Locus 'decontraction' and centromeric recruitment contribute to allelic exclusion of the immunoglobulin heavy-chain gene

Allelic exclusion of immunoglobulin genes ensures the expression of a single antibody molecule in B cells through mostly unknown mechanisms. Large-scale contraction of the immunoglobulin heavy-chain (Igh) locus facilitates rearrangements between Igh variable (V(H)) and diversity gene segments in pro-B cells. Here we show that these long-range interactions are mediated by 'looping' of individual Igh subdomains. The Igk locus also underwent contraction by looping in small pre-B and immature B cells, demonstrating that immunoglobulin loci are in a contracted state in rearranging cells. Successful Igh recombination induced the rapid reversal of locus contraction in response to pre-B cell receptor signaling, which physically separated the distal V(H) genes from the proximal Igh domain, thus preventing further rearrangements. In the absence of locus contraction, only the four most proximal V(H) genes escaped allelic exclusion in immature mu-transgenic B lymphocytes. Pre-B cell receptor signaling also led to rapid repositioning of one Igh allele to repressive centromeric domains in response to downregulation of interleukin 7 signaling. These data link both locus 'decontraction' and centromeric recruitment to the establishment of allelic exclusion at the Igh locus.     

Evaluation of sodium T1 relaxation times in human heart

PURPOSE: To evaluate the sodium longitudinal relaxation (T(1)) characteristics for myocardium and blood in humans. MATERIALS AND METHODS: Eleven healthy volunteers were examined by using a (23)Na heart surface coil at a 1.5 T clinical scanner equipped with a broadband spectroscopy option. (23)Na MR measurements were performed by using a three-dimensional spoiled gradient echo sequence (in-plane resolution, 3.5 mm x 7 mm; slice thickness, 24 mm; TE, 3.1 msec; bandwidth, 65 Hz/pixel; TR, 21 to 150 msec). RESULTS: Longitudinal T(1) relaxation time components were 31.6+/-7.0 msec and 31.1+/-7.5 msec for myocardium and blood, respectively. CONCLUSION: (23)Na T(1) relaxation times of myocardium and blood can be determined in humans. The results are in agreement with values obtained from animal studies.     

Cardiac and skeletal muscle involvement in myotonic dystrophy type 2 (DM2): a quantitative 31P-MRS and MRI study

In myotonic dystrophy type 2 (DM2/PROMM), cardiac muscle involvement is usually more benign than in DM1, but clinically severe cardiomyopathy has been reported in some patients. Using a novel method of magnetic resonance spectroscopy (MRS), we examined the left ventricular myocardium and the left gastrocnemius muscle in 11 unselected DM2/PROMM patients without overt cardiac disease. Data on cardiac morphology and function were obtained by gradient echo two-dimensional cine magnetic resonance imaging (MRI); no significant differences were found between DM2 patients and healthy controls, but using a median split approach older patients showed mildly increased left ventricular (LV) volumes, i.e., 59% increase of end-systolic volume index (ESVI) and 35% increase of end-diastolic volume index (EDVI), and an increase of LV mass (26%). On cardiac MRS, DM2/PROMM patients showed a reduction of phosphocreatine (PCr) and adenosine triphosphate (ATP) by 25 and 20% compared to matched healthy controls. No significant differences were found between younger and older patients. In skeletal muscle of the DM2 patients, no significant decrease of PCr and ATP concentrations was found. However, in older patients, who commonly show overt hip flexor muscle weakness, we observed reduced values for PCr and ATP. Our MRS and MRI findings reveal evidence for subclinical cardiomyopathy in DM2/PROMM patients without overt heart disease. Future prospective studies are needed to clarify the risk of developing overt cardiac disease in DM2 and to define prognostic factors.     

Impact of point-of-care testing for CYP2C19 on platelet inhibition in patients with acute coronary syndrome and early dual antiplatelet therapy in the emergency setting

Aims

Only limited data exist about the role of point of care CYP2C19 testing in the acute setting in the early phase of acute coronary syndromes (ACS). Therefore, the present study was designed to investigate the impact of CYP2C19 loss-of–function point-of-care (POC) genotyping in patients presenting with acute coronary syndromes (ACS) and treated with dual antiplatelet therapy in the emergency setting.

Methods and Results

137 subjects with ACS scheduled for percutaneous coronary intervention were consecutively enrolled. Pre- and on-treatment platelet aggregation was assessed by multiple electrode aggregometry (MEA) after stimulation with adenosine diphosphate (ADP). Patients were loaded according to current guideline adherent indications and contraindications for use of P2Y₁₂ inhibitors in ACS. POC genotyping for CYP2C19*2 was performed in the emergency room after obtaining a buccal swab using the Spartan RX CYP2C19 system and obtaining patient’s informed consent. Prasugrel and ticagrelor treated patients had significantly lower PR compared to clopidogrel-treated patients. The benefits of prasugrel and ticagrelor compared to clopidogrel treated patients in terms of platelet inhibition were more pronounced in CYP2C19*2 carriers. Non-carriers showed similar inhibition regardless of particular P2Y₁₂ inhibitor treatment. Statistical analyses adjusting for factors associated with response (e.g. smoking) revealed that CYP2C19*2 allele carrier status and loading with different type of P2Y12 receptor blockers were significant predictors of on-treatment platelet reactivity in the early phase of ACS.

