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- ItemGlycerylphytate as an ionic crosslinker for 3D printing of multi-layered scaffolds with improved shape fidelity and biological features(London : Royal Society of Chemistry, 2020) Mora-Boza, A.; Włodarczyk-Biegun, M.K.; Del Campo, A.; Vázquez-Lasa, B.; Román, J.S.The fabrication of intricate and long-term stable 3D polymeric scaffolds by a 3D printing technique is still a challenge. In the biomedical field, hydrogel materials are very frequently used because of their excellent biocompatibility and biodegradability, however the improvement of their processability and mechanical properties is still required. This paper reports the fabrication of dual crosslinked 3D scaffolds using a low concentrated (<10 wt%) ink of gelatin methacryloyl (GelMA)/chitosan and a novel crosslinking agent, glycerylphytate (G1Phy) to overcome the current limitations in the 3D printing field using hydrogels. The applied methodology consisted of a first ultraviolet light (UV) photopolymerization followed by a post-printing ionic crosslinking treatment with G1Phy. This crosslinker provides a robust framework and avoids the necessity of neutralization with strong bases. The blend ink showed shear-thinning behavior and excellent printability in the form of a straight and homogeneous filament. UV curing was undertaken simultaneously to 3D deposition, which enhanced precision and shape fidelity (resolution ≈150 μm), and prevented the collapse of the subsequent printed layers (up to 28 layers). In the second step, the novel G1Phy ionic crosslinker agent provided swelling and long term stability properties to the 3D scaffolds. The multi-layered printed scaffolds were mechanically stable under physiological conditions for at least one month. Preliminary in vitro assays using L929 fibroblasts showed very promising results in terms of adhesion, spreading, and proliferation in comparison to other phosphate-based traditional crosslinkers (i.e. TPP). We envision that the proposed combination of the blend ink and 3D printing approach can have widespread applications in the regeneration of soft tissues.
- ItemDenoising for Improved Parametric MRI of the Kidney: Protocol for Nonlocal Means Filtering(Totowa, NJ : Humana Press, 2021) Starke, Ludger; Tabelow, Karsten; Niendorf, Thoralf; Pohlmann, Andreas; Pohlmann, Andreas; Niendorf, ThoralfIn order to tackle the challenges caused by the variability in estimated MRI parameters (e.g., T2* and T2) due to low SNR a number of strategies can be followed. One approach is postprocessing of the acquired data with a filter. The basic idea is that MR images possess a local spatial structure that is characterized by equal, or at least similar, noise-free signal values in vicinities of a location. Then, local averaging of the signal reduces the noise component of the signal. In contrast, nonlocal means filtering defines the weights for averaging not only within the local vicinity, bur it compares the image intensities between all voxels to define “nonlocal” weights. Furthermore, it generally compares not only single-voxel intensities but small spatial patches of the data to better account for extended similar patterns. Here we describe how to use an open source NLM filter tool to denoise 2D MR image series of the kidney used for parametric mapping of the relaxation times T2* and T2. This chapter is based upon work from the COST Action PARENCHIMA, a community-driven network funded by the European Cooperation in Science and Technology (COST) program of the European Union, which aims to improve the reproducibility and standardization of renal MRI biomarkers.
- ItemConsequences of nano and microplastic exposure in rodent models: the known and unknown(London : BioMed Central, 2022) da Silva Brito, Walison Augusto; Mutter, Fiona; Wende, Kristian; Cecchini, Alessandra Lourenco; Schmidt, Anke; Bekeschus, SanderThe ubiquitous nature of micro- (MP) and nanoplastics (NP) is a growing environmental concern. However, their potential impact on human health remains unknown. Research increasingly focused on using rodent models to understand the effects of exposure to individual plastic polymers. In vivo data showed critical exposure effects depending on particle size, polymer, shape, charge, concentration, and exposure routes. Those effects included local inflammation, oxidative stress, and metabolic disruption, leading to gastrointestinal toxicity, hepatotoxicity, reproduction disorders, and neurotoxic effects. This review distillates the current knowledge regarding rodent models exposed to MP and NP with different experimental designs assessing biodistribution, bioaccumulation, and biological responses. Rodents exposed to MP and NP showed particle accumulation in several tissues. Critical responses included local inflammation and oxidative stress, leading to microbiota dysbiosis, metabolic, hepatic, and reproductive disorders, and diseases exacerbation. Most studies used MP and NP commercially provided and doses higher than found in environmental exposure. Hence, standardized sampling techniques and improved characterization of environmental MP and NP are needed and may help in toxicity assessments of relevant particle mixtures, filling knowledge gaps in the literature.
