In vivo and ex vivo measurements: noninvasive assessment of alcoholic fatty liver using 1H-MR spectroscopy

PURPOSEWe aimed to evaluate the ability of 1H-magnetic resonance spectroscopy (1H-MRS) to detect and quantify hepatic fat content in vivo and ex vivo in an experimental rat model of alcoholic fatty liver using histopathology, biochemistry, and laboratory analyses as reference.METHODSAlcoholic fatty liver was induced within 48 hours in 20 Lewis rats; 10 rats served as control. Intrahepatic fat content determined by 1H-MRS was expressed as the percent ratio of the lipid and water peaks and was correlated with intrahepatic fat content determined histologically and biochemically. Liver enzymes were measured in serum.RESULTSFatty liver could be detected in vivo as well as ex vivo using 1H-MRS, in all 20 animals. Histologic analysis showed a fatty liver in 16 of 20 animals. Histology and 1H-MRS results were highly correlated (in vivo, r=0.93, P = 0.0005; ex vivo, r=0.92, P = 0.0006). Also a strong correlation was noted between in vivo 1H-MRS measurements and the fat content determined biochemically (r=0.96, P = 0.0003). Ex vivo results showed a similarly strong correlation between 1H-MRS and biochemistry (r=0.89, P = 0.0011).CONCLUSION1H-MRS can be carried out in ex vivo models, as well as in vivo, to detect and quantify intrahepatic fat content in the acute fatty liver

Influence of myocardial hypertrophy and neurohumoral factors on contraction capability of the isolated ventricular heart muscle cells

In der vorliegenden Arbeit wurde auf zellulärer Ebene die kausalverknüpfung von Hypertrophie und Kontraktion belegt. Dazu wurden in einem ersten experimentellen Modell adulte ventrikuläre Herzmuskelzellen aus der Ratte isoliert und für 24 h mit Phenylephrin inkubiert. Phenylephrin hat eine starke hypertrophe Wirkung auf Herzmuskelzellen. Es steigert die Proteinsynthese durch Aktivierung der Proteinkinase C und des PI 3-Kinase/Akt-Signalweges. Daneben kommt es durch Stimulation mitogen aktivierter Kinasen (p42-MAP) zu phänotypischen Veränderungen. In der vorliegenden Arbeit wurde der Einfluss einer chronischen Stimulation alpha-adrenerger Rezeptoren auf das Kontraktionsverhalten adulter Herzmuskelzellen der Ratte und der Beitrag der oben genannten Signaltransduktionswege durch spezifische pharmakologische Hemmung untersucht. Eine signifikante kontraktile Dysfunktion der hypertrophierten Herzmuskelzellen wurde sichtbar, obwohl diese eine erhöhte Relaxationsgeschwindigkeit zeigen. Die Übertragbarkeit dieser Befunde auf Herzmuskelzellen aus hypertrophierten Herzen wurde nachfoglend analysiert. Dazu wurden drei verschiedene Modelle verwendet. Zunächst wurden Herzmuskelzellen aus transgenen Mäusen isoliert, die eine myokardiale Expression einer konstitutiv aktivierten Akt besitzen. Dieses Modell entspricht einer Hypertrophie ohne erkennbare phänotypischen Veränderungen, wie sie durch MAP-Kinasen induziert werden. Als zweites wurden Kardiomyozyten aus spontan hypertensiven Ratten isoliert. Diese Tiere zeigen in vivo eine druckinduzierte Myokardhypertrophie, wahrscheinlich basierend auf einer Aktivierung sowohl des sympathischen Nervensystems als auch des Renin-Angiotensin-Systems. Phänotypische Veränderungen sind hier wahrscheinlich. Diese Herzmuskelzellen zeigten einen kontraktile Dysfunktion, welche nicht durch schnellere Relaxations- und Kontraktionsgeschwindigkeit ausgeglichen werden konnte, während bei den Akt exprimierenden Herzmuskelzellen ein Funktionsausgleich stattfand. Als drittes wurden Kardiomyozyten aus transgenen Mäusen isoliert, welche TGF-beta überexprimieren. Diese Tiere haben eine moderate Myokardhypertrohie aber keine Druckbelastung. TGF-beta wird dabei als Endpunkt einer Angiotensin II –Wirkung verstanden, welche anschließend durch eine veränderte Ankopplung der beta-adrenergen Rezeptoren zu einer Myokardhypertrophie beiträgt. In dem hier verwendeten Modell konnte dieser Teil der postulierten Kausalkette (Angiotensin II – TGF-beta – beta-adrenerge Rezeptoren) von den Eigeneffekten des Angiotensin II entkoppelt werden. Dies wurde im weiteren belegt durch eine Anzahl an Experimenten, in denen auf jeder der drei Ebenen der Kausalkette in den transgenen Tieren eingegriffen wurde. Deshalb wurden die TGF-beta überexprierenden Mäuse mit Telmisartan (einem AT1-Rezeptoranagonisten), TGF-beta-Antikörper und beta-Blockern behandelt und nachfolgend auf ihre beta-adrenerge Ansprechbarkeit untersucht. Die Untersuchungen belegen die Bedeutung von TGF-beta für die verminderte Ansprechbarkeit beta-adrenerger Rezeptoren im Rahmen einer druckinduzierten Myokardhypertrophie. Sie zeigen erstmals, das die Gabe von beta-Blockern TGF-beta induzierte funktionelle Veränderungen auf zellulärer Ebene korrigieren kann.The following paper demonstrates the causal link between hypertrophy and contraction at the cellular level. In a first experimental model, adult ventricular rat cardiac muscle cells were isolated, and incubated for 24 hours with phenylephrine. Phenylephrine has a strong hypertrophic effect on cardiac muscle cells. It increases the rate of protein synthesis by activating protein kinase C and the PI 3 kinase / Akt signaling pathway. In addition, it causes phenotypic changes through stimulation of mitogen-activated kinases (p42 MAP). This paper examines the effect of chronic stimulation of alpha-adrenergic receptors on the contractile behaviour of adult rat cardiac muscle cells and examines the contribution made by the above signal transduction pathway through the use of specific pharmacological inhibition. Significant contractile dysfunction of the hypertrophied cardiac muscle cells was observed, although the cells exhibited an increased relaxation speed. The transferability of these findings to cardiac muscle cells from hypertrophied hearts was then analysed using three different models. Firstly, cardiac muscle cells from transgenic mice expressing constitutively active Akt kinase in the myocard were isolated. This model is equivalent to hypertrophy without the recognisable phenotypic changes induced by MAP kinases. Secondly cardiomyocytes from spontaneously hypertensive rats were isolated. These animals in vivo show pressure induced myocardial hypertrophy, probably as a result of the activation of both the sympathetic nervous system and the renin-angiotensin system. Phenotypic changes are likely. These cardiac muscle cells exhibit contractile dysfunction, which cannot be compensated for by more rapid relaxation and contraction speeds, whereas in the Akt expressing cardiac muscle cells functional compensation had taken place. Thirdly, cardiomyocytes from transgenic mice overexpressing TGF-beta were isolated. These mice have a moderate myocardial hypertrophy, but no raised pressure load. TGF-beta is assumed to be the end point of an angiotensin II effect, which then contributes to myocardial hypertrophy as a result of altered beta;-adrenergic receptor coupling. In the model used here, it is possible to decouple this part of the postulated chain of cause and effect (angiotensin II – TGF-beta – beta-adrenergic receptors) from the individual effect of angiotensin II. This was further demonstrated by a series of experiments which interfered at each of the three stages of the chain of cause and effect in the transgenic animals. The TGF-beta overexpressing mice were treated with Telmisartan (an ATI receptor antagonist), TGF-beta antibodies and beta-blockers and subsequently examined for their beta-adrenergic responsiveness. These experiments demonstrate the significance of TGF-beta in reduced responsiveness of beta-adrenergic receptors in pressure induced myocardial hypertrophy. They show, for the first time, that the administration of beta-blockers can correct TGF-beta induced functional changes at the cellular level

