Characterization of microcirculation in multiple sclerosis lesions by dynamic texture parameter analysis (DTPA)

OBJECTIVE Texture analysis is an alternative method to quantitatively assess MR-images. In this study, we introduce dynamic texture parameter analysis (DTPA), a novel technique to investigate the temporal evolution of texture parameters using dynamic susceptibility contrast enhanced (DSCE) imaging. Here, we aim to introduce the method and its application on enhancing lesions (EL), non-enhancing lesions (NEL) and normal appearing white matter (NAWM) in multiple sclerosis (MS). METHODS We investigated 18 patients with MS and clinical isolated syndrome (CIS), according to the 2010 McDonald's criteria using DSCE imaging at different field strengths (1.5 and 3 Tesla). Tissues of interest (TOIs) were defined within 27 EL, 29 NEL and 37 NAWM areas after normalization and eight histogram-based texture parameter maps (TPMs) were computed. TPMs quantify the heterogeneity of the TOI. For every TOI, the average, variance, skewness, kurtosis and variance-of-the-variance statistical parameters were calculated. These TOI parameters were further analyzed using one-way ANOVA followed by multiple Wilcoxon sum rank testing corrected for multiple comparisons. RESULTS Tissue- and time-dependent differences were observed in the dynamics of computed texture parameters. Sixteen parameters discriminated between EL, NEL and NAWM (pAVG = 0.0005). Significant differences in the DTPA texture maps were found during inflow (52 parameters), outflow (40 parameters) and reperfusion (62 parameters). The strongest discriminators among the TPMs were observed in the variance-related parameters, while skewness and kurtosis TPMs were in general less sensitive to detect differences between the tissues. CONCLUSION DTPA of DSCE image time series revealed characteristic time responses for ELs, NELs and NAWM. This may be further used for a refined quantitative grading of MS lesions during their evolution from acute to chronic state. DTPA discriminates lesions beyond features of enhancement or T2-hypersignal, on a numeric scale allowing for a more subtle grading of MS-lesions.

Characterization of a novel CYP19A1 (aromatase) R192H mutation causing virilization of a 46,XX newborn, undervirilization of the 46,XY brother, but no virilization of the mother during pregnancies.

BACKGROUND P450 aromatase (CYP19A1) is essential for the biosynthesis of estrogens from androgen precursors. Mutations in the coding region of CYP19A1 lead to autosomal recessive aromatase deficiency. To date over 20 subjects have been reported with aromatase deficiency which may manifest during fetal life with maternal virilization and virilization of the external genitalia of a female fetus due to low aromatase activity in the steroid metabolizing fetal-placental unit and thus high androgen levels. During infancy, girls often have ovarian cysts and thereafter fail to enter puberty showing signs of variable degree of androgen excess. Moreover, impact on growth, skeletal maturation and other metabolic parameters is seen in both sexes. OBJECTIVE AND HYPOTHESIS We found a novel homozygous CYP19A1 mutation in a 46,XX girl who was born at term to consanguineous parents. Although the mother did not virilize during pregnancy, the baby was found to have a complex genital anomaly at birth (enlarged genital tubercle, fusion of labioscrotal folds) with elevated androgens at birth, normalizing thereafter. Presence of 46,XX karyotype and female internal genital organs (uterus, vagina) together with biochemical findings and follow-up showing regression of clitoral hypertrophy, as well as elevated FSH suggested aromatase deficiency. Interestingly, her older brother presented with mild hypospadias and bilateral cryptorchidism and was found to carry the same homozygous CYP19A1 mutation. To confirm the clinical diagnosis, genetic, functional and computational studies were performed. METHODS AND RESULTS Genetic analysis revealed a homozygous R192H mutation in the CYP19A1 gene. This novel mutation was characterized for its enzymatic activity (Km, Vmax) in a cell model and found to have markedly reduced catalytic activity when compared to wild-type aromatase; thus explaining the phenotype. Computational studies suggest that R192H disrupts the substrate access channel in CYP19A1 that may affect binding of substrates and exit of catalytic products. CONCLUSION R192H is a novel CYP19A1 mutation which causes a severe phenotype of aromatase deficiency in a 46,XX newborn and maybe hypospadias and cryptorchidism in a 46,XY, but no maternal androgen excess during pregnancy.