- ItemBreast Cancer Stem Cell–Derived Tumors Escape from γδ T-cell Immunosurveillance In Vivo by Modulating γδ T-cell Ligands(Philadelphia, Pa. : AACR, 2023) Raute, Katrin; Strietz, Juliane; Parigiani, Maria Alejandra; Andrieux, Geoffroy; Thomas, Oliver S.; Kistner, Klaus M.; Zintchenko, Marina; Aichele, Peter; Hofmann, Maike; Zhou, Houjiang; Weber, Wilfried; Boerries, Melanie; Swamy, Mahima; Maurer, Jochen; Minguet, SusanaThere are no targeted therapies for patients with triple-negative breast cancer (TNBC). TNBC is enriched in breast cancer stem cells (BCSC), which play a key role in metastasis, chemoresistance, relapse, and mortality. γδ T cells hold great potential in immunotherapy against cancer and might provide an approach to therapeutically target TNBC. γδ T cells are commonly observed to infiltrate solid tumors and have an extensive repertoire of tumor-sensing mechanisms, recognizing stress-induced molecules and phosphoantigens (pAgs) on transformed cells. Herein, we show that patient-derived triple-negative BCSCs are efficiently recognized and killed by ex vivo expanded γδ T cells from healthy donors. Orthotopically xenografted BCSCs, however, were refractory to γ δ T-cell immunotherapy. We unraveled concerted differentiation and immune escape mechanisms: xenografted BCSCs lost stemness, expression of γ δ T-cell ligands, adhesion molecules, and pAgs, thereby evading immune recognition by γ δ T cells. Indeed, neither promigratory engineered γ δ T cells, nor anti–PD-1 checkpoint blockade, significantly prolonged overall survival of tumor-bearing mice. BCSC immune escape was independent of the immune pressure exerted by the γ δ T cells and could be pharmacologically reverted by zoledronate or IFNα treatment. These results pave the way for novel combinatorial immunotherapies for TNBC.
- ItemRepeated exposure of the oral mucosa over 12 months with cold plasma is not carcinogenic in mice(London : Nature Publishing Group, 2021) Evert, K.; Kocher, T.; Schindler, A.; Müller, M.; Müller, K.; Pink, C.; Holtfreter, B.; Schmidt, A.; Dombrowski, F.; Schubert, A.; von Woedtke, T.; Rupf, S.; Calvisi, D. F.; Bekeschus, S.; Jablonowski, L.Peri-implantitis may result in the loss of dental implants. Cold atmospheric pressure plasma (CAP) was suggested to promote re-osseointegration, decrease antimicrobial burden, and support wound healing. However, the long-term risk assessment of CAP treatment in the oral cavity has not been addressed. Treatment with two different CAP devices was compared against UV radiation, carcinogen administration, and untreated conditions over 12 months. Histological analysis of 406 animals revealed that repeated CAP exposure did not foster non-invasive lesions or squamous cell carcinoma (SCCs). Carcinogen administration promoted non-invasive lesions and SCCs. Molecular analysis by a qPCR screening of 144 transcripts revealed distinct inflammatory profiles associated with each treatment regimen. Interestingly, CAP treatment of carcinogen-challenged mucosa did not promote but instead left unchanged or reduced the proportion of non-invasive lesions and SCC formation. In conclusion, repeated CAP exposure of murine oral mucosa was well tolerated, and carcinogenic effects did not occur, motivating CAP applications in patients for dental and implant treatments in the future.
