Integrative Biology最新文献

{"title":"Spatiotemporal model of cellular mechanotransduction via Rho and YAP.","authors":"Javor K Novev,&nbsp;Mathias L Heltberg,&nbsp;Mogens H Jensen,&nbsp;Amin Doostmohammadi","doi":"10.1093/intbio/zyab012","DOIUrl":"https://doi.org/10.1093/intbio/zyab012","url":null,"abstract":"<p><p>How cells sense and respond to mechanical stimuli remains an open question. Recent advances have identified the translocation of Yes-associated protein (YAP) between nucleus and cytoplasm as a central mechanism for sensing mechanical forces and regulating mechanotransduction. We formulate a spatiotemporal model of the mechanotransduction signalling pathway that includes coupling of YAP with the cell force-generation machinery through the Rho family of GTPases. Considering the active and inactive forms of a single Rho protein (GTP/GDP-bound) and of YAP (non-phosphorylated/phosphorylated), we study the cross-talk between cell polarization due to active Rho and YAP activation through its nuclear localization. For fixed mechanical stimuli, our model predicts stationary nuclear-to-cytoplasmic YAP ratios consistent with experimental data at varying adhesive cell area. We further predict damped and even sustained oscillations in the YAP nuclear-to-cytoplasmic ratio by accounting for recently reported positive and negative YAP-Rho feedback. Extending the framework to time-varying mechanical stimuli that simulate cyclic stretching and compression, we show that the YAP nuclear-to-cytoplasmic ratio's time dependence follows that of the cyclic mechanical stimulus. The model presents one of the first frameworks for understanding spatiotemporal YAP mechanotransduction, providing several predictions of possible YAP localization dynamics, and suggesting new directions for experimental and theoretical studies.</p>","PeriodicalId":80,"journal":{"name":"Integrative Biology","volume":"13 8","pages":"197-209"},"PeriodicalIF":2.5,"publicationDate":"2021-08-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"39198597","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Bioengineered 3D electrospun nanofibrous scaffold with human liver cells to study alcoholic liver disease in vitro.","authors":"Prativa Das,&nbsp;Michael D DiVito,&nbsp;Jason A Wertheim,&nbsp;Lay Poh Tan","doi":"10.1093/intbio/zyab011","DOIUrl":"https://doi.org/10.1093/intbio/zyab011","url":null,"abstract":"<p><p>Alcohol injury induces hepatic fibrosis which gradually progresses to cirrhosis, sometimes may lead to liver cancer. Animal models are less efficient in mimicking responses of human liver cells, whereas in vitro models discussed so far are majorly based on rodent cells. In this work, a coculture of primary human hepatocytes (PHHs) with LX-2 cells was established on the unmodified (C:F_0:0), collagen-I modified (C:F_1:0), fibronectin modified (C:F_0:1) and 3:1 collagen-I to fibronectin modified (C:F_3:1) 3D electrospun fibrous scaffolds. The effect of alcohol injury was evaluated on this cell-scaffold model at 0-40 μl/ml alcohol concentrations over 14 days of culture period by using the gold standard sandwich culture as the control. Among all the culture groups, C:F_3:1 scaffold was able to maintain translational and transcriptional properties of human liver cells at all concentrations of alcohol treatment. The study reveals that, PHHs on C:F_3:1 were able to maintain ~4-fold and ~1.6-fold higher secretion of albumin than the gold standard sandwich culture on Day 3 and Day 7, respectively. When treated with alcohol, at concentrations of 20 and 40 μl/ml, albumin secretion was also observed to be higher (~2-fold) when compared to the gold standard sandwich culture. Again as expected, in C:F_3:1 culture group on 40 μl/ml alcohol treatment, albumin gene expression decreased by ~2-fold due to alcohol toxicity, whereas CYP2C9, CYP3A4, CYP2E1 and CYP1A2 gene expressions upregulated by ~3.5, ~~4, ~5 and ~15-fold, respectively in response to the alcohol injury. LX-2 cells also acquire more quiescent phenotype on C:F_3:1 scaffolds when compared to the gold standard sandwich culture upon alcohol treatment. Thus, C:F_3:1 scaffold with human liver cells was established as the potential platform to scan alcohol toxicity at varied alcohol concentrations. Thus, it can pave a promising path not only to support functional healthy human liver cells for liver tissue engineering but also to examine potential drugs to study the progression or inhibition of alcoholic liver fibrosis in vitro.