Materials and Methods. 86 2.1 PARTICIPANTS 87 We used baseline measurements from a convenience sample of participants in previous (3) and 88 ongoing cohort studies investigating the effects of rehabilitation on balance responses (Table 1). PD 89 participants were mild-moderate with bilateral symptoms (Xxxxx and Xxxx stage 2-3 (13)). All 90 participants provided written informed consent and all study procedures were approved by Institutional 91 Review Boards at the Georgia Institute of Technology and Emory University.
Materials and Methods. Description of Simulations. We used a lattice-based stochastic patch occu- pancy model (26) to simulate spatially explicit dynamics of large numbers of species with dissimilar ecological traits. The model assumes that different lattice cells may represent different habitat type, and that habitat type across the lattice may be spatially correlated (26). In the present simulations, we assumed a high degree of spatial correlation in habitat type (parameter ω = 2 in ref. 26). Interspecific interactions are not modeled, but each species has distinct ecological traits defined by five parameters: colonization rate (c), extinction rate (e), average dispersal distance (1/α), mean phenotype (φ), and niche width (γ). Colonization rate and extinction rate control the probability of a species populating an unoccupied lattice cell and the probability of extinction in an occupied cell in unit time, respectively. These probabilities are affected by the habitat type of the cell in relation to the mean pheno- type and niche width of the species (26). Cells with habitat type close to the species mean phenotype support populations best, whereas niche width controls the sensitivity of the species to habitat type. In the present simu- lations, a species typically reaches the stochastic stationary state from a few dozen to a few hundred time steps (26). We run the simulations for at least 500 steps to ensure that most species had reached the quasi-stationary state. Increasing the simulation time did not qualitatively alter the results, al- though it resulted in a few additional extinctions as expected due to sto- xxxxxxxxxx. Further details of the simulations are given in SI Text.
Materials and Methods. Materials T-cell leukemia cells (Jurkat, Clone E6-1, ATCC® TIB152™) were provided by Leiden University Medical Centre (LUMC) as frozen 1-ml aliquots at a total cell concentration of 107 cells/ml, and were stored at -140 oC in the freezer prior to usage. The Jurkat cells were formulated in high-glucose RPMI 1640 (RPMI medium; ThermoFisher, Waltham, USA) supplemented with 10% fetal bovine serum (Life Technologies, USA) and 10% dimethyl sulfoxide (DMSO) (Life Technologies, USA). Dynabeads Human T-Activator CD3/CD28 for T Cell Expansion and Activation and low-protein binding collection tubes were purchased from ThermoFisher (Waltham, USA). Sterile 5-ml Eppendorf tubes were purchased from VWR (Ismaning, Germany). Sample preparation Jurkat cells used in this study as model T- cells were thawed and freshly prepared in RPMI medium prior to analysis. Frozen cell aliquots were thawed at 36oC and resuspended in ca. 40 ml of RPMI medium. To remove residual fetal bovine serum and DMSO, the cell suspension was centrifuged at 300 rcf for 10 minutes at 20oC. The supernatant was removed and the pellet was resuspended in 10 ml of RPMI medium, unless otherwise stated. The mean concentration of (live and dead) cells was 477,188 ± 85,914 per ml with a mean viability of 81% ± 9% (n=8) as determined by hemocytometry (described below), unless otherwise stated. Cell containing samples were measured up to 4 hours post thawing, a time window during which the cell viability was not affected (data not shown). Dynabeads were diluted to an intermediate stock concentration of 106 beads/ml (based on the dilution factor of the nominal Dynabead concentration) in RPMI medium and stored at 2-8 oC for up to 1 month. The required volume of the intermediate stock was added to cell samples to reach the desired Dynabead concentrations. Reference concentration of Dynabeads stated in the results section is the expected concentration of Dynabeads in the sample derived from dilution calculations and the original bead concentration stated by the manufacturer. It must be noted that the manufacturer does Particulates impurities in CBMPs traced by FIM-CNN not use FlowCam for quantification of Dynabeads; therefore, a systematic deviation between reference concentrations and measured concentrations should be anticipated. Hemocytometry Cell viability and total cell concentration were determined by using a Bright-Line hemocytometer glass (Merck, Darmstadt, Germany) and an Axiostar Plus microscope (Zeiss, Jena...