Deoxynucleoside triphosphates bearing histamine, carboxylic acid, and hydroxyl residues--synthesis and biochemical characterization.

Modified nucleoside triphosphates (dA(Hs)TP, dU(POH)TP, and dC(Val)TP) bearing imidazole, hydroxyl, and carboxylic acid residues connected to the purine and pyrimidine bases through alkyne linkers were prepared. These modified dN*TPs were excellent substrates for various DNA polymerases in primer extension reactions. Moreover, the combined use of terminal deoxynucleotidyl transferase (TdT) and the modified dNTPs led to efficient tailing reactions that rival those of natural counterparts. Finally, the triphosphates were tolerated by polymerases under PCR conditions, and the ensuing modified oligonucleotides served as templates for the regeneration of unmodified DNA. Thus, these modified dN*TPs are fully compatible with in vitro selection methods and can be used to develop artificial peptidases based on DNA.

Characterization of the inflammatory infiltrate and cytokine expression in the skin of horses with recurrent urticaria.

BACKGROUND Recurrent urticaria (RU) is a common skin disease of horses, but little is known about its pathogenesis. HYPOTHESIS/OBJECTIVE The aim of this study was to characterize the inflammatory cell infiltrate and cytokine expression pattern in the skin of horses with RU. ANIMALS   Biopsies of lesional and nonlesional skin of horses with RU (n = 8) and of skin from healthy control horses (n = 8) were evaluated. METHODS The inflammatory cell infiltrate was analysed by routine histology. Immunohistochemistry was used to identify T cells (CD3), B  ells (CD79), macrophages (MAC387) and mast cells (tryptase). Expression of T-helper 2 cytokines (interleukins IL-4, IL-5 and IL-13), a T-helper 1 cytokine (interferon-γ), IL-4 receptor α and thymic stromal lymphopoietin was assessed by quantitative RT-PCR. Results -  In subepidermal lesional skin of RU-affected horses, increased numbers of eosinophils (P ≤ 0.01), CD79-positive (P ≤ 0.01), MAC387-positive (P ≤ 0.01) and tryptase-positive cells (P ≤ 0.05) were found compared with healthy horses. Subepidermal lesional skin of RU-affected horses contained more eosinophils (P ≤ 0.05) and tryptase-positive cells (P ≤ 0.05) compared with nonlesional skin. There was no significant difference in infiltrating cells between nonlesional skin and skin of healthy horses. Expression of IL-4 (P ≤ 0.01), IL-13 (P ≤ 0.05), thymic stromal lymphopoietin (P ≤ 0.05) and IL-4 receptor α (P ≤ 0.05) was increased in lesional skin of RU-affected horses compared with control horses. Expression of IL-4 was higher (P ≤ 0.05) in lesional compared with nonlesional RU skin. CONCLUSIONS AND CLINICAL IMPORTANCE Analysis of cytokine expression and inflammatory infiltrate suggests that T-helper 2 cytokines, eosinophils, mast cells and presumptive macrophages play a role in the pathogenesis of equine RU.

Microstructural and mechanical characterization of contemporary lingual orthodontic brackets.

