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The optical imaging plays an increasing role in preclinical studies, particularly in cancer biology. The combined ultrasound and optical imaging, named photoacoustic imaging (PAI), is an emerging hybrid technique for real-time molecular imaging in preclinical research and recently expanding into clinical setting. PAI can be performed using endogenous contrast, particularly from oxygenated and deoxygenated hemoglobin and melanin, or exogenous contrast agents, sometimes targeted for specific biomarkers, providing comprehensive morphofunctional and molecular information on tumor microenvironment. Overall, PAI has revealed notable opportunities to improve knowledge on tumor pathophysiology and on the biological mechanisms underlying therapy. The aim of this review is to introduce the principles of PAI and to provide a brief overview of current PAI applications in preclinical research, highlighting also on recent advances in clinical translation for cancer diagnosis, staging, and therapy.

Organic acids are known phytotoxins in some anaerobic waterlogged soils. In this work, we tested the influence of hexanoic acid (HxA) on pea roots. HxA had an inhibitory effect on root tissue O2 consumption but only at pH 5.0, not at pH 6.0. Intact root tissues exposed to 20 mM HxA for 1 h showed solute leakage. Mitochondria were isolated from pea roots after 1 h incubation in the presence or absence of HxA, at either pH 5.0 or 6.0. The maximum effect of HxA on  was detected at pH 5.0 in the presence of succinic acid and without BSA in the assay buffer. A smaller effect was induced by HxA at pH 5.0 with malic plus glutamic acids, while at pH pH 6.0 no effect was observed. Mitochondrial respiration was affected by HxA after root incubation at pH 5.0. The decrease of respiration was induced by HxA with both succinic acid and malic plus glutamic acids but BSA in the assay buffer enhanced the effect of HxA, at least with succinic acid. These findings strongly suggest that the toxicity of HxA is caused by its un-dissociated form, affecting plasmalemma permeability and mitochondrial activities.

Dexmedetomidine (DEX) and Acepromazine (ACP) are powerful sedatives with remarkable hemodynamic effects, Several phenothiazines and α2-agonist molecules have long been used in translational research separately or in combination. However, their individual or synergistic peripheral hemodynamic effects have not been fully described in different laboratory animal species in spite of their relevance in cardiovascular studies. Some authors reported an attenuation of α2-adrenergic agonist pressor response with the acepromazine-xylazine combination in dogs. Laser Doppler Imaging (LDPI) and High frequency ultrasound echocardiography (HRE) provides best noninvasive measurement of cardiovascular function in mice. The aim of the study was to investigate noninvasively the cardiovascular effects of DEX, of ACP and of their combination in isoflurane (ISO) anesthetized mice. Methods Forty five age-matched and sex-paired CD1 mice, 8 to 10 weeks old, were randomly assigned to one of three experimental groups and underwent 1.5% ISO anesthesia, followed by intraperitoneal injection of either 5 mg/kg ACP, or 1 mg/kg DEX, or by their combination. Body temperature was adjusted to 36 °C. Heart (HR) and respiratory (RR) rates were monitored. Hind paws blood flow (Perfusion Units, volt) was recorded by LDPI apparatus (Periscan®, laser beam power = 1 mV; wavelength = 670 nm; pixel size = 0.25x0.25 mm2; scanner head distance =15 cm; scanning area = 3x2 cm2; scanning time = 2 minutes). An high-resolution ultrasound imaging system (Vevo 770, VisualSonics, transducer frequency 40 MHz, focal length 6 mm, frame rate 30 Hz, spatial resolution 30 μm) synchronized to the electrocardiographic signal was used. A two-dimensional (B-mode) and motion-mode (M-mode) imaging of the left ventricular short axis was taken just at the level of the papillary muscles. Peripheral perfusion (PP), ejection fraction (EF) and fractional shortening (FS) were recorded 10’ and 20’ before and at different intervals after treatments. A further measurement was recorded after reversing dexmedetomidine by the α2-antagonist atipamezole (ATP). We compared PP, HR, RR, EF and FS values at different times by paired non parametric Wilcoxon test and inter-groups differences by One Way Friedman ANOVA. When appropriate, a post hoc analysis by Mann Whitney test was made. Significance was set at P 0.05). ACP increased PP (P=0.005) and reduced FS (P0.05); ATP brought back values such close to baseline (P>0.05). Conclusions In mice ACP+DEX produced more temperate hemodynamic values compared to those following single agents, lessening DEX biphasic hemodynamic pattern. Traslational imaging allows noninvasive and accurate measurements of hemodynamic effects of different sedatives in mice models.

