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- Beal, Jacob, et al.
(författare)
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Robust estimation of bacterial cell count from optical density
- 2020
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Ingår i: Communications Biology. - : Springer Science and Business Media LLC. - 2399-3642. ; 3:1
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Tidskriftsartikel (refereegranskat)abstract
- Optical density (OD) is widely used to estimate the density of cells in liquid culture, but cannot be compared between instruments without a standardized calibration protocol and is challenging to relate to actual cell count. We address this with an interlaboratory study comparing three simple, low-cost, and highly accessible OD calibration protocols across 244 laboratories, applied to eight strains of constitutive GFP-expressing E. coli. Based on our results, we recommend calibrating OD to estimated cell count using serial dilution of silica microspheres, which produces highly precise calibration (95.5% of residuals <1.2-fold), is easily assessed for quality control, also assesses instrument effective linear range, and can be combined with fluorescence calibration to obtain units of Molecules of Equivalent Fluorescein (MEFL) per cell, allowing direct comparison and data fusion with flow cytometry measurements: in our study, fluorescence per cell measurements showed only a 1.07-fold mean difference between plate reader and flow cytometry data.
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| 2. |
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| 3. |
- Pang, Mei-Fong
(författare)
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Studies of the tumor-vasculature interface : role of TGF-beta 1-induced epithelial to mesenchymal transition
- 2013
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Tumor metastasis is a complex multistep process. Among key steps that occur during metastatic spread are acquisition of tumor cell motility, intravasation of tumor cells into blood or lymphatic vessels and extravasation of tumor cells at distal sites. However, the precise mechanisms that govern these metastatic steps remain elusive. This thesis aimed to bridge the fields of tumor and vascular biology to provide new insights into the metastatic process. Results are presented indicating a role of the cytokine transforming growth factor beta (TGF-β) in activating breast cancer cells for dissemination through the lymphatic system through re-activation of a latent development process termed epithelial to mesenchymal transition (EMT). Furthermore, essential roles of the coxsackie-and adenovirus receptor (CAR) for lymph vessel development, and the sphingosine-1-phosphate receptor (S1PR1) for blood vessel stabilization are presented. We expect the findings to have impact on our understanding of the interface between tumor and vascular biology and to influence future strategies to target cancer metastasis. In paper I, we present data identifying an essential role of CAR for normal development of lymphatic vessels in the mouse. We show that genetic deletion of the CAR gene (Cxadr) from E12.5 during mouse development leads to subcutaneous edema, hemorrhage and embryonic death. The lymphatic vessels in CAR-deficient mice were dilated and structurally abnormal with the presence of gaps and holes at lymphatic endothelial cell-cell junctions. In addition, blood-filled lymphatics were observed in CAR-deficient mice suggesting an incomplete separation between the blood and lymphatic vascular systems. The data demonstrate that CAR plays a crucial role in the development of lymphatic vasculature in mice through formation of lymphatic endothelial cell-cell junctions. In paper II, we demonstrate that S1PR1 plays critical role in suppressing angiogenesis and promoting vascular stability during mouse development. S1PR1 signaling promotes cell-cell adhesion and prevents sprouting angiogenesis whereas S1PR1-deficiency leads to hypersprouting angiogenesis. These data suggest that S1PR1 signaling might protect developing blood vessels from abnormal angiogenic signals through promotion of vascular stability. In paper III, we show that TGF-β-induced EMT promotes chemotactic migration of tumor cells through the lymphatic system by mediating crosstalk between tumor cells and lymphatic endothelial cells through the chemokine receptor 7 (CCR7) and its chemokine ligand, CCL21. Reversal of EMT process through p38 MAPK inhibition inhibited tumor cell invasion in vitro and migration towards the lymphatics in vivo suggesting that p38 MAPK inhibition may be a useful therapeutic approach to inhibit tumor cell dissemination through the lymphatic system. In paper IV, we describe development of a novel co-culture system to study tumor cell migration and interaction with lymphatic endothelial cells within a 3-dimensional matrix component. This assay allows manipulation of tumor properties or matrix components and can be used as a platform to screen for pharmacological agents which inhibit tumor-endothelial interactions.
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| 4. |
- Porsbjerg, Celeste M., et al.
(författare)
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Association between pre-biologic T2-biomarker combinations and response to biologics in patients with severe asthma
- 2024
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Ingår i: Frontiers in Immunology. - : Frontiers Media S.A.. - 1664-3224. ; 15
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Tidskriftsartikel (refereegranskat)abstract
- Background: To date, studies investigating the association between pre-biologic biomarker levels and post-biologic outcomes have been limited to single biomarkers and assessment of biologic efficacy from structured clinical trials.Aim: To elucidate the associations of pre-biologic individual biomarker levels or their combinations with pre-to-post biologic changes in asthma outcomes in real-life.Methods: This was a registry-based, cohort study using data from 23 countries, which shared data with the International Severe Asthma Registry (May 2017-February 2023). The investigated biomarkers (highest pre-biologic levels) were immunoglobulin E (IgE), blood eosinophil count (BEC) and fractional exhaled nitric oxide (FeNO). Pre- to approximately 12-month post-biologic change for each of three asthma outcome domains (i.e. exacerbation rate, symptom control and lung function), and the association of this change with pre-biologic biomarkers was investigated for individual and combined biomarkers.Results: Overall, 3751 patients initiated biologics and were included in the analysis. No association was found between pre-biologic BEC and pre-to-post biologic change in exacerbation rate for any biologic class. However, higher pre-biologic BEC and FeNO were both associated with greater post-biologic improvement in FEV1 for both anti-IgE and anti-IL5/5R, with a trend for antiI-IL4R alpha. Mean FEV1 improved by 27-178 mL post-anti-IgE as pre-biologic BEC increased (250 to 1000 cells/mu L), and by 43-216 mL and 129-250 mL post-anti-IL5/5R and - anti- IL4R alpha, respectively along the same BEC gradient. Corresponding improvements along a FeNO gradient (25-100 ppb) were 41-274 mL, 69-207 mL and 148-224 mL for anti-IgE, anti-IL5/5R, and anti-IL4R alpha, respectively. Higher baseline BEC was also associated with lower probability of uncontrolled asthma (OR 0.392; p=0.001) post-biologic for anti-IL5/5R. Pre-biologic IgE was a poor predictor of subsequent pre-to-post-biologic change for all outcomes assessed for all biologics. The combination of BEC + FeNO marginally improved the prediction of post-biologic FEV1 increase (adjusted R-2: 0.751), compared to BEC (adjusted R-2: 0.747) or FeNO alone (adjusted R-2: 0.743) (p=0.005 and <0.001, respectively); however, this prediction was not improved by the addition of IgE.Conclusions: The ability of higher baseline BEC, FeNO and their combination to predict biologic-associated lung function improvement may encourage earlier intervention in patients with impaired lung function or at risk of accelerated lung function decline.
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