Cell-biomaterial mechanical interactions: from cancer mechanobiology to cell therapies

The physical characteristics of the cell microenvironment greatly affect cellular processes such as survival, proliferation, migration, and differentiation. Biomaterials with well-defined physical and chemical properties have been used to better understand the cell microenvironment. Furthermore, the physicochemical properties of biomaterials can be modulated to induce host body responses and therapeutic cell behaviour. Based on these premises, the work presented in this thesis investigated the effect of soluble polymers commonly used in cell therapies on the physical properties of the extracellular microenvironment. It was found, that viscosity-enhancing polymers induce mesenchymal migration in liver cancer cells, an effect derived from changes in integrin-dependent cell – substrate adhesion dynamics and mechanosensing. Also, the role of extracellular polymers on endothelial-derived cell alignment was explored indicating that molecular soluble polymers enhance cell elongation and alignment in a molecular weight-dependent manner. In addition, the effect of hydrogel density and crosslinking causing mechanical confinement was found to affect cancer cell growth and cell cycle progression, leading to an enhanced content in polyploid cells. Finally, the effect of mechanical confinement on liver cancer cells was taken advantage of to improve the production of biomass for a bioartificial liver device by modulating the degree of crosslinking of alginate hydrogel. In conclusion, the work presented here indicates that physical properties of both the extracellular fluid and matrix greatly affect cell behaviour and can be exploited to improve biomaterial design for in vitro testing and clinical applications

MMP14 in Sarcoma: A Regulator of Tumor Microenvironment Communication in Connective Tissues

Sarcomas are deadly malignant tumors of mesenchymal origin occurring at all ages. The expression and function of the membrane-type matrix metalloproteinase MMP14 is closely related to the mesenchymal cell phenotype, and it is highly expressed in most sarcomas. MMP14 regulates the activity of multiple extracellular and plasma membrane proteins, influencing cell–cell and cell–extracellular matrix (ECM) communication. This regulation mediates processes such as ECM degradation and remodeling, cell invasion, and cancer metastasis. Thus, a comprehensive understanding of the biology of MMP14 in sarcomas will shed light on the mechanisms controlling the key processes in these diseases. Here, we provide an overview of the function and regulation of MMP14 and we discuss their relationship with clinical and pre-clinical MMP14 data in both adult and childhood sarcomas

Analytical Evaluation of Dried Blood Spot and Rapid Diagnostic Test as a New Strategy for Serological Community Screening for Chronic Chagas Disease

Trypanosoma cruzi; Mancha de sangre seca (DBS); Cribado serológicoTrypanosoma cruzi; Taca de sang seca (DBS); Cribratge serològicTrypanosoma cruzi; Dried blood spot (DBS); Serological screeningBackground: Chagas disease is a public health problem not only in Latin America, but also in other regions, including Spain, due to migration movements. Conventional serological diagnosis requires an invasive sample (plasma or serum) and a well-equipped laboratory. To circumvent those limitations, blood samples dried on filter paper (DBS) or Rapid Diagnostic Test (RDT) could be a practical alternative to reference protocol for serological screening in epidemiological studies. We evaluated the usefulness of dried blood sampling and a rapid diagnostic test (Trypanosoma Detect™) for the detection of antibodies against T. cruzi for their use in community-based screening. Methodology/Principal Findings: A total of 162 stored paired whole-blood and serum samples from Latin American migrants and 25 negative-control blood samples were included. Diagnosis of chronic Chagas disease was performed in serum according to WHO algorithms. Blood samples were retrospectively collected as dried spots and then analyzed using two different serological techniques, enzyme-linked immunosorbent assay (ELISA) and electrochemiluminescence immunoassay (E-CLIA). Whole-blood samples were also used to evaluate a rapid diagnostic test based on immunochromatography. A better correlation with conventional serum was observed in dried blood elutes using E-CLIA than ELISA (97% vs. 77% sensitivity, respectively). Both assays reported 100% specificity. The median cut-off index values of E-CLIA for dried blood were significantly lower than those for serum (138.1 vs. 243.3, P<0.05). The Trypanosoma Detect™ test presented a sensitivity and specificity of 89.6% and 100%, respectively. Conclusions: The detection of antibodies against T. cruzi in dried blood samples shows a higher sensitivity when using E-CLIA compared with ELISA. Trypanosoma Detect™ is easier to use but has a lower sensitivity. Hence, we propose a sequential strategy based on performing the rapid test first, and a negative result will be confirmed by DBS-ECLIA for use in community Chagas disease screening programs.This work has been supported by the Fundació la Marató TV3 (project number 20182610)