We performed a multistage genome-wide association study of melanoma. In a discovery cohort of 1804 melanoma cases and 1026 controls, we identified loci at chromosomes 15q13.1 (HERC2/OCA2 region) and 16q24.3 (MC1R) regions that reached genome-wide significance within this study and also found strong evidence for genetic effects on susceptibility to melanoma from markers on chromosome 9p21.3 in the p16/ARF region and on chromosome 1q21.3 (ARNT/LASS2/ANXA9 region). The most significant single-nucleotide polymorphisms (SNPs) in the 15q13.1 locus (rs1129038 and rs12913832) lie within a genomic region that has profound effects on eye and skin color; notably, 50% of variability in eye color is associated with variation in the SNP rs12913832. Because eye and skin colors vary across European populations, we further evaluated the associations of the significant SNPs after carefully adjusting for European substructure. We also evaluated the top 10 most significant SNPs by using data from three other genome-wide scans. Additional in silico data provided replication of the findings from the most significant region on chromosome 1q21.3 rs7412746 (P = 6 × 10(-10)). Together, these data identified several candidate genes for additional studies to identify causal variants predisposing to increased risk for developing melanoma.

Different animal models have been used to reproduce coronary heart disease but in the last years mice became the animals of choice, because of their short life cycle and the possibility of genetic manipulation. Various techniques are currently used for cardiovascular imaging in mice, including high-resolution ultrasound, X-ray computed tomography (CT), magnetic resonance imaging and nuclear medicine procedures. In particular, molecular imaging with cardiac positron emission tomography (PET) allows to evaluate noninvasively changes in myocardial perfusion, metabolism, apoptosis, inflammation, and gene expression or to measure changes in left ventricular functional parameters. With technological advancements, dedicated small laboratory PET/CT imaging has emerged in cardiovascular research, providing in vivo a non-invasive, serial and quantitative assessment of left ventricular function, myocardial perfusion and metabolism at a molecular level. This non-invasive methodology might be useful in longitudinal studies monitoring cardiac biochemical parameters and might facilitate studies to assess the effect of different interventions after acute myocardial ischemia.

Anesthetics can alter microvascular perfusion, affecting tissue oxygenation and delivery of vital substrates. The ??2???adrenergic agonist dexmedetomidine and the ?????blocker acepromazine are powerful sedatives with remarkable hemodynamic effects. Some authors reported an attenuation of the ??2???adrenergic agonist pressor response by an acepromazine???xylazine combination in dogs. We investigated non???invasively the microcirculatory effects of dexmedetomidine, of acepromazine and of their combination in isoflurane anesthetized mice by Laser Doppler Perfusion Imaging (LDPI). Thirty???two age???matched and sex???paired CD1 mice underwent 1.5% isoflurane anesthesia, followed by intraperitoneal injection of either 5 mg/kg acepromazine, or 1 mg/kg dexmedetomidine, or by their combination. Body temperature was adjusted to 36 °C. Heart (HR) and breath (BR) rate were recorded. Hind paws blood flow (Perfusion Units, volt) was recorded by LDPI 10 and 20 minutes after isoflurane induction, at different intervals after treatments, and after reversing dexmedetomidine by the ??2??? antagonist atipamezole. BR decreased in all groups without significant differences to baseline (P>0.05). Dexmedetomidine sharply reduced over time HR (P0.05). Acepromazine+dexmedetomidine decreased HR (P0.05); atipamezole gradually raised HR close to baseline (P>0.05). Peripheral perfusion under isoflurane anesthesia showed an increasing trend after 10 and 20 minutes, without differences among groups (P=0.1). Acepromazine increased perfusion between 10 and 20 minutes (P=0.005). Dexmedetomidine reduced blood perfusion after 5 minutes (P=0.0001), followed by an increase after 15 minutes (P=0.008). No significant changes were seen 5 minutes after atipamezole (P=0.9). Acepromazine+dexmedetomidine resulted in steady perfusion values over time (P=0.44), which after atipamezole increased very close to baseline (P=0.237). Acepromazine+dexmedetomidine in mice produced more temperate, steady peripheral perfusion values compared to those following single agent, reducing the entity of the ??2 ???agonist biphasic hemodynamic pattern. Our translational approach by LDPI in a mouse model allows an easy, accurate and non invasive measurement of the effects of anesthetics on peripheral microcirculation. 1. Alvaides RK, Neto FJ, Aguiar AJ, et al. Sedative and cardiorespiratory effects of acepromazine or atropine given before Dexmedetomidine in dogs. Vet Rec 2008; 26: 852???856. 2. B. J. A. Janssen, T. De Celle, J. J. M. Debets, A. E. et al, ???Effects of anesthetics on systemic hemodynamics in mice,??? Am J Physiol Heart Circ Physiol, vol. 4, no. 287, pp. 1618???1624, 2004. 3. Adelaide Greco, Monica Ragucci, Raffaele Liuzzi, Sara Gargiulo, Matteo Gramanzini, e al. Reproducibility and Standardization of Laser doppler Imaging technique for the evaluation of normal mice hindlimbs