Materials and Methods. Participants and Recruitment Participants included two hundred and six Fédération Interna- tionale de Football Association (FIFA) registered contracted professional football players (x±s; age=24.1±5.4-years, body mass=78.8±8.4 kg, stretched stature=180.1±7.0 cm and whole body density=1.075±0.010 (g.ml-1) who were all over 18-years of age and free from disease. They were recruited from eight profes- sional football clubs that represented Barclays Premiership, npow- er Championship, npower League One, npower League Two and Blue Square Premier Leagues during the 2007-2008, 2008-2009 and 2009-2010 playing seasons. All participants completed a health screening questionnaire and agreed to take part in the study by giv- ing their informed consent. Ethical approval was granted from the University of Gloucestershire (UK) Research Ethics Committee (MIL-UOGREC/10).
Materials and Methods. The Ethics Committee of Xxxx University approved the study protocol. Informed consent was obtained from all participants or participant´s guardian, after the nature of the procedures had been fully explained. This is a prospective observational study of real-time live ultrasound examinations of adnexal masses. Consecutive patients referred for an ultrasound examination and found to have an adnexal mass judged to need surgical removal were scanned according to the research protocol by sonologist 1 (PS) as part of the clinical ultrasound examination. A second ultrasound examination was carried out before surgery by sonologist 2 (LV). Both examiners used the standardized IOTA examination and measurement technique and the IOTA terminology (11) to describe their ultrasound findings and noted their results in a dedicated paper form. Sonologist 2 was blinded to the results of sonologist 1. Information on the clinical variables included in LR1 and LR2 (personal history of ovarian cancer, current hormonal therapy, age of the patient), was obtained at the preoperative ultrasound examination by sonologist 2. All patients were operated on within 90 days after the preoperative ultrasound examination performed by sonologist 2. The excised tissues underwent histological examination and tumors were classified according to the criteria recommended by the International Federation of Gynecology and Obstetrics (16). Borderline tumors were classified as malignant. The patients were examined in the lithotomy position with an empty urinary bladder (11). Abdominal ultrasound examination was added when needed. The ultrasound variables assessed with regard to interobserver reproducibility are shown in Table 1. The size of the lesion and that of its largest solid component were measured (largest diameter and mean of three orthogonal diameters) using calipers on the frozen ultrasound image. A color score was assigned on the basis of subjective assessment of the color content of the tumor scan at power Doppler ultrasound examination. A color score of 1 indicates absence of color Doppler signals, a color score of 2 a minimal amount of color Doppler signals, a color score of 3 a moderate amount of color Doppler signals and a color score of 4 a large amount of color Doppler signals in the tumor (11). The ultrasound systems used were GE Voluson 730 Expert or GE Voluson E8 (GE Healthcare, Xxxx, Austria) with a 5–9-MHz transvaginal transducer. For power Doppler ultrasound examinations the f...
Materials and Methods. Generation of molecular maps We generated detailed molecular maps for the TCR and TLR5 signaling pathways using the software CellDesigner (xxxx://xxx.xxxxxxxxxxxx.xxx/). To do this, we relied on data gathered from the literature and from databases such as KEGG (xxxx://xxx.xxxxxx.xx/kegg/), Reactome (xxxx://xxx.xxxxxxxx.xxx/), and DC-ATLAS (xxxx://xxxxxxxxxxxxxx.xxxxx.xx/DCATLAS.php). In CellDesigner, there are three possible types of Components, which are Species, Reactions, and Compartments. A Species represents a protein, a complex, or some other molecule in a biochemical or regulatory network. On the other hand, a Reaction can be a chemical reaction, a physical interaction between Species, or a regulatory relation between genes. Finally, a Compartment represents a container for other components, such as a cell or an intracellular compartment. All species and reactions of the molecular maps are annotated with textual comments and hyperlinks to record the supporting information and its source (see Files 1 and 3). Generation of logical models We built our logical models using the software GINsim (xxx.xxxxxx.xxx), relying on the molecular maps previously generated. XXXxxx implements the multivalued logical modeling formalism introduced by Xxxxxx and D’Ari (80). This formalism relies on the delineation of a regulatory graph (LRG), where each component (protein or more abstract biological function) is represented by a logical node (taking the values 0 or 1, or additional values when justified), and each influence (activation or inhibition) between a pair of components is represented by a signed arc. Next, a logical rule is assigned to each node in the network, which determines its activation level according to the levels of its regulators. These logical rules involve literals (component values) and the logical operators AND, OR and NOT (81, 82). We confirmed the expression of all model components in unstimulated naïve CD4+ T cells using RNA-seq and ChIP-seq data reported by the BLUEPRINT consortium (Table S1). A gene was considered expressed if it could be associated with regions with chromatin marks denoting active promoters or enhancers, together with an expression value of at least 1 RPKM (Reads Per Kilobase of transcripts, per Million mapped). ChIP-seq peaks of histone modifications were given as input to the software ChromHMM (xxxx://xxxxxxx.xxx.xxx/ChromHMM/) (with the parameters BinerizeBed –center option, assembly hg38; LearnModel 10 states) to segmen...