OBJECTIVES To investigate the composition and the microstructural and mechanical characterization of three different types of lingual brackets. MATERIALS AND METHODS Incognito™ (3M Unitek), In-Ovation L (DENTSPLY GAC) and STb™ (Light Lingual System, ORMCO) lingual brackets were studied under the scanning electron microscope employing backscattered electron imaging and their elemental composition was analysed by energy-dispersive X-ray microanalysis. Additionally, Vickers hardness was assessed using a universal hardness-testing machine, and the indentation modulus was measured according to instrumented indentation test. Two-way analysis of variance was conducted employing bracket type and location (base and wing) as discriminating variable. Significant differences among groups were allocated by post hoc Student-Newman-Keuls multiple comparison analysis at 95% level of significance. RESULTS Three different phases were identified for Incognito and In-Ovation L bracket based on mean atomic number contrast. On the contrary, STb did not show mean atomic contrast areas and thus it is recognized as a single phase. Incognito is a one-piece bracket with the same structure in wing and base regions. Incognito consists mainly of noble metals while In-Ovation L and STb show similar formulations of ferrous alloys in wing and base regions. No significant differences were found between ferrous brackets in hardness and modulus values, but there were significant differences between wing and base regions. Incognito illustrated intermediate values with significant differences from base and wing values of ferrous brackets. CONCLUSIONS/IMPLICATIONS Significant differences exist in microstructure, elemental composition, and mechanical properties among the brackets tested; these might have a series of clinical implications during mechanotherapy.

In vitro characterization of a synthetic calcium phosphate bone graft on periodontal ligament cell and osteoblast behavior and its combination with an enamel matrix derivative.

OBJECTIVES Recent studies suggest that a combination of enamel matrix derivative (EMD) with grafting material may improve periodontal wound healing/regeneration. Newly developed calcium phosphate (CaP) ceramics have been demonstrated a viable synthetic replacement option for bone grafting filler materials. AIMS This study aims to test the ability for EMD to adsorb to the surface of CaP particles and to determine the effect of EMD on downstream cellular pathways such as adhesion, proliferation, and differentiation of primary human osteoblasts and periodontal ligament (PDL) cells. MATERIALS AND METHODS EMD was adsorbed onto CaP particles and analyzed for protein adsorption patterns via scanning electron microscopy and high-resolution immunocytochemistry with an anti-EMD antibody. Cell attachment and cell proliferation were quantified using CellTiter 96 One Solution Cell Assay (MTS). Cell differentiation was analyzed using real-time PCR for genes encoding Runx2, alkaline phosphatase, osteocalcin, and collagen1α1, and mineralization was assessed using alizarin red staining. RESULTS Analysis of cell attachment revealed significantly higher number of cells attached to EMD-adsorbed CaP particles when compared to control and blood-adsorbed samples. EMD also significantly increased cell proliferation at 3 and 5 days post-seeding. Moreover, there were significantly higher mRNA levels of osteoblast differentiation markers including collagen1α1, alkaline phosphatase, and osteocalcin in osteoblasts and PDL cells cultured on EMD-adsorbed CaP particles at various time points. CONCLUSION The present study suggests that the addition of EMD to CaP grafting particles may influence periodontal regeneration by stimulating PDL cell and osteoblast attachment, proliferation, and differentiation. Future in vivo and clinical studies are required to confirm these findings. CLINICAL RELEVANCE The combination of EMD and CaP may represent an option for regenerative periodontal therapy in advanced intrabony defects.

Postmortem MR quantification of the heart for characterization and differentiation of ischaemic myocardial lesions.

OBJECTIVES Recently, an MRI quantification sequence has been developed which can be used to acquire T1- and T2-relaxation times as well as proton density (PD) values. Those three quantitative values can be used to describe soft tissue in an objective manner. The purpose of this study was to investigate the applicability of quantitative cardiac MRI for characterization and differentiation of ischaemic myocardial lesions of different age. MATERIALS AND METHODS Fifty post-mortem short axis cardiac 3 T MR examinations have been quantified using a quantification sequence. Myocardial lesions were identified according to histology and appearance in MRI images. Ischaemic lesions were assessed for mean T1-, T2- and proton density values. Quantitative values were plotted in a 3D-coordinate system to investigate the clustering of ischaemic myocardial lesions. RESULTS A total of 16 myocardial lesions detected in MRI images were histologically characterized as acute lesions (n = 8) with perifocal oedema (n = 8), subacute lesions (n = 6) and chronic lesions (n = 2). In a 3D plot comprising the combined quantitative values of T1, T2 and PD, the clusters of all investigated lesions could be well differentiated from each other. CONCLUSION Post-mortem quantitative cardiac MRI is feasible for characterization and discrimination of different age stages of myocardial infarction. KEY POINTS • MR quantification is feasible for characterization of different stages of myocardial infarction. • The results provide the base for computer-aided MRI cardiac infarction diagnosis. • Diagnostic criteria may also be applied for living patients.

