was associated with an increase in left ventricular dimensions as assessed by MRI (left ventricular end diastolic volume 104 (27) ml to 118 (27) ml, p = 0.003). Late after ASD closure NT-proBNP returned to baseline concentrations (102 pg/ml, 82-188 pg/ml, p = 0.004 versus early follow up). CONCLUSION: These findings suggest the presence of transitory haemodynamic stress during adaptation of the left ventricle after ASD closure, which may contribute to the understanding of the pathological mechanism of acute heart failure and delayed improvement of exercise capacity after ASD closure.