The cytoskeleton is the physical and biochemical interface for a large variety of cellular processes. Its complex regulation machinery is involved upstream and downstream in various signaling pathways. The cytoskeleton determines the mechanical properties of a cell. Thus, cell elasticity could serve as a parameter reflecting the behavior of the system rather than reflecting the specific properties of isolated components. In this study, we used atomic force microscopy to perform real-time monitoring of cell elasticity unveiling cytoskeletal dynamics of living bronchial epithelial cells. In resting cells, we found a periodic activity of the cytoskeleton. Amplitude and frequency of this spontaneous oscillation were strongly affected by intracellular calcium. Experiments reveal that basal cell elasticity and superimposed elasticity oscillations are caused by the collective action of myosin motor proteins. We characterized the cell as a mechanically multilayered structure, and followed cytoskeletal dynamics in the different layers with high time resolution. In conclusion, the collective activities of the myosin motor proteins define overall mechanical cell dynamics, reflecting specific changes of the chemical and mechanical environment.

Abstract-Elevation of C-reactive protein (CRP) in human blood accompanies inflammatory processes, including cardiovascular diseases. There is increasing evidence that the acute-phase reactant CRP is not only a passive marker protein for systemic inflammation but also affects the vascular system. Further, CRP is an independent risk factor for atherosclerosis and the development of hypertension. Another crucial player in atherosclerotic processes is the mineralocorticoid hormone aldosterone. Even in low physiological concentrations, it stimulates the expression and membrane insertion of the epithelial sodium channel, thereby increasing the mechanical stiffness of endothelial cells. This contributes to the progression of endothelial dysfunction. In the present study, the hypothesis was tested that the acute application of CRP (25 mg/L), in presence of aldosterone (0.5 nmol/L; 24 hour incubation), modifies the mechanical stiffness and permeability of the endothelium. We found that endothelial cells stiffen in response to CRP. In parallel, endothelial epithelial sodium channel is inserted into the plasma membrane, while, surprisingly, the endothelial permeability decreases. CRP actions are prevented either by the inhibition of the intracellular aldosterone receptors using spironolactone (5 nmol/L) or by the inactivation of epithelial sodium channel using specific blockers. In contrast, inhibition of the release of the vasodilating gas nitric oxide via blockade of the phosphoinositide 3-kinase/Akt pathway has no effect on the CRP-induced stiffening of endothelial cells. The data indicate that CRP enhances the effects of aldosterone on the mechanical properties of the endothelium. Thus, CRP could counteract any decrease in arterial blood pressure that accompanies severe acute inflammatory processes. (Hypertension. 2011; 57:231-237.)Key Words: aldosterone Ⅲ CRP Ⅲ ENaC Ⅲ AFM Ⅲ immunofluorescence Ⅲ PI3K Ⅲ NO C -Reactive protein (CRP) is considered to be a stable and powerful inflammatory marker of future cardiovascular risk 1 and, as an acute-phase reactant, originally considered to be a mere marker of vascular inflammation. However, CRP may also participate directly in the inflammatory process. 2 During inflammation and sepsis, the production of NO is increased, which leads to vasodilation and thus to a drop in blood pressure. 3,4 Recently, CRP was shown to decrease endothelial NO synthase expression 5 via inhibition of the phosphoinositide 3-kinase (PI3K)/Akt signaling pathway, 6 which suggests that it also has a role in endothelial (dys)function. In clinical studies, it has been shown that in patients with essential hypertension, there is a positive correlation between CRP levels and pulse wave velocity, which is a functional indicator for arterial stiffness. 7,8 When the paradigm shift occurred that CRP was not merely a "reporting" but also an "acting" protein, another shift in understanding took place, namely that the mineralocorticoid hormone aldosterone not only acts on the kidney but also on the cardiovascular sys...

In normal airway epithelium, the cystic fibrosis transmembrane conductance regulator (CFTR) transports Cl^- ions to the apical surface of the epithelium paralleled by the flow of water through transcellular and paracellular pathways. The hypothesis was tested whether CFTR not only regulates the transcellular but also the paracellular shunt pathway. Therefore, we performed measurements of transepithelial electrical resistance (TER) and paracellular ¹⁴C-mannitol permeability in wtCFTR (16HBE14o^-) and delF508-CFTR (CFBE41o^-) expressing human bronchial epithelial cells. Under resting conditions, CFBE41o^- cell monolayers exhibit a higher paracellular permeability and lower TER as compared to 16HBE14o^- monolayers. Stimulation of CFTR by cAMP induces opposite effects in the two cell lines. 16HBE14o^- monolayers show a sharp decrease of TER, in parallel with a concomitant increase of paracellular permeability. The change in paracellular permeability is mediated by a myosin II dependent mechanism because it can be blocked by the myosin light chain kinase inhibitor ML-7. In contrast, CFBE41o^- cells respond to cAMP stimulation with a decrease of paracellular permeability, paralleled by slight increase of TER. We conclude that stimulation of wtCFTR increases vectorial transcellular salt transport and, simultaneously, the paracellular permeability allowing water to follow through the paracellular pathway. In contrast, in CF epithelium cAMP stimulation increases neither vectorial salt transport nor paracellular permeability which is likely to contribute to the CF pulmonary phenotype. Taken together, our results link CFTR dysfunction to an improper regulation of the paracellular transport route.