Detection of EGFR mutations with mutation-specific antibodies in stage IV non-small-cell lung cancer

<p>Abstract</p> <p>Background</p> <p>Immunohistochemistry (IHC) with mutation-specific antibodies may be an ancillary method of detecting EGFR mutations in lung cancer patients.</p> <p>Methods</p> <p>EGFR mutation status was analyzed by DNA assays, and compared with IHC results in five non-small-cell lung cancer (NSCLC) cell lines and tumor samples from 78 stage IV NSCLC patients.</p> <p>Results</p> <p>IHC correctly identified del 19 in the H1650 and PC9 cell lines, L858R in H1975, and wild-type EGFR in H460 and A549, as well as wild-type EGFR in tumor samples from 22 patients. IHC with the mAb against EGFR with del 19 was highly positive for the protein in all 17 patients with a 15-bp (ELREA) deletion in exon 19, whereas in patients with other deletions, IHC was weakly positive in 3 cases and negative in 9 cases. IHC with the mAb against the L858R mutation showed high positivity for the protein in 25/27 (93%) patients with exon 21 EGFR mutations (all with L858R) but did not identify the L861Q mutation in the remaining two patients.</p> <p>Conclusions</p> <p>IHC with mutation-specific mAbs against EGFR is a promising method for detecting EGFR mutations in NSCLC patients. However these mAbs should be validated with additional studies to clarify their possible role in routine clinical practice for screening EGFR mutations in NSCLC patients.</p

Spontaneous formation of spiral-like patterns with distinct periodic physical properties by confined electrodeposition of Co-In disks

Golvano-Escobal, Irati et al.Spatio-temporal patterns are ubiquitous in different areas of materials science and biological systems. However, typically the motifs in these types of systems present a random distribution with many possible different structures. Herein, we demonstrate that controlled spatio-temporal patterns, with reproducible spiral-like shapes, can be obtained by electrodeposition of Co-In alloys inside a confined circular geometry (i.e., in disks that are commensurate with the typical size of the spatio-temporal features). These patterns are mainly of compositional nature, i.e., with virtually no topographic features. Interestingly, the local changes in composition lead to a periodic modulation of the physical (electric, magnetic and mechanical) properties. Namely, the Co-rich areas show higher saturation magnetization and electrical conductivity and are mechanically harder than the In-rich ones. Thus, this work reveals that confined electrodeposition of this binary system constitutes an effective procedure to attain template-free magnetic, electric and mechanical surface patterning with specific and reproducible shapes.This work has been partially funded by the 2014-SGR-1015, 2014-SGR-1216 and 2014-SGR-753 projects from the Generalitat de Catalunya, the MAT2014-57960-C3-1-R, MAT2014-51778-C2-1-R and FIS2013-48668-C2-1-P from the Spanish Ministerio de Economía y Competitividad (MINECO) and the ‘Fondo Europeo de Desarrollo Regional’ (FEDER). Dr. Eva Pellicer and Dr. Neus Domingo are also grateful to MINECO for the “Ramon y Cajal” contracts (RYC-2012-10839 and RYC-2010-06365 respectively). ICN2 and ICMAB acknowledge support from the Severo Ochoa Program (MINECO, Grant SEV-2013-0295 and Grant SEV-2015-0496, respectively). The PEEM experiments were performed at CIRCE beamline at ALBA Synchrotron with the collaboration of ALBA staff.Peer reviewe