Conclusion

The results of this pilot study of treatment of patients in the early phase of ACS indicate that CYP2C19*2 POC genotyping might help to identify patients at risk with poor response to clopidogrel treatment, thereby benefiting from reloading and switching to alternative P2Y₁₂ receptor inhibition.     

Auto-SENSE perfusion imaging of the whole human heart

PURPOSE: To show the application of auto-sensitivity encoding (SENSE)-a self-calibrating parallel imaging technique-to first pass perfusion imaging of the whole human heart. MATERIALS AND METHODS: The self-calibrating parallel imaging method auto-SENSE was implemented for a saturation recovery turbo-fast low-angle shot (FLASH) sequence on a 1.5-T scanner using a standard four-element body phased array coil. By reducing the acquisition time per slice by a factor of two compared to conventional turbo FLASH imaging, the number of imaged slices could be doubled to six to ten with an unchanged temporal resolution of one image per heartbeat. This technique has been tested in eight healthy volunteers for contrast-enhanced heart perfusion imaging. RESULTS: Auto-SENSE heart perfusion imaging with improved coverage of the human heart could be performed successfully in all volunteers. A first quantitative comparison of perfusion values between the auto-SENSE and the non-SENSE techniques shows good agreement. CONCLUSION: Auto-SENSE allows perfusion imaging of the whole human heart without gaps.     

Molecular and functional characterization of the extracellular calcium-sensing receptor in human colon cancer cells

Presence of a functional extracellular calcium-sensing receptor (CaR) is of particular relevance for the growth-inhibitory action of Ca2+ on human colon carcinoma cells. In order to detect CaR gene alterations that may have occurred during the tumorigenic process, we applied Southern blot, DNA sequence, and RT-PCR analysis to DNA from normal human colon mucosa and from cancerous lesions of different grading, as well as from primary cultured and established colonic carcinoma cell lines (e.g., Caco-2). No evidence was obtained for mutations or other sequence alterations in the CaR gene in any of the colon carcinoma cells analyzed. Only a differential expression of two splice variants of the CaR gene, which are generated by usage of different promoters in the 5'-untranslated region, was detected in colon carcinomas of different grade. From Western blot analysis a tendency towards lower CaR protein levels in carcinoma cells in parallel with tumor progression became apparent. Activation of the CaR by extracellular Ca2+ or by specific receptor agonists resulted in substantial growth inhibition in Caco-2 cells. Activation of the CaR was transduced into inhibition of phospholipase A2-mediated arachidonic acid formation, but also into increased production of cAMP and IP3. This provides evidence for a cell type-specific function of the CaR in human colonocytes. We conclude that neoplastic colon epithelial cells can respond to antimitogenic signals generated by activation of the CaR as long as they express sufficient amounts of the CaR protein. This provides a rationale for the use of calcium in chemoprevention of colon tumor development.     

Adenosine stress cardiac magnetic resonance imaging for the assessment of ischemic heart disease

Aims  This prospective study was designed to determine the diagnostic value of adenosine stress cardiac magnetic resonance imaging (CMRI) in patients referred to elective coronary angiography. Methods and results  Myocardial perfusion measurements at rest and adenosine stress were performed in 141 patients (105 men, 36 women, mean age 63.4 years) at 1.5 T with a Turbo Flash sequence. Stress-induced perfusion deficits were correlated to angiographic stenoses ≥75%. The overall sensitivity for CMRI depicting coronary artery disease (CAD) with relevant stenoses was 90.4%, the specificity was 77.4%, the positive predictive value was 85.9%, the negative predictive value was 84.2% and the accuracy 85.2%. Subgroup analysis was performed for 3-vessel disease (n = 44, sensitivity 92.3%, specificity 75.0%), 2-vessel disease (n = 43, sensitivity 92.6%, specificity 92.9%), 1-vessel disease (n = 27, sensitivity 93.1%, specificity 71.4%) and patients without CAD (n = 27, specificity 70.4%) as well as for patients with prior myocardial infarction (n = 44, sensitivity 92.9%, specificity 86.7%), prior coronary artery bypass surgery (n = 21, sensitivity 88.2%, specificity 66.7%), prior coronary interventions (n = 88, sensitivity 91.9%, specificity 75.0%), or diabetics (n = 27, sensitivity 90.5%, specificity 83.3%). Conclusion  Our study shows that stress perfusion CMRI can accurately predict relevant CAD and contributes to the identification of hemodynamic relevant stenoses in patients scheduled for coronary angiography. C. Doesch and A. Seeger have equally contributed to this publication.