</p>","PeriodicalId":80,"journal":{"name":"Integrative Biology","volume":"13 7","pages":"184-195"},"PeriodicalIF":2.5,"publicationDate":"2021-07-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"39118187","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A time-resolved experimental-mathematical model for predicting the response of glioma cells to single-dose radiation therapy.","authors":"Junyan Liu,&nbsp;David A Hormuth,&nbsp;Tessa Davis,&nbsp;Jianchen Yang,&nbsp;Matthew T McKenna,&nbsp;Angela M Jarrett,&nbsp;Heiko Enderling,&nbsp;Amy Brock,&nbsp;Thomas E Yankeelov","doi":"10.1093/intbio/zyab010","DOIUrl":"https://doi.org/10.1093/intbio/zyab010","url":null,"abstract":"<p><strong>Purpose: </strong>To develop and validate a mechanism-based, mathematical model that characterizes 9L and C6 glioma cells' temporal response to single-dose radiation therapy in vitro by explicitly incorporating time-dependent biological interactions with radiation.</p><p><strong>Methods: </strong>We employed time-resolved microscopy to track the confluence of 9L and C6 glioma cells receiving radiation doses of 0, 2, 4, 6, 8, 10, 12, 14 or 16 Gy. DNA repair kinetics are measured by γH2AX expression via flow cytometry. The microscopy data (814 replicates for 9L, 540 replicates for C6 at various seeding densities receiving doses above) were divided into training (75%) and validation (25%) sets. A mechanistic model was developed, and model parameters were calibrated to the training data. The model was then used to predict the temporal dynamics of the validation set given the known initial confluences and doses. The predictions were compared to the corresponding dynamic microscopy data.</p><p><strong>Results: </strong>For 9L, we obtained an average (± standard deviation, SD) Pearson correlation coefficient between the predicted and measured confluence of 0.87 ± 0.16, and an average (±SD) concordance correlation coefficient of 0.72 ± 0.28. For C6, we obtained an average (±SD) Pearson correlation coefficient of 0.90 ± 0.17, and an average (±SD) concordance correlation coefficient of 0.71 ± 0.24.</p><p><strong>Conclusion: </strong>The proposed model can effectively predict the temporal development of 9L and C6 glioma cells in response to a range of single-fraction radiation doses. By developing a mechanism-based, mathematical model that can be populated with time-resolved data, we provide an experimental-mathematical framework that allows for quantitative investigation of cells' temporal response to radiation. Our approach provides two key advances: (i) a time-resolved, dynamic death rate with a clear biological interpretation, and (ii) accurate predictions over a wide range of cell seeding densities and radiation doses.</p>","PeriodicalId":80,"journal":{"name":"Integrative Biology","volume":"13 7","pages":"167-183"},"PeriodicalIF":2.5,"publicationDate":"2021-07-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8271006/pdf/zyab010.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"39038280","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Cell surface integrin α5ß1 clustering negatively regulates receptor tyrosine kinase signaling in colorectal cancer cells via glycogen synthase kinase 3.","authors":"Alina Starchenko,&nbsp;Ramona Graves-Deal,&nbsp;Douglas Brubaker,&nbsp;Cunxi Li,&nbsp;Yuping Yang,&nbsp;Bhuminder Singh,&nbsp;Robert J Coffey,&nbsp;Douglas A Lauffenburger","doi":"10.1093/intbio/zyab009","DOIUrl":"https://doi.org/10.1093/intbio/zyab009","url":null,"abstract":"<p><p>As a key process within the tissue microenvironment, integrin signaling can influence cell functional responses to growth factor stimuli. We show here that clustering of integrin α5ß1 at the plasma membrane of colorectal cancer-derived epithelial cells modulates their ability to respond to stimulation by receptor tyrosine kinase (RTK)-activating growth factors EGF, NRG and HGF, through GSK3-mediated suppression of Akt pathway. We observed that integrin α5ß1 is lost from the membrane of poorly organized human colorectal tumors and that treatment with the integrin-clustering antibody P4G11 is sufficient to induce polarity in a mouse tumor xenograft model. While adding RTK growth factors (EGF, NRG and HGF) to polarized colorectal cancer cells induced invasion and loss of monolayer formation in 2D and 3D, this pathological behavior could be blocked by P4G11. Phosphorylation of ErbB family members as well as MET following EGF, NRG and HGF treatment was diminished in cells pretreated with P4G11. Focusing on EGFR, we found that blockade of integrin α5ß1 increased EGFR phosphorylation. Since activity of multiple downstream kinase pathways were altered by these various treatments, we employed computational machine learning techniques to ascertain the most important effects. Partial least-squares discriminant analysis identified GSK3 as a major regulator of EGFR pathway activities influenced by integrin α5ß1. Moreover, we used partial correlation analysis to examine signaling pathway crosstalk downstream of EGF stimulation and found that integrin α5ß1 acts as a negative regulator of the AKT signaling cascade downstream of EGFR, with GSK3 acting as a key mediator. We experimentally validated these computational inferences by confirming that blockade of GSK3 activity is sufficient to induce loss of polarity and increase of oncogenic signaling in the colonic epithelial cells.