Background: Anesthetic agents alter microcirculation, influencing tissue oxygenation and delivery of vital substrates. Laser Doppler perfusion imaging is a widespread technique in the field of microvascular research that can evaluate noninvasively and in real time the effects of environmental conditions, physical manipulations, diseases and treatments on peripheral perfusion. This study aims to evaluate laser Doppler perfusion imaging as a means to detect changes in skin microcirculation induced by some popular anesthetic agents in a murine model. Twenty-four age- and gender-matched healthy CD1 mice were examined by laser Doppler perfusion imaging. The skin microcirculatory response was measured at the level of plantar surfaces during isoflurane anesthesia with or without subsequent dexmedetomidine or acepromazine. At the end of the procedure, dexmedetomidine was reversed by atipamezole administration. Results: In all mice, skin blood flow under isoflurane anesthesia did not show significant differences over time (P = 0.1). The serial perfusion pattern and values following acepromazine or dexmedetomidine administration differed significantly (P < 0.05). Conclusions: We standardized a reliable laser Doppler perfusion imaging protocol to non-invasively assess changes in skin microcirculation induced by anesthesia in mice, considering the advantages and drawbacks of this technique and its translational value. Keywords: Microvascular perfusion, Anesthesia, Murine model, Laser Doppler perfusion imaging

Human glioblastoma is the most frequent and aggressive form of brain tumour in the adult population. Proteolytic turnover of tumour suppressors by the ubiquitin-proteasome system is a mechanism that tumour cells can adopt to sustain their growth and invasiveness. However, the identity of ubiquitin-proteasome targets and regulators in glioblastoma are still unknown. Here we report that the RING ligase praja2 ubiquitylates and degrades Mob, a core component of NDR/LATS kinase and a positive regulator of the tumour-suppressor Hippo cascade. Degradation of Mob through the ubiquitin-proteasome system attenuates the Hippo cascade and sustains glioblastoma growth in vivo. Accordingly, accumulation of praja2 during the transition from low- to high-grade glioma is associated with significant downregulation of the Hippo pathway. These findings identify praja2 as a novel upstream regulator of the Hippo cascade, linking the ubiquitin proteasome system to deregulated glioblastoma growth.

Introduction: The translocator protein 18 kDa (TSPO), a biochemical marker of neuroinflammation, is highly expressed in the brain activated microglia and it is also expressed by peripheral inflammatory cells and normal peripheral tissues. Thus, development of radioligands for the TSPO may contribute to further understanding the in vivo TSPO function in central and peripheral inflammatory processes and other pathologies. Here, we report the biodistribution, the specific binding and the radiometabolites of [(18)F]DPA-714, a promising fluorinated PET radiotracer, in normal mice using a microPET/CT scanner. Methods: The in vivo biodistribution and kinetics of [(18)F]DPA-714 were measured in mice brain and peripheral tissues. Specific binding to TSPO sites was assessed using pharmacological competitive studies by means of saturation experiments performed by i.v. injection of 1mg/kg of unlabeled DPA-714 or 3mg/kg of unlabeled PK11195. A region of interest analysis was performed to generate time-activity curves in the brain, heart, lung, kidney, spleen and liver. Metabolites assay was performed in the plasma and peripheral organs by radio-HPLC. Results: [(18)F]DPA-714 reached high concentration in lung, heart, kidney and spleen, tissues well known to be rich in TSPO sites. [(18)F]DPA-714 kinetics were faster in the lung and slower in the kidney. Pre-injection of unlabeled DPA-714 or PK11195 inhibited about 80% of [(18)F]DPA-714 uptake in the lung and heart (p

Animal experiments are necessary for a better understanding of diseases and for developing new therapeutic strategies. The mouse (Mus musculus) is currently the most popular laboratory animal in biomedical research. Mice imaging procedures are increasingly used in preclinical research because they allow in vivo monitoring and they are readily available for longitudinal and noninvasive studies as well as investigations into the evolution of diseases and the effects of new therapies. New imaging techniques and sophisticated laboratory animal imaging tools are currently producing a large body of evidence about the possible interference of anesthesia with different imaging methods that have the potential to compromise the results of in vivo studies. The purpose of this article is to review the existing literature on molecular imaging studies in mice, to describe the effects of different anesthetic protocols on their outcome, and to report our own experience with such studies.