Comparison of gadoteric acid and gadobutrol for detection as well as morphologic and dynamic characterization of lesions on breast dynamic contrast-enhanced magnetic resonance imaging

OBJECTIVE In contrast to conventional breast imaging techniques, one major diagnostic benefit of breast magnetic resonance imaging (MRI) is the simultaneous acquisition of morphologic and dynamic enhancement characteristics, which are based on angiogenesis and therefore provide insights into tumor pathophysiology. The aim of this investigation was to intraindividually compare 2 macrocyclic MRI contrast agents, with low risk for nephrogenic systemic fibrosis, in the morphologic and dynamic characterization of histologically verified mass breast lesions, analyzed by blinded human evaluation and a fully automatic computer-assisted diagnosis (CAD) technique. MATERIALS AND METHODS Institutional review board approval and patient informed consent were obtained. In this prospective, single-center study, 45 women with 51 histopathologically verified (41 malignant, 10 benign) mass lesions underwent 2 identical examinations at 1.5 T (mean time interval, 2.1 days) with 0.1-mmol kg doses of gadoteric acid and gadobutrol. All magnetic resonance images were visually evaluated by 2 experienced, blinded breast radiologists in consensus and by an automatic CAD system, whereas the morphologic and dynamic characterization as well as the final human classification of lesions were performed based on the categories of the Breast imaging reporting and data system MRI atlas. Lesions were also classified by defining their probability of malignancy (morpho-dynamic index; 0%-100%) by the CAD system. Imaging results were correlated with histopathology as gold standard. RESULTS The CAD system coded 49 of 51 lesions with gadoteric acid and gadobutrol (detection rate, 96.1%); initial signal increase was significantly higher for gadobutrol than for gadoteric acid for all and the malignant coded lesions (P < 0.05). Gadoteric acid resulted in more postinitial washout curves and fewer continuous increases of all and the malignant lesions compared with gadobutrol (CAD hot spot regions, P < 0.05). Morphologically, the margins of the malignancies were different between the 2 agents, whereas gadobutrol demonstrated more spiculated and fewer smooth margins (P < 0.05). Lesion classifications by the human observers and by the morpho-dynamic index compared with the histopathologic results did not significantly differ between gadoteric acid and gadobutrol. CONCLUSIONS Macrocyclic contrast media can be reliably used for breast dynamic contrast-enhanced MRI. However, gadoteric acid and gadobutrol differed in some dynamic and morphologic characterization of histologically verified breast lesions in an intraindividual, comparison. Besides the standardization of technical parameters and imaging evaluation of breast MRI, the standardization of the applied contrast medium seems to be important to receive best comparable MRI interpretation.

Characterization of small ruminant lentivirus A4 subtype isolates and assessment of their pathogenic potential in naturally infected goats.