</p>","PeriodicalId":80,"journal":{"name":"Integrative Biology","volume":"13 6","pages":"153-166"},"PeriodicalIF":2.5,"publicationDate":"2021-06-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/e5/af/zyab009.PMC8204629.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"39019625","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Hyperglycemia minimally alters primary self-renewing human colonic epithelial cells while TNFα-promotes severe intestinal epithelial dysfunction.","authors":"Johanna S Dutton, Samuel S Hinman, Raehyun Kim, Peter J Attayek, Mallory Maurer, Christopher S Sims, Nancy L Allbritton","doi":"10.1093/intbio/zyab008","DOIUrl":"10.1093/intbio/zyab008","url":null,"abstract":"<p><p>Hyperglycemia is thought to increase production of inflammatory cytokines and permeability of the large intestine. Resulting intestinal inflammation is then often characterized by excess secretion of tumor necrosis factor alpha (TNFα). Thus, hyperglycemia in hospitalized patients suffering from severe trauma or disease is frequently accompanied by TNFα secretion, and the combined impact of these insults on the intestinal epithelium is poorly understood. This study utilized a simple yet elegant model of the intestinal epithelium, comprised of primary human intestinal stem cells and their differentiated progeny, to investigate the impact of hyperglycemia and inflammatory factors on the colonic epithelium. When compared to epithelium cultured under conditions of physiologic glucose, cells under hyperglycemic conditions displayed decreased mucin-2 (MUC2), as well as diminished alkaline phosphatase (ALP) activity. Conditions of 60 mM glucose potentiated secretion of the cytokine IL-8 suggesting that cytokine secretion during hyperglycemia may be a source of tissue inflammation. TNFα measurably increased secretion of IL-8 and IL-1β, which was enhanced at 60 mM glucose. Surprisingly, intestinal permeability and paracellular transport were not altered by even extreme levels of hyperglycemia. The presence of TNFα increased MUC2 presence, decreased ALP activity, and negatively impacted monolayer barrier function. When TNFα hyperglycemia and ≤30 mM glucose and were combined, MUC2 and ALP activity remained similar to that of TNFα alone, although synergistic effects were seen at 60 mM glucose. An automated image analysis pipeline was developed to assay changes in properties of the zonula occludens-1 (ZO-1)-demarcated cell boundaries. While hyperglycemia alone had little impact on cell shape and size, cell morphologic properties were extraordinarily sensitive to soluble TNFα. These results suggest that TNFα acted as the dominant modulator of the epithelium relative to glucose, and that control of inflammation rather than glucose may be key to maintaining intestinal homeostasis.</p>","PeriodicalId":80,"journal":{"name":"Integrative Biology","volume":"13 6","pages":"139-152"},"PeriodicalIF":2.5,"publicationDate":"2021-06-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8204630/pdf/zyab008.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"38982518","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Insights into therapeutic targets and biomarkers using integrated multi-'omics' approaches for dilated and ischemic cardiomyopathies.","authors":"Austė Kanapeckaitė,&nbsp;Neringa Burokienė","doi":"10.1093/intbio/zyab007","DOIUrl":"https://doi.org/10.1093/intbio/zyab007","url":null,"abstract":"<p><p>At present, heart failure (HF) treatment only targets the symptoms based on the left ventricle dysfunction severity; however, the lack of systemic 'omics' studies and available biological data to uncover the heterogeneous underlying mechanisms signifies the need to shift the analytical paradigm towards network-centric and data mining approaches. This study, for the first time, aimed to investigate how bulk and single cell RNA-sequencing as well as the proteomics analysis of the human heart tissue can be integrated to uncover HF-specific networks and potential therapeutic targets or biomarkers. We also aimed to address the issue of dealing with a limited number of samples and to show how appropriate statistical models, enrichment with other datasets as well as machine learning-guided analysis can aid in such cases. Furthermore, we elucidated specific gene expression profiles using transcriptomic and mined data from public databases. This was achieved using the two-step machine learning algorithm to predict the likelihood of the therapeutic target or biomarker tractability based on a novel scoring system, which has also been introduced in this study. The described methodology could be very useful for the target or biomarker selection and evaluation during the pre-clinical therapeutics development stage as well as disease progression monitoring. In addition, the present study sheds new light into the complex aetiology of HF, differentiating between subtle changes in dilated cardiomyopathies (DCs) and ischemic cardiomyopathies (ICs) on the single cell, proteome and whole transcriptome level, demonstrating that HF might be dependent on the involvement of not only the cardiomyocytes but also on other cell populations. Identified tissue remodelling and inflammatory processes can be beneficial when selecting targeted pharmacological management for DCs or ICs, respectively.