La Biomicroscopia ad Ultrasuoni (UBM) rappresenta una metodica di Diagnostica per immagini sperimentale che permette di eseguire studi morfologici e ultrastrutturali in maniera non invasiva e ripetibili nel tempo in modelli animali di malattia umana. Uno dei campi in crescente espansione è quello legato all’ecografia applicata alla diagnosi precoce, alla stadiazione della gravidanza e alla microiniezione in utero. La microiniezione ecoguidata in utero nel periodo embrionale sta diventando unametodologia molto attrattiva per la prevenzione e la cura di malattie genetiche. Inoltre la terapia genica in età fetale offre altri vantaggi, in quanto può indurre nel feto una tolleranza immunitaria contro i transgeni introdottigrazie al fatto che il sistema immunitario del feto non è maturo. Inoltre in età fetale è possible bypassare la barriera emato-encefalica per influenzare la differenziazione delle cellule neuronali e delle staminali. La microiniezione in utero ecoguidata nel topo di laboratorio trova diverseapplicazioni: veicolare a scopo terapeutico marcatori fluorescenti, materiale genico,cellule, farmaci, vettori virali; studi di embriogenesi e/o di lineage cellulare nel l’encefalo, nel cuore, nell’occhio del feto, nella cavità amniotica e nei vasi del cordone ombelicale. Esistono ad oggi, due metodiche descritte in letteratura per l’ersecuzione della microiniezione in utero nel topo di laboratorio.

L???Ecografia ad altissima risoluzione, detta anche Biomicroscopia ad Ultrasuoni (UBM) è una moderna tecnica che utilizza gli ultrasuoni per fornire immagini dei tessuti in tempo reale e ad altissima risoluzione. Questa tecnica ha il vantaggio di essere non invasiva, di avere costi contenuti rispetto alle altre metodiche avanzate di Imaging e di essere ripetibile, consentendo un monitoraggio nel tempo delle strutture corporee da esaminare. Recentemente, questa metodica sta avendo un crescente utilizzo nel topo di laboratorio perché può consentire di individuare biomarkers tumorali molecolari, di studiare la differenziazione neoplastica e di valutare l???effetto delle terapie sulla crescita e sulla regressione dei tumori. Inoltre, l???utilizzo del Doppler consente di quantificare la neoangiogenesi tumorale . Altre applicazioni dell???UBM sono la diagnosi precoce di gravidanza nel topo, il monitoraggio dello sviluppo dell???apparato cardiovascolare dell???embrione e l???esecuzione di microiniezioniecoguidate di farmaci, geni e altre molecole. Recentemente, diversi gruppi di ricerca stanno utilizzando le microbolle dei mezzi di contrasto ecografici come supporto per il rilascio di geni e farmaci direttamente nel tessuto d???interesse, in sostituzione di metodiche meno efficienti. Alla luce dei progressi fatti nella Diagnostica per Immagini sperimentale, risulta interessante pensare a possibili applicazioni future dell???UBM nella Clinica Chirurgica Veterinaria, in particolare nellostudio di strutture superficiali come l???occhio, in quanto gli elementi costituitivi del segmento anteriore dell???occhio e le porzioni periferiche della retina sono localizzate entro la profonditàconsentita dal potere di penetrazione tessutale (5-10 mm) che caratterizza questa metodica. Pochi sono, infatti, i dati presenti in letteratura riguardanti l???uso dell???UBM nel cane e nel gatto.Scopo del presente lavoro è di descrivere i principi base dell???ecografia ad alta risoluzione nellasperimentazione animale, alcune applicazioni sperimentali maturate nella nostra esperienza, ed un excursus delle possibili applicazioni dell???UBM nella clinica del cane e del gatto.