BACKGROUND Small ruminant lentiviruses escaping efficient serological detection are still circulating in Swiss goats in spite of a long eradication campaign that essentially eliminated clinical cases of caprine arthritis encephalitis in the country. This strongly suggests that the circulating viruses are avirulent for goats.To test this hypothesis, we isolated circulating viruses from naturally infected animals and tested the in vitro and in vivo characteristics of these field isolates. METHODS Viruses were isolated from primary macrophage cultures. The presence of lentiviruses in the culture supernatants was monitored by reverse transcriptase assay. Isolates were passaged in different cells and their cytopathogenic effects monitored by microscopy. Proviral load was quantified by real-time PCR using customized primer and probes. Statistical analysis comprised Analysis of Variance and Bonferroni Multiple Comparison Test. RESULTS The isolated viruses belonged to the small ruminant lentiviruses A4 subtype that appears to be prominent in Switzerland. The 4 isolates replicated very efficiently in macrophages, displaying heterogeneous phenotypes, with two isolates showing a pronounced cytopathogenicity for these cells. By contrast, all 4 isolates had a poor replication capacity in goat and sheep fibroblasts. The proviral loads in the peripheral blood and, in particular, in the mammary gland were surprisingly high compared to previous observations. Nevertheless, these viruses appear to be of low virulence for goats except for the mammary gland were histopathological changes were observed. CONCLUSIONS Small ruminant lentiviruses continue to circulate in Switzerland despite a long and expensive caprine arthritis encephalitis virus eradication campaign. We isolated 4 of these lentiviruses and confirmed their phylogenetic association with the prominent A4 subtype. The pathological and histopathological analysis of the infected animals supported the hypothesis that these A4 viruses are of low pathogenicity for goats, with, however, a caveat about the potentially detrimental effects on the mammary gland. Moreover, the high proviral load detected indicates that the immune system of the animals cannot control the infection and this, combined with the phenotypic plasticity observed in vitro, strongly argues in favour of a continuous and precise monitoring of these SRLV to avoid the risk of jeopardizing a long eradication campaign.

Development and characterization of a scaffold-free 3D spheroid model of iPSC-derived human cardiomyocytes

Purpose: Cardiomyocytes are terminally differentiated cells in the adult heart and ischemia and cardiotoxic compounds can lead to cell death and irreversible decline of cardiac function. As testing platforms, isolated organs and primary cells from rodents have been the standard in research and toxicology, but there is a need for better models that more faithfully recapitulate native human biology. Hence, a new in vitro model comprising the advantages of 3D cell culture and the availability of induced pluripotent stem cells (iPSC) from human origin was developed and characterized. Methods: Human cardiomyocytes (CMs) derived from induced pluripotent stem cells (iPSCs) were studied in standard 2D culture and as cardiac microtissues (MTs) formed in hanging drops. 2D cultures were examined using immunofluorescence microscopy and Western blotting while the cardiac MTs were subjected to immunofluorescence, contractility, and pharmacological investigations. Results: iPSC-derived CMs in 2D culture showed well-formed myofibrils, cell-cell contacts positive for connexin-43, and other typical cardiac proteins. The cells reacted to pro-hypertrophic growth factors with a substantial increase in myofibrils and sarcomeric proteins. In hanging drop cultures, iPSC-derived cardiomyocytes formed spheroidal MTs within 4 days showing a homogeneous tissue structure with well-developed myofibrils extending throughout the whole spheroid without a necrotic core. MTs showed spontaneous contractions for more than 4 weeks that were recorded by optical motion tracking, sensitive to temperature, and responsive to electrical pacing. Contractile pharmacology was tested with several agents known to modulate cardiac rate and viability. Calcium-transients underlay the contractile activity and were also responsive to electrical stimulation, caffeine-induced Ca2+-release, extracellular calcium levels. Conclusions: 3D culture using iPSC-derived human cardiomyocytes provides an organoid human-based cellular platform that is free of necrosis and recapitulates vital cardiac functionality, thereby providing new and promising relevant model for the evaluation and development of new therapies and detection of cardiotoxicity.

