NorthernLights in slide-based cytometry and microscopy.

In recent years, slide-based cytometry has become a key technology for polychromatic cytometric investigations, and many efforts have been made to increase the number of measurable fluorochromes for multiparametric analysis. Sequential photobleaching of fluorochromes next to very photostable dyes is one approach for this technology. As the ALEXA dyes are known to be photostable as compared to the conventional fluorochromes FITC, PE (Riggs et al., Am J Pathol 1958;34:1081-1097), and APC, a differentiation within a fluorochrome pair is possible. Here, we have analyzed the newly available NorthernLights secondary antibodies for use in slide-based cytometry and microscopy. Currently, these fluorochrome-conjugates are now available with three distinct excitation- and emission maxima (NL493, NL557, NL637). Their spectral properties are similar to the frequently used fluorochromes FITC, PE, and APC and can, therefore, be used with most common excitation sources of cytometers or microscopes. As the NorthernLights are bright, resistant to photobleaching, stable in alcohols and xylene and of affordable price, these dyes are promising candidates for use with most laser- and HBO/XBA-based fluorescence microscopy-like techniques.

Characterization of fibroblasts responsible for cartilage destruction in arthritis.

In the pathogenesis of rheumatoid arthritis (RA), synovial fibroblasts (SF) play a key role as they secrete distinct patterns of cytokines and express variable levels of costimulatory and adhesion molecules. The murine fibroblast cell line LS48 has been shown to be invasive in the cartilage destruction models in vivo and in vitro. The purpose of this study was to examine in detail the LS48 phenotype, to obtain a better understanding of the SF-mediated cartilage destruction in RA. The destructive fibroblasts line LS48 and the nondestructive 3T3 cells were cultured and characterized with slide-based and flow cytometry, using antibodies against several adhesion molecules, immunological acting molecules, and marker proteins. The invasive LS48 fibroblasts are characterized by significantly higher expression of adhesion molecules such as CD47 (IAP), CD51 (integrin alpha V), CD61 (GPIIIa), and CD147 (EMMPRIN), and immunological acting molecules such as CD40 (Bp50), CD55 (DAF), and TLR-2. The results from the slide-based and flow cytometry analyses were exactly the same, except for the selected CD147 and TLR-2. This study demonstrated that the destructive fibroblast cell line LS48 has the characteristics of RA SFs. The high expression of specific costimulatory and adhesion molecules underlines the aberrant phenotype of these cells when compared with noninvasive fibroblasts. Furthermore, slide-based and flow cytometry complement each other in fibroblast phenotyping. Overall, this study shows that LS48 is an excellent tool to gain a deeper understanding of SF in RA.

Implantation of CARILLON® Mitral Contour System with transvenous left ventricular lead in place.

BACKGROUND: Cardiac resynchronisation therapy (CRT) is an established treatment option for chronic heart failure patients with left bundle branch block. Although a concomitant functional mitral regurgitation is often reduced by CRT, many patients need additional mitral valve repair. Placing a CARILLON® Mitral Contour System (CMCS) over a transvenous CRT lead is currently not recommended, since both of them are implanted in the coronary sinus (CS). The aim of this study was to investigate the feasibility of sequential implantation of a transvenous LV lead followed by CMCS implantation, and to assess LV lead performance and possibility of extraction. METHODS AND RESULTS: Standard transvenous LV leads were implanted in the CS of five female sheep. After establishing regular anatomical position with stable electrical parameters of the LV lead, a CMCS was additionally implanted in the CS. After an observation period of 100 days, lead performance and positions of lead and CMCS were studied. Sequential implantation of the two components was feasible in sheep. After 100 days, all leads showed regular measurements of impedance, threshold, and sensing. There was no migration of either the LV lead or the CMCS. In all cases, the LV lead could be completely extracted without migration of the CMCS. There were no acute or long-term complications. CONCLUSIONS: In an animal model of healthy adult sheep, implantation of CMCS with a transvenous LV lead already in place was feasible and without major problems with either the CMCS or the LV lead. Electrical performance of the LV leads was excellent. All LV leads could be extracted without migration of the CMCS.

The importance of right atrial pacing electrode position and pacing configuration for intra-atrial and inter-atrial conduction times.

Pace prevention of atrial tachyarrhythmias is based in part on the reduction of intra-atrial (IAA) and/or inter-atrial (IEA) conduction. We previously introduced a novel pacing mode using floating atrial ring electrodes on a VDD-lead (BIdirectional MO nophasic impulSe: BIMOS). The effects of BIMOS pacing on IAA and IEA conduction times has not been studied. In nine Merino sheep electrode catheters were placed at the His-Bundle (HBE), high right atrium (HRA), coronary sinus ostium (Cs-Os), and left lateral atrium (LLA). A VDD-lead was introduced with floating electrodes in the high and mid right atrium (Floating). IAA (S/P-HRA, S/P-Cs-Os, S/P-HBE, S/P-Floating), IEA conduction times (S/P-LLA), and P-wave duration (PD) were measured during sinus rhythm (S), during bipolar cathodal pacing (P) in the HRA, in the Cs-Os position, as well as during BIMOS floating pacing. The mean PD during S was significantly shorter than during HRA- (66. 6+/-12.8ms; vs. 116.2+/-11.1ms; p<0.05) and Cs-Os-P (66.6+/-12.8ms vs. 94.4+/-9.0ms; p<0.05). In comparison to HRA-P, BIMOS configuration lead to a significant reduction of the P-wave duration (116.2+/-11.1ms vs. 85. 4+/-8.8ms; p<0.05). During BIMOS pacing, the global atrial conduction time was significantly shorter than during pacing in the HRA and Cs-Os position. The results of this study demonstrate a clear reduction of IAA and IEA conduction times using BIMOS configurations compared to conventional HRA-P. Furthermore, BIMOS pacing produced a more homogeneous atrial activation when compared with conventional HRA- and Cs-Os-P.