Animal experiments are necessary for a better understanding of diseases and for developing new therapeutic strategies. The mouse (Mus musculus) is currently the most popular laboratory animal in biomedical research. Mice imaging procedures are increasingly used in preclinical research because they allow in vivo monitoring and they are readily available for longitudinal and noninvasive studies as well as investigations into the evolution of diseases and the effects of new therapies. New imaging techniques and sophisticated laboratory animal imaging tools are currently producing a large body of evidence about the possible interference of anesthesia with different imaging methods that have the potential to compromise the results of in vivo studies. The purpose of this article is to review the existing literature on molecular imaging studies in mice, to describe the effects of different anesthetic protocols on their outcome, and to report our own experience with such studies.

Different species have been used to reproduce myocardial infarction models but in the last years mice became the animals of choice for the analysis of several diseases, due to their short life cycle and the possibility of genetic manipulation. Many techniques are currently used for cardiovascular imaging in mice, including X-ray computed tomography (CT), high-resolution ultrasound, magnetic resonance imaging, and nuclear medicine procedures. Cardiac positron emission tomography (PET) allows to examine noninvasively, on a molecular level and with high sensitivity, regional changes in myocardial perfusion, metabolism, apoptosis, inflammation, and gene expression or to measure changes in anatomical and functional parameters in heart diseases. Currently hybrid PET/CT scanners for small laboratory animals are available, where CT adds high-resolution anatomical information. This paper reviews mouse models of myocardial infarction and discusses the applications of dedicated PET/CT systems technology, including animal preparation, anesthesia, radiotracers, and images postprocessing.

L’innovativo approccio molecolare ha contribuito significativamente a migliorare la tempestività delle diagnosi, il monitoraggio dell’evoluzione delle malattie e lo sviluppo di nuove terapie. Le patologie cardiovascolari, ed in particolare la cardiomiopatia ischemica, sono attualmente annoverate tra le principali cause di morbilità e mortalità nel mondo occidentale. Pertanto, la comunità scientifica nutre un forte interesse nel perfezionare l’iter diagnostico e sviluppare programmi preventivi e terapeutici innovativi per le cardiopatie. La comprensione della patogenesi e dell’evoluzione dell’ischemia del miocardio e lo sviluppo di nuove terapie si sono evolute di pari passo con la sperimentazione biomedica sui modelli animali. Le potenzialità di manipolazione genetica nei roditori da laboratorio hanno consentito di migliorare la comprensione di fondamentali aspetti genetici e fisiopatologici delle malattie cardiovascolari umane. In particolare, la crescente disponibilità di numerosi ceppi di topo (Mus Musculus) geneticamente modificati, ha condotto alla loro progressiva affermazione rispetto ad altri modelli animali. L’imaging preclinico nei topi da laboratorio ha apportato un contributo rilevante alla ricerca cardiovascolare. La traslazione delle metodiche di imaging dal modello animale all’uomo non è semplice ed in particolare, il loro impiego nel topo per lo studio del cuore è complicata da una serie di fattori, come le ridotte dimensioni e l’elevata frequenza cardiaca. Pertanto, gli studi di diagnostica per immagini sul cuore del topo richiedono alta risoluzione spaziale e temporale. Diverse metodiche di imaging, come l’Ecocardiografia, la Tomografia Computerizzata a raggi X (TC), la Risonanza Magnetica (RM), la Tomografia ad Emissione di Positroni (PET) e la Tomografia ad Emissione di Singolo Fotone (SPECT) consentono di studiare in vivo, in modo non invasivo e longitudinale, la morfologia, il metabolismo e la funzionalità cardiaca. In particolare, l’impiego di tecniche di Imaging morfo-funzionale integrato, come PET/TC o PET/RM, permettono di valutare in vivo processi molecolari che avvengono a livello cellulare e contemporaneamente ottenere informazioni anatomiche ad alta risoluzione.

Amelanotic melanoma (AM) is a rare subtype of melanoma with little or no clinically visible pigment; it is more difficult to diagnose than pigmented melanoma (PM), and has a worse prognosis. In the attempt to find a genetic explanation for the distinction between AM and PM, we conducted a case-case study, matching AM and PM patients, and testing them for germline mutations in high(p16INK4A, p14ARF, CDK4) and low-penetrance (MC1R) melanoma susceptibility genes. Similar CDKN2A mutations were found in both sets of melanomas. A p14ARF splice germline mutation was detected for the first time in an Italian family with AM. This rare mutation, which has been described only once previously, may be involved in predisposition to the amelanotic phenotype in combination with germline MC1R variants and coordinate somatic expression of pigmentation genes and their regulators. Melanoma Res 19:142-145 (C) 2009 Wolters Kluwer Health vertical bar Lippincott Williams & Wilkins.