Real-Time In Vivo Characterization of Primary Liver Tumors With Diffuse Optical Spectroscopy During Percutaneous Needle Interventions: Feasibility Study in Woodchucks

OBJECTIVE This study presents the first in vivo real-time optical tissue characterization during image-guided percutaneous intervention using near-infrared diffuse optical spectroscopy sensing at the tip of a needle. The goal of this study was to indicate transition boundaries from healthy tissue to tumors, namely, hepatic carcinoma, based on the real-time feedback derived from the optical measurements. MATERIALS AND METHODS Five woodchucks with hepatic carcinoma were used for this study. The woodchucks were imaged with contrast-enhanced cone beam computed tomography with a flat panel detector C-arm system to visualize the carcinoma in the liver. In each animal, 3 insertions were performed, starting from the skin surface toward the hepatic carcinoma under image guidance. In 2 woodchucks, each end point of the insertion was confirmed with pathologic examination of a biopsy sample. While advancing the needle in the animals under image guidance such as fluoroscopy overlaid with cone beam computed tomography slice and ultrasound, optical spectra were acquired at the distal end of the needles. Optical tissue characterization was determined by translating the acquired optical spectra into clinical parameters such as blood, water, lipid, and bile fractions; tissue oxygenation levels; and scattering amplitude related to tissue density. The Kruskal-Wallis test was used to study the difference in the derived clinical parameters from the measurements performed within the healthy tissue and the hepatic carcinoma. Kurtoses were calculated to assess the dispersion of these parameters within the healthy and carcinoma tissues. RESULTS Blood and lipid volume fractions as well as tissue oxygenation and reduced scattering amplitude showed to be significantly different between the healthy part of the liver and the hepatic carcinoma (P < 0.05) being higher in normal liver tissue. A decrease in blood and lipid volume fractions and tissue oxygenation as well as an increase in scattering amplitude were observed when the tip of the needle crossed the margin from the healthy liver tissue to the carcinoma. The kurtosis for each derived clinical parameter was high in the hepatic tumor as compared with that in the healthy liver indicating intracarcinoma variability. CONCLUSIONS Tissue blood content, oxygenation level, lipid content, and tissue density all showed significant differences when the needle tip was guided from the healthy tissue to the carcinoma and can therefore be used to identify tissue boundaries during percutaneous image-guided interventions.

Development and characterization of chemical cochleostomy in the Guinea pig

OBJECTIVES Creation of an atraumatic, hearing-preservation cochleostomy is integral to the future of minimally invasive inner ear surgery. The goal of this study was to develop and characterize a novel chemical approach to cochleostomy. STUDY DESIGN Prospective animal study. SETTING Laboratory. METHODS Experimental animal study in which phosphoric acid gel (PAG) was used to decalcify the otic capsule in 25 Hartley guinea pigs. Five animals in each of 5 surgical groups were studied: (1) mechanically opening the auditory bulla alone, (2) PAG thinning of the basal turn otic capsule, leaving endosteum covered by a layer of bone, (3) micro-pick manual cochleostomy, (4) PAG chemical cochleostomy, exposing the endosteum, and (5) combined PAG/micro-pick cochleostomy, with initial chemical thinning and subsequent manual removal of the last osseous layer. Preoperative and postoperative auditory brainstem responses and otoacoustic emissions were obtained at 2, 6, 10, and 16 kHz. Hematoxylin and eosin-stained paraffin sections were compared. RESULTS Surgical and histologic findings confirmed that application of PAG provided reproducible local bone removal, and cochlear access was enabled. Statistically significant auditory threshold shifts were observed at 10 kHz (P = .048) and 16 kHz (P = .0013) following cochleostomy using PAG alone (group 4) and at 16 kHz using manual cochleostomy (group 3) (P = .028). No statistically significant, postoperative auditory threshold shifts were observed in the other groups, including PAG thinning with manual completion cochleostomy (group 5). CONCLUSION Hearing preservation cochleostomy can be performed in an animal model using a novel technique of thinning cochlear bone with PAG and manually completing cochleostomy.