Suprapubic drainage in robot-assisted radical prostatectomy shows significantly decreased pain levels during the catheterization period compared to the transurethral diversion without compromising long-term functional results. Intraoperative placement of a suprapubic tube should be discussed as a standard procedure for further improvement of patients' postoperative comfort.

It has been reported recently that the cystic fibrosis transmembrane conductance regulator (CFTR) besides transcellular chloride transport, also controls the paracellular permeability of bronchial epithelium. The aim of this study was to test whether overexpressing wtCFTR solely regulates paracellular permeability of cell monolayers. To answer this question we used a CFBE41o– cell line transfected with wtCFTR or mutant F508del-CFTR and compered them with parental line and healthy 16HBE14o– cells. Transepithelial electrical resistance (TER) and paracellular fluorescein flux were measured under control and CFTR-stimulating conditions. CFTR stimulation significant decreased TER in 16HBE14o– and also in CFBE41o– cells transfected with wtCFTR. In contrast, TER increased upon stimulation in CFBE41o– cells and CFBE41o– cells transfected with F508del-CFTR. Under non-stimulated conditions, all four cell lines had similar paracellular fluorescein flux. Stimulation increased only the paracellular permeability of the 16HBE14o– cell monolayers. We observed that 16HBE14o– cells were significantly smaller and showed a different structure of cell-cell contacts than CFBE41o– and its overexpressing clones. Consequently, 16HBE14o– cells have about 80% more cell-cell contacts through which electrical current and solutes can leak. Also tight junction protein composition is different in ‘healthy’ 16HBE14o– cells compared to ‘cystic fibrosis’ CFBE41o– cells. We found that claudin-3 expression was considerably stronger in 16HBE14o– cells than in the three CFBE41o– cell clones and thus independent of the presence of functional CFTR. Together, CFBE41o– cell line transfection with wtCFTR modifies transcellular conductance, but not the paracellular permeability. We conclude that CFTR overexpression is not sufficient to fully reconstitute transport in CF bronchial epithelium. Hence, it is not recommended to use those cell lines to study CFTR-dependent epithelial transport.

While ICG-ePLND seems to be beneficial for a better understanding of the lymphatic drainage and a more meticulous diagnostic approach, the sensitivity is not sufficient to recommend stand-alone ICG lymph node dissection.

The aim of this study was to assess clinically meaningful differences of preoperative lower urinary tract symptoms (LUTS) and quality of life (QoL) before and after robot-assisted radical prostatectomy (RARP). Therefore we identified 5506 RARP patients from 2007 to 2018 with completed International Prostate Symptom Score (IPSS) and -QoL questionnaires before and 12 months after RARP in our institution. Marked clinically important difference (MCID) was defined by using the strictest IPSS-difference of − 8 points. Multivariable logistic regression analyses (LRM) aimed to predict ∆IPSS ≤ − 8 and were restricted to RARP patients with preoperatively moderate (IPSS 8–19) vs. severe (IPSS 20–35) LUTS burden (n = 2305). Preoperative LUTS was categorized as moderate and severe in 37% (n = 2014) and 5.3% of the complete cohort (n = 291), respectively. Here, a postoperative ∆IPSS ≤ − 8, was reported in 38% vs. 90%. In LRM, younger age (OR 0.98, 95%CI 0.97–0.99; p = 0.007), lower BMI (OR 0.94, 95%CI 0.92–0.97; p < 0.001), higher preoperative LUTS burden (severe vs. moderate [REF.] OR 15.6, 95%CI 10.4–23.4; p < 0.001), greater prostate specimen weight (per 10 g, OR 1.12, 95%CI 1.07–1.16; p < 0.001) and the event of urinary continence recovery (OR 1.66 95%CI 1.25–2.21; p < 0.001) were independent predictors of a marked LUTS improvement after RARP. Less rigorous IPSS-difference of − 5 points yielded identical predictors. To sum up, in substantial proportions of patients with preoperative moderate or severe LUTS a marked improvement of LUTS and QoL can be expected at 12 months after RARP. LRM revealed greatest benefit in those patients with preoperatively greatest LUTS burden, prostate enlargement, lower BMI, younger age and the event of urinary continence recovery.

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