Materials and Methods. Patients After approval by our institutional ethics committee, untreated pa- tients with biopsy-confirmed SCCA of the upper aerodigestive tract (the oral cavity, oropharynx, larynx, and hypopharynx) were pro- spectively enrolled into a study investigating the potential of CTP for induction-chemotherapy monitoring. Written informed consent was obtained from all the participants before the study. Patients were included if they met the following criteria: stage III or IVA-B tumor according to tumor node metastasis system,11 ≥18 and ≤75 years of age, Eastern Cooperative Oncology Group perfor- Table 1: Enrolled patients and tumor characteristics No. % Total 30 100 Site Oral cavity 1 3.3 Oropharynx 17 56.7 Hypopharynx-larynx 12 40 IVA 20 66.7 IVB TNM 4 13.3 T1N3M0 1 3.3 T1N2M0 1 3.3 T2N1M0 3 10 T2N2M0 11 36.7 T3N0M0 3 10 T3N2M0 3 10 T4N0M0 2 6.7 T4N1M0 3 10 T4N2M0 2 6.7 T4N3M0 1 3.3 constant table position, 1-second gantry rotation time, 120 kilovolt (peak), and 100 mA. Images were acquired every second for 45 sec- onds for first-pass enhancement assessment (first-pass scanning) and every 10 seconds up to 122 seconds (prolonged scanning) for late- enhancement assessment. Immediately after completion of CTP scanning, diagnostic CT of the head and neck was performed by using 16 × 0.625 (10 mm) collimation; section thickness and increment, 1.25 mm; standard re- construction filter; 180 mA; 120 kV(p); rotation time, 0.6 seconds; speed, 9.38 mm/rotation; FOV, 18 cm; intravenous contrast material (Iomeron [400 mg I/mL]); injection rate, 2 mL/s; volume, 90 mL; acquisition delay, 70 seconds). The average total examination dose- length product for the combined CTP and diagnostic CT of the head and neck was 1196.13 mGy cm. Preliminary noncontrast CT of the head and neck and CTP and diagnostic CT of the head and neck were performed by using quiet respiration. The patients were warned to avoid swallowing and taking a deep breath when experiencing the “hot flush” resulting from the rapid bolus of iodinated contrast material. xxxxx status from zero to 2, white blood cell count ≥3000 mm3, neutrophil count ≥1500 mm3, platelet count ≥100,000 mm3, serum creatinine level ≤1 mg/dL, and serum bilirubin level ≤1.25 mg/dL. Patients with previous neoplasms in other sites not treatable for re- covery, pregnancy, or allergy to iodinated contrast media were ex- cluded from the study. Local tumor staging was based on clinical evaluation by endos- copy and contrast-enhanced CT of the head a...