- ItemTumor cytotoxicity and immunogenicity of a novel V-jet neon plasma source compared to the kINPen(London : Nature Publishing Group, 2021) Miebach, Lea; Freund, Eric; Horn, Stefan; Niessner, Felix; Sagwal, Sanjeev Kumar; von Woedtke, Thomas; Emmert, Steffen; Weltmann, Klaus-Dieter; Clemen, Ramona; Schmidt, Anke; Gerling, Torsten; Bekeschus, SanderRecent research indicated the potential of cold physical plasma in cancer therapy. The plethora of plasma-derived reactive oxygen and nitrogen species (ROS/RNS) mediate diverse antitumor effects after eliciting oxidative stress in cancer cells. We aimed at exploiting this principle using a newly designed dual-jet neon plasma source (Vjet) to treat colorectal cancer cells. A treatment time-dependent ROS/RNS generation induced oxidation, growth retardation, and cell death within 3D tumor spheroids were found. In TUM-CAM, a semi in vivo model, the Vjet markedly reduced vascularized tumors' growth, but an increase of tumor cell immunogenicity or uptake by dendritic cells was not observed. By comparison, the argon-driven single jet kINPen, known to mediate anticancer effects in vitro, in vivo, and in patients, generated less ROS/RNS and terminal cell death in spheroids. In the TUM-CAM model, however, the kINPen was equivalently effective and induced a stronger expression of immunogenic cancer cell death (ICD) markers, leading to increased phagocytosis of kINPen but not Vjet plasma-treated tumor cells by dendritic cells. Moreover, the Vjet was characterized according to the requirements of the DIN-SPEC 91315. Our results highlight the plasma device-specific action on cancer cells for evaluating optimal discharges for plasma cancer treatment.
- ItemRestricted differentiative capacity of Wt1-expressing peritoneal mesothelium in postnatal and adult mice(London : Nature Publishing Group, 2021) Wilm, Thomas P.; Tanton, Helen; Mutter, Fiona; Foisor, Veronica; Middlehurst, Ben; Ward, Kelly; Benameur, Tarek; Hastie, Nicholas; Wilm, BettinaPreviously, genetic lineage tracing based on the mesothelial marker Wt1, appeared to show that peritoneal mesothelial cells have a range of differentiative capacities and are the direct progenitors of vascular smooth muscle in the intestine. However, it was not clear whether this was a temporally limited process or continued throughout postnatal life. Here, using a conditional Wt1-based genetic lineage tracing approach, we demonstrate that the postnatal and adult peritoneum covering intestine, mesentery and body wall only maintained itself and failed to contribute to other visceral tissues. Pulse-chase experiments of up to 6 months revealed that Wt1-expressing cells remained confined to the peritoneum and failed to differentiate into cellular components of blood vessels or other tissues underlying the peritoneum. Our data confirmed that the Wt1-lineage system also labelled submesothelial cells. Ablation of Wt1 in adult mice did not result in changes to the intestinal wall architecture. In the heart, we observed that Wt1-expressing cells maintained the epicardium and contributed to coronary vessels in newborn and adult mice. Our results demonstrate that Wt1-expressing cells in the peritoneum have limited differentiation capacities, and that contribution of Wt1-expressing cells to cardiac vasculature is based on organ-specific mechanisms.
- ItemAccurate in vivo tumor detection using plasmonic-enhanced shifted-excitation Raman difference spectroscopy (SERDS)(Wyoming, NSW : Ivyspring, 2021) Strobbia, Pietro; Cupil-Garcia, Vanessa; Crawford, Bridget M.; Fales, Andrew M.; Pfefer, T. Joshua; Liu, Yang; Maiwald, Martin; Sumpf, Bernd; Vo-Dinh, TuanFor the majority of cancer patients, surgery is the primary method of treatment. In these cases, accurately removing the entire tumor without harming surrounding tissue is critical; however, due to the lack of intraoperative imaging techniques, surgeons rely on visual and physical inspection to identify tumors. Surface-enhanced Raman scattering (SERS) is emerging as a non-invasive optical alternative for intraoperative tumor identification, with high accuracy and stability. However, Raman detection requires dark rooms to work, which is not consistent with surgical settings. Methods: Herein, we used SERS nanoprobes combined with shifted-excitation Raman difference spectroscopy (SERDS) detection, to accurately detect tumors in xenograft murine model. Results: We demonstrate for the first time the use of SERDS for in vivo tumor detection in a murine model under ambient light conditions. We compare traditional Raman detection with SERDS, showing that our method can improve sensitivity and accuracy for this task. Conclusion: Our results show that this method can be used to improve the accuracy and robustness of in vivo Raman/SERS biomedical application, aiding the process of clinical translation of these technologies. © The author(s). This is an open access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/). See http://ivyspring.com/terms for full terms and conditions.