Quantitative imaging of focal adhesion dynamics and their regulation by HGF and Rap1 signaling.

Cell migration is crucial in development, tissue repair and immunity and frequently aberrant in pathological processes including tumor metastasis. Focal adhesions (FAs) are integrin-based adhesion complexes that form the link between the cytoskeleton and the extracellular matrix and are thought to orchestrate cell migration. Understanding the regulation of FAs by (oncogenic) signaling pathways may identify strategies to target pathological cell migration. Here we describe the development of a robust FA tracker that enables the automatic, multi-parametric analysis of FA dynamics, morphology and composition from time-lapse image series generated by total internal reflection fluorescence (TIRF) microscopy. In control prostate carcinoma cells, this software recapitulates previous findings that relate morphological characteristics of FAs to their lifetime and their cellular location. We then investigated how FAs are altered when cell migration is induced by the metastasis-promoting hormone HGF and subsequently inhibited by activation of the small GTPase Rap1. We performed a detailed analysis of individual FA parameters, which identified FA size, sliding and intensity as primary targets of Rap1. HGF did not have strong effects on any of the FA parameters within the first hours of its addition. Subsequent Bayesian network inference (BNI), using all measured parameters as input, revealed little correlation between changes in cell migration and FA characteristics in this prostate carcinoma cell line. Instead BNI indicated a concerted coordination of cell size and FA parameters. Thus our results did not reveal a direct relation between the regulation of cell migration and the regulation of FA dynamics.

Medical attention seeking by suspected stroke patients: Emergency medical services or general practitioner?

OBJECTIVE: Awareness campaigns advise the public to call emergency medical services (EMS) directly in case of suspected stroke. We aimed to explore patient and notification characteristics that influence direct EMS notification, the time to alert, and the time to treatment. METHODS: We performed a secondary analysis with data from the PRESTO study, a multi-center prospective observational cohort study that included patients with suspected stroke. We used multivariable binary logistic regression analyses to assess the association with direct EMS notification and multivariable linear regression analyses to assess the association with the onset-to-alert time, onset-to-needle time and onset-to-groin time. RESULTS: Of 436 included patients, 208 patients (48%) contacted EMS directly. FAST scores (aOR 1.45 for every point increase, 95%CI: 1.14-1.86), alert outside office hours (aOR 1.64 [1.05-2.55]), and onset-to-alert time (aOR for every minute less [≤55 min]: 0.96 [0.95-0.97]) were independently associated with direct EMS notification. Direct EMS call was independently associated with shorter onset-to-alert times (27 min [54-0.84]) and with shorter onset-to-needle times (-30 min [-51 to -10]). The association between direct EMS call and the onset-to-groin time was almost similar to the association with onset-to-needle time, though not statistically significant (univariable analysis: 23.7 min decrease [-103.7 to 56.2]). CONCLUSION: More than half of all patients with suspected stroke do not call EMS directly but call their GP instead. Patients with higher FAST scores, alert outside office hours, and a rapid alert, more often call EMS directly. Patients who call EMS directly are treated with IVT 30 min faster than patients who call the GP first. TRIAL REGISTRATION NUMBER: Netherlands Trial Register: NL7387, (www.trialregister.nl).

Troponin I release at rest and after exercise in patients with hypertrophic cardiomyopathy and the effect of betablockade.

PURPOSE OF THE WORK: In patients with hypertrophic cardiomyopathy ischemia may occur due to massive heart weight, myocyte disarray or small vessel disease. We detected elevated troponin levels in some of these patients and hypothesized that troponin release would rise after exercise and diminish after betablockade. METHODS AND RESULTS: In 5 of 7 young patients (6 males) with hypertrophic cardiomyopathy and no overt coronary artery disease we found elevated troponin levels after physical exercise; the peak was between 6 and 9 hours and levels returned to pre-exercise values within 24 hours. Troponin release was consistently diminished after use of a betablocker. CONCLUSIONS: Increased troponin release may be present in patients with hypertrophic cardiomyopathy and is temporarily enhanced by exercise and diminishes with betablockade.

[Idiopathic ventricular tachycardia, an arrhythmia with good prognosis]. / Idiopathische ventrikeltachycardie, een ritmestoornis met gunstige prognose.

Three patients, one experiencing palpitations and two complaining of chest pain in stressful situations, appeared to have monomorphic wide complex tachycardia. After excluding channelopathy, structural abnormalities and ischaemia of the heart, this arrhythmia was classified as idiopathic. Symptoms disappeared in one patient after using metoprolol, a ß-adrenoceptor blocker. The other two patients were treated with radiofrequency ablation of the focus from which the tachycardias arose. Idiopathic ventricular tachycardia mostly arises from the right ventricular outflow tract. The diagnosis is made upon history taking, including family history, echocardiography, 12-lead ECG, exercise testing and 24-hour Holter monitoring. The prognosis is excellent and sudden cardiac death is rarely seen.

The Rap1 guanine nucleotide exchange factor C3G is required for preservation of larval muscle integrity in Drosophila melanogaster.

C3G is a guanine nucleotide exchange factor (GEF) and modulator of small G-protein activity, which primarily acts on members of the Rap GTPase subfamily. Via promotion of the active GTP bound conformation of target GTPases, C3G has been implicated in the regulation of multiple cellular and developmental events including proliferation, differentiation and apoptosis. The Drosophila C3G orthologue exhibits a domain organization similar to that of vertebrate C3G. Through deletion of the C3G locus, we have observed that loss of C3G causes semi-lethality, and that escaping adult flies are characterized by a reduction in lifespan and general fitness. In situ hybridization reveals C3G expression in the developing embryonic somatic and visceral muscles, and indeed analysis of C3G mutants suggests essential functions of C3G for normal body wall muscle development during larval stages. C3G mutants display abnormal muscle morphology and attachment, as well as failure to properly localize betaPS integrins to muscle attachment sites. Moreover, we show that C3G stimulates guanine nucleotide exchange on Drosophila Rap GTPases in vitro. Taken together, we conclude that Drosophila C3G is a Rap1-specific GEF with important functions in maintaining muscle integrity during larval stages.

Dual site coronary sinus pacing in a patient with an artificial tricuspid valve prosthesis.

For safety reasons, two leads for left ventricular pacing were implanted in the coronary sinus of a pacemaker dependent patient with an artificial tricuspid valve prosthesis.

Troponin I release at rest and after exercise in patients with hypertrophic cardiomyopathy and the effect of betablockade

PURPOSE OF THE WORK: In patients with hypertrophic cardiomyopathy ischemia may occur due to massive heart weight, myocyte disarray or small vessel disease. We detected elevated troponin levels in some of these patients and hypothesized that troponin release would rise after exercise and diminish after betablockade. METHODS AND RESULTS: In 5 of 7 young patients (6 males) with hypertrophic cardiomyopathy and no overt coronary artery disease we found elevated troponin levels after physical exercise; the peak was between 6 and 9 hours and levels returned to pre-exercise values within 24 hours. Troponin release was consistently diminished after use of a betablocker. CONCLUSIONS: Increased troponin release may be present in patients with hypertrophic cardiomyopathy and is temporarily enhanced by exercise and diminishes with betablockade.