Materials and Methods. Twenty-six patients with SCCA were enrolled in this prospective study and underwent CTP. Data were analyzed by 2 expert readers and by an inexperienced reader for interobserver agreement and by the 2 expert readers for intraobserver agreement assessment, by using the ECA ipsilateral to tumor site as arterial input. All 3 readers repeated their analysis by using the ECA contralateral to tumor site as arterial input. Inter- and intraobserver agreement was assessed by using the Xxxxx-Xxxxxx approach; CTP measurements by using ipsilateral-versus-contralateral or left-versus-right ECA were compared by using the Wilcoxon signed rank test. RESULTS: The geometric mean of the ratios (95% limits of agreement) for inter- and intraobserver agreement ranged from 0.96 (0.75–1.23) to 1.00 (0.92–1.10) for blood flow (BF), from 0.88 (0.63–1.21) to 1.00 (0.88 –1.14) for blood volume (BV), from 0.96 (0.64 –1.44) to 0.98 (0.76 –1.27) for mean transit time (MTT), and from 0.85 (0.41–1.76) to 1.14 (0.70 –1.86) for permeability surface area product (PS). Significantly higher tumor PS and MTT for 2 readers and lower tumor BF for 1 of 3 readers were observed when the arterial input was placed in the left ECA. HEAD & NECK ORIGINAL RESEARCH CONCLUSIONS: BF, BV, and MTT demonstrated higher inter- and intraobserver agreement than PS. The selection of arterial input, right-versus-left ECA, may determine changes in CTP measurements in patients with SCCA of the upper aerodigestive tract. C T perfusion (CTP) of squamous cell carcinoma (SCCA) of the upper aerodigestive tract has demonstrated potential for tumor diagnosis, with significantly different perfusion pa- rameters compared with normal structures,1,2 and therapy monitoring, correlating with endoscopic response following induction chemotherapy.3 In addition, CTP seems to predict local outcome in patients with SCCA of the upper aerodiges- tive tract undergoing radiation therapy4 and induction
Materials and Methods. Materials The humanized mAb, isotype IgG1, trastuzumab (Herzuma®) was purchased from Leiden University Medical Centre (Leiden, The Netherlands). Pharmaceutical-grade surcose was purchased from VWR (Bruchsal, Germany) and RPMI 1640 medium was supplied by ThermoFisher Scientific (Waltham, USA). Polyvinylidene fluoride (PDVF) syringe filters with a pore size of 0.22 µm were purchased from Merck (Darmstadt, Germany). Labeled monoclonal antibodies used for flow cytometry were purchased from BD Biosciences (New Jersey, USA). A cytokine and chemokine multiplex assay (U-PLEX) was purchased from MSD (Rockville, USA). Highly purified water (conductivity: 18.2 mUΩ*cm) obtained from a Milli- Q® Advantage A10 system (Merck, Newark, USA) was used throughout the study. Preparation of NPIs Preparation of NPIs was performed as described before.7 Briefly, a 50% w/v of sucrose solution in water was prepared and submitted to an ultra-/dia-filtration process by using a Minimate II Tangential Flow Filtration (TFF) system (Pall, Crailsheim, Germany) equipped with a 30-kDa TFF capsule (Pall). Diafiltration of the sucrose solution was performed against 20 volumes of water. Further, the sucrose-free retentate (confirmed by DLS, data not shown) was upconcentrated 1000-fold and filtered by using a 0.22-µm PDVF filter. Immunological evaluation in vitro of NPIs Upconcentrated stock retentate containing NPIs at a concentration 1x1011 particles/mL (as determined by NTA) was stored at -80 oC until needed. Water without addition of sucrose was submitted to the same process described above and served as a control sample (Ctl) throughout the study. Sample preparation Stock suspensions of NPIs (two-fold higher in concentration compared to the target concentration) were prepared in RPMI medium and spiked into trastuzumab samples to target concentrations stated in the results section. Trastuzumab was diluted with RPMI medium to 2 mg/mL. Further addition of NPIs and RPMI medium was performed to reach a final protein concentration of 1 mg/mL. Trastuzumab was incubated in presence of NPIs or control (water) for 1 hour at room temperature. The formulations were added to moDC suspensions to reach a target IgG concentration of 0.1 mg/mL.19 Nanoparticle tracking analysis (NTA) NanoSight (Model LM20, Malvern Instrument, Malvern, UK) instrument equipped with a 405-nm laser (blue) was used to quantify and determine the size of nanometer sized particles (size range ̴50 – 1000 nm). Samples were injected in...
Materials and Methods. 2.1 Study population A total of 137 consecutive CTO patients, diagnosed by coronary angiography (CAG) from May 2015 to June 2017, were enrolled in this study. As shown in Fig. 1, patients were excluded due to the contradictions of CMR (N = 33) or incomplete CMR data (N = 1). The rest of the patients were referred to echocardiography examination, and further excluded due to poor-quality images (N = 5), incomplete 2DTTE data (N = 4), or lack of 2DTTE images (N = 6), leaving 88 patients eligible for the study. All enrolled patients underwent both echocardiography and CMR imaging within the same week. Coronary artery interventions were not performed until both types of imaging were finished.