</p>","PeriodicalId":80,"journal":{"name":"Integrative Biology","volume":"13 5","pages":"121-137"},"PeriodicalIF":2.5,"publicationDate":"2021-05-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1093/intbio/zyab007","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"38885786","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Mechanistic dissection of spatial organization in NF-κB signaling pathways by hybrid simulations.","authors":"Yinghao Wu,&nbsp;Kalyani Dhusia,&nbsp;Zhaoqian Su","doi":"10.1093/intbio/zyab006","DOIUrl":"https://doi.org/10.1093/intbio/zyab006","url":null,"abstract":"<p><p>The nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) is one of the most important transcription factors involved in the regulation of inflammatory signaling pathways. Inappropriate activation of these pathways has been linked to autoimmunity and cancers. Emerging experimental evidences have been showing the existence of elaborate spatial organizations for various molecular components in the pathways. One example is the scaffold protein tumor necrosis factor receptor associated factor (TRAF). While most TRAF proteins form trimeric quaternary structure through their coiled-coil regions, the N-terminal region of some members in the family can further be dimerized. This dimerization of TRAF trimers can drive them into higher-order clusters as a response to receptor stimulation, which functions as a spatial platform to mediate the downstream poly-ubiquitination. However, the molecular mechanism underlying the TRAF protein clustering and its functional impacts are not well-understood. In this article, we developed a hybrid simulation method to tackle this problem. The assembly of TRAF-based signaling platform at the membrane-proximal region is modeled with spatial resolution, while the dynamics of downstream signaling network, including the negative feedbacks through various signaling inhibitors, is simulated as stochastic chemical reactions. These two algorithms are further synchronized under a multiscale simulation framework. Using this computational model, we illustrated that the formation of TRAF signaling platform can trigger an oscillatory NF-κB response. We further demonstrated that the temporal patterns of downstream signal oscillations are closely regulated by the spatial factors of TRAF clustering, such as the geometry and energy of dimerization between TRAF trimers. In general, our study sheds light on the basic mechanism of NF-κB signaling pathway and highlights the functional importance of spatial regulation within the pathway. The simulation framework also showcases its potential of application to other signaling pathways in cells.</p>","PeriodicalId":80,"journal":{"name":"Integrative Biology","volume":"13 5","pages":"109-120"},"PeriodicalIF":2.5,"publicationDate":"2021-05-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8129552/pdf/zyab006.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"38903857","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A reconfigurable microscale assay enables insights into cancer-associated fibroblast modulation of immune cell recruitment.","authors":"Jiaquan Yu,&nbsp;Amber Piazza,&nbsp;Sidney Sparks,&nbsp;Laurel E Hind,&nbsp;David J Niles,&nbsp;Patrick N Ingram,&nbsp;Wei Huang,&nbsp;William A Ricke,&nbsp;David F Jarrard,&nbsp;Anna Huttenlocher,&nbsp;Hirak Basu,&nbsp;David J Beebe","doi":"10.1093/intbio/zyab004","DOIUrl":"https://doi.org/10.1093/intbio/zyab004","url":null,"abstract":"<p><p>Innate immune cell infiltration into neoplastic tissue is the first line of defense against cancer and can play a deterministic role in tumor progression. Here, we describe a series of assays, using a reconfigurable microscale assay platform (i.e. Stacks), which allows the study of immune cell infiltration in vitro with spatiotemporal manipulations. We assembled Stacks assays to investigate tumor-monocyte interactions, re-education of activated macrophages, and neutrophil infiltration. For the first time in vitro, the Stacks infiltration assays reveal that primary tumor-associated fibroblasts from specific patients differ from that associated with the benign region of the prostate in their ability to limit neutrophil infiltration as well as facilitate monocyte adhesion and anti-inflammatory monocyte polarization. These results show that fibroblasts play a regulatory role in immune cell infiltration and that Stacks has the potential to predict individual patients' cancer-immune response.