Characterization of Enhancing MS Lesions by Dynamic Texture Parameter Analysis of Dynamic Susceptibility Perfusion Imaging

Purpose. The purpose of this study was to investigate statistical differences with MR perfusion imaging features that reflect the dynamics of Gadolinium-uptake in MS lesions using dynamic texture parameter analysis (DTPA). Methods. We investigated 51 MS lesions (25 enhancing, 26 nonenhancing lesions) of 12 patients. Enhancing lesions () were prestratified into enhancing lesions with increased permeability (EL+; ) and enhancing lesions with subtle permeability (EL−; ). Histogram-based feature maps were computed from the raw DSC-image time series and the corresponding texture parameters were analyzed during the inflow, outflow, and reperfusion time intervals. Results. Significant differences () were found between EL+ and EL− and between EL+ and nonenhancing inactive lesions (NEL). Main effects between EL+ versus EL− and EL+ versus NEL were observed during reperfusion (mainly in mean and standard deviation (SD): EL+ versus EL− and EL+ versus NEL), while EL− and NEL differed only in their SD during outflow. Conclusion. DTPA allows grading enhancing MS lesions according to their perfusion characteristics. Texture parameters of EL− were similar to NEL, while EL+ differed significantly from EL− and NEL. Dynamic texture analysis may thus be further investigated as noninvasive endogenous marker of lesion formation and restoration.

Three-dimensional culture and characterization of mononuclear cells from human bone marrow

BACKGROUND AIMS The diverse phenotypic changes and clinical and economic disadvantages associated with the monolayer expansion of bone marrow-derived mesenchymal stromal cells (MSCs) have focused attention on the development of one-step intraoperative cells therapies and homing strategies. The mononuclear cell fraction of bone marrow, inclusive of discrete stem cell populations, is not well characterized, and we currently lack suitable cell culture systems in which to culture and investigate the behavior of these cells. METHODS Human bone marrow-derived mononuclear cells were cultured within fibrin for 2 weeks with or without fibroblast growth factor-2 supplementation. DNA content and cell viability of enzymatically retrieved cells were determined at days 7 and 14. Cell surface marker profiling and cell cycle analysis were performed by means of multi-color flow cytometry and a 5-ethynyl-2'-deoxyuridine incorporation assay, respectively. RESULTS Total mononuclear cell fractions, isolated from whole human bone marrow, was successfully cultured in fibrin gels for up to 14 days under static conditions. Discrete niche cell populations including MSCs, pericytes and hematopoietic stem cells were maintained in relative quiescence for 7 days in proportions similar to that in freshly isolated cells. Colony-forming unit efficiency of enzymatically retrieved MSCs was significantly higher at day 14 compared to day 0; and in accordance with previously published works, it was fibroblast growth factor-2-dependant. CONCLUSIONS Fibrin gels provide a simple, novel system in which to culture and study the complete fraction of bone marrow-derived mononuclear cells and may support the development of improved bone marrow cell-based therapies.

Treatment planning aspects and Monte Carlo methods in proton therapy

Over the last years, the interest in proton radiotherapy is rapidly increasing. Protons provide superior physical properties compared with conventional radiotherapy using photons. These properties result in depth dose curves with a large dose peak at the end of the proton track and the finite proton range allows sparing the distally located healthy tissue. These properties offer an increased flexibility in proton radiotherapy, but also increase the demand in accurate dose estimations. To carry out accurate dose calculations, first an accurate and detailed characterization of the physical proton beam exiting the treatment head is necessary for both currently available delivery techniques: scattered and scanned proton beams. Since Monte Carlo (MC) methods follow the particle track simulating the interactions from first principles, this technique is perfectly suited to accurately model the treatment head. Nevertheless, careful validation of these MC models is necessary. While for the dose estimation pencil beam algorithms provide the advantage of fast computations, they are limited in accuracy. In contrast, MC dose calculation algorithms overcome these limitations and due to recent improvements in efficiency, these algorithms are expected to improve the accuracy of the calculated dose distributions and to be introduced in clinical routine in the near future.