Early assessment of high-intensity focused ultrasound treatment of benign thyroid nodules by scintigraphic means

Background: High-intensity focused ultrasound (HIFU) allows to inflict intracorporal thermal lesions without penetrating the skin or damaging the surrounding tissue. This analysis intends to assess the magnitude of HIFU-induced ablations within benign thyroid nodules using scintigraphic imaging with 99mTc. Methods: Ten cold, hot, or indifferent nodules were treated using multiple pulses of HIFU to induce temperatures of around 85°C within the ablation zone. Pre- and posttreatment, uptake values of 99mTc pertechnetate or 99mTc-MIBI were recorded. The pre-post reduction of nodular uptake was evaluated to assess ablation magnitude. Results: Relative nodular uptake in relation to total thyroidal uptake decreased after one session of HIFU in all cases. Median 99mTc-MIBI uptake reduction was 35.5% (ranging from 11% to 57%; p < 0.1), while 99mTc-pertechnetate scintigraphy showed a median uptake reduction of 27% (range 10% to 44%; p < 0.1). No major complications were observed. Conclusions: HIFU appears to be safe and is an easy to perform means of thermal ablation. This study shows that HIFU treatment in thyroidal nodules can be evaluated by scintigraphic means shortly after the intervention. Due to small sample size, the exact magnitude of HIFU ablation efficiency in thyroidal nodules remains a value to beassessed in a larger study

Reduced efficacy of the plk1 inhibitor bi 2536 on the progression of hepatocellular carcinoma due to low intratumoral drug levels

Highly promising preclinical data obtained in cultured cells and in nude mice bearing xenografts contrast with the rather modest clinical efficacy of Polo-like kinase 1 (Plk1) inhibitors. In the present study, we investigated if Plk1 might be a suitable target in hepatocellular carcinoma (HCC) and if a genetically engineered mouse tumor model that well reflects the tumor cell and micro-environmental features of naturally occurring cancers might be suitable to study anti-Plk1 therapy. Analysis of Plk1 expression in human HCC samples confirmed that HCC express much higher Plk1 levels than the adjacent normal liver tissue. Inhibition of Plk1 by an adenovirus encoding for a short hairpin RNA against Plk1 or by the small-molecule inhibitor BI 2536 reduced the viability of HCC cell lines and inhibited HCC xenograft progression in nude mice. Treatment of transforming growth factor (TGF) α/c-myc bitransgenic mice with BI 2536 during hepatocarcinogenesis reduced the number of dysplastic foci and of Ki-67-positive cells within the foci, indicating diminished tumorigenesis. In contrast, BI 2536 had no significant effect on HCC progression in the transgenic mouse HCC model as revealed by magnetic resonance imaging. Measurement of BI 2536 by mass spectrometry revealed considerably lower BI 2536 levels in HCC compared with the adjacent normal liver tissue. In conclusion, low intratumoral levels are a novel mechanism of resistance to the Plk1 inhibitor BI 2536. Plk1 inhibitors achieving sufficient intratumoral levels are highly promising in HCC treatment