</p>","PeriodicalId":80,"journal":{"name":"Integrative Biology","volume":"13 4","pages":"87-97"},"PeriodicalIF":2.5,"publicationDate":"2021-04-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8793920/pdf/nihms-1762984.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"25564238","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Microbial-based magnetic nanoparticles production: a mini-review.","authors":"Victor Chmykhalo,&nbsp;Anna Belanova,&nbsp;Mariya Belousova,&nbsp;Vera Butova,&nbsp;Yuriy Makarenko,&nbsp;Vera Khrenkova,&nbsp;Alexander Soldatov,&nbsp;Peter Zolotukhin","doi":"10.1093/intbio/zyab005","DOIUrl":"https://doi.org/10.1093/intbio/zyab005","url":null,"abstract":"<p><p>The ever-increasing biomedical application of magnetic nanoparticles (MNPs) implies increasing demand in their scalable and high-throughput production, with finely tuned and well-controlled characteristics. One of the options to meet the demand is microbial production by nanoparticles-synthesizing bacteria. This approach has several benefits over the standard chemical synthesis methods, including improved homogeneity of synthesis, cost-effectiveness, safety and eco-friendliness. There are, however, specific challenges emanating from the nature of the approach that are to be accounted and resolved in each manufacturing instance. Most of the challenges can be resolved by proper selection of the producing organism and optimizing cell culture and nanoparticles extraction conditions. Other issues require development of proper continuous production equipment, medium usage optimization and precursor ions recycling. This mini-review focuses on the related topics in microbial synthesis of MNPs: producing organisms, culturing methods, nanoparticles characteristics tuning, nanoparticles yield and synthesis timeframe considerations, nanoparticles isolation as well as on the respective challenges and possible solutions.</p>","PeriodicalId":80,"journal":{"name":"Integrative Biology","volume":"13 4","pages":"98-107"},"PeriodicalIF":2.5,"publicationDate":"2021-04-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1093/intbio/zyab005","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"25572157","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Oxygenation as a driving factor in epithelial differentiation at the air-liquid interface.","authors":"Sonya Kouthouridis, Julie Goepp, Carolina Martini, Elizabeth Matthes, John W Hanrahan, Christopher Moraes","doi":"10.1093/intbio/zyab002","DOIUrl":"10.1093/intbio/zyab002","url":null,"abstract":"<p><p>Culture at the air-liquid interface is broadly accepted as necessary for differentiation of cultured epithelial cells towards an in vivo-like phenotype. However, air-liquid interface cultures are expensive, laborious and challenging to scale for increased throughput applications. Deconstructing the microenvironmental parameters that drive these differentiation processes could circumvent these limitations, and here we hypothesize that reduced oxygenation due to diffusion limitations in liquid media limits differentiation in submerged cultures; and that this phenotype can be rescued by recreating normoxic conditions at the epithelial monolayer, even under submerged conditions. Guided by computational models, hyperoxygenation of atmospheric conditions was applied to manipulate oxygenation at the monolayer surface. The impact of this rescue condition was confirmed by assessing protein expression of hypoxia-sensitive markers. Differentiation of primary human bronchial epithelial cells isolated from healthy patients was then assessed in air-liquid interface, submerged and hyperoxygenated submerged culture conditions. Markers of differentiation, including epithelial layer thickness, tight junction formation, ciliated surface area and functional capacity for mucociliary clearance, were assessed and found to improve significantly in hyperoxygenated submerged cultures, beyond standard air-liquid interface or submerged culture conditions. These results demonstrate that an air-liquid interface is not necessary to produce highly differentiated epithelial structures, and that increased availability of oxygen and nutrient media can be leveraged as important strategies to improve epithelial differentiation for applications in respiratory toxicology and therapeutic development.</p>","PeriodicalId":80,"journal":{"name":"Integrative Biology","volume":"13 3","pages":"61-72"},"PeriodicalIF":2.5,"publicationDate":"2021-03-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7965686/pdf/zyab002.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"25442115","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}