Differential effects of the second SARS-CoV-2 mRNA vaccine dose on T cell immunity in naive and COVID-19 recovered individuals

The rapid development of mRNA-based vaccines against the severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) led to the design of accelerated vaccination schedules that have been extremely effective in naive individuals. While a two-dose immunization regimen with the BNT162b2 vaccine has been demonstrated to provide a 95% efficacy in naive individuals, the effects of the second vaccine dose in individuals who have previously recovered from natural SARS-CoV-2 infection has not been investigated in detail. In this study, we characterize SARS-CoV-2 spike-specific humoral and cellular immunity in naive and previously infected individuals during and after two doses of BNT162b2 vaccination. Our results demonstrate that, while the second dose increases both the humoral and cellular immunity in naive individuals, COVID-19 recovered individuals reach their peak of immunity after the first dose. These results suggests that a second dose, according to the current standard regimen of vaccination, may be not necessary in individuals previously infected with SARS-CoV-2.Research reported in this publication was supported in part by the National Cancer Institute of the NIH (5R01HD102614-02; R01CA249204 and R01CA248984) and an ISMMS seed fund to E.G. The authors gratefully acknowledge use of the services and facilities of the Tisch Cancer Institute supported by a NCI Cancer Center Support Grant (P30 CA196521). M.S. was supported by a NCI training grant (T32CA078207). This work was supported by an ISMMS seed fund to J.O.; Instituto de Salud Carlos III (COV20-00668) to R.C.R.; the Instituto de Salud Carlos III, Spanish Ministry of Science and Innovation (COVID-19 research call COV20/00181) co-financed by the European Development Regional Fund “A way to achieve Europe” to E.P.; the Instituto de Salud Carlos III, Spain (COV20/00170); the Government of Cantabria, Spain (2020UIC22-PUB-0019) to M.L.H.; the Instituto de Salud Carlos III (PI16CIII/00012) to P.P.; the Fondo Social Europeo e Iniciativa de Empleo Juvenil YEI (Grant PEJ2018-004557-A) to M.P.E.; and by REDInREN 016/009/009 ISCIII. This project has received funding from the European Union Horizon 2020 research and innovation programs VACCELERATE and INsTRuCT under grant agreements 101037867 and 860003.S

Th1 and Th17 hypercytokinemia as early host response signature in severe pandemic influenza

Abstract Introduction Human host immune response following infection with the new variant of A/H1N1 pandemic influenza virus (nvH1N1) is poorly understood. We utilize here systemic cytokine and antibody levels in evaluating differences in early immune response in both mild and severe patients infected with nvH1N1. Methods We profiled 29 cytokines and chemokines and evaluated the haemagglutination inhibition activity as quantitative and qualitative measurements of host immune responses in serum obtained during the first five days after symptoms onset, in two cohorts of nvH1N1 infected patients. Severe patients required hospitalization (n = 20), due to respiratory insufficiency (10 of them were admitted to the intensive care unit), while mild patients had exclusively flu-like symptoms (n = 15). A group of healthy donors was included as control (n = 15). Differences in levels of mediators between groups were assessed by using the non parametric U-Mann Whitney test. Association between variables was determined by calculating the Spearman correlation coefficient. Viral load was performed in serum by using real-time PCR targeting the neuraminidase gene. Results Increased levels of innate-immunity mediators (IP-10, MCP-1, MIP-1β), and the absence of anti-nvH1N1 antibodies, characterized the early response to nvH1N1 infection in both hospitalized and mild patients. High systemic levels of type-II interferon (IFN-γ) and also of a group of mediators involved in the development of T-helper 17 (IL-8, IL-9, IL-17, IL-6) and T-helper 1 (TNF-α, IL-15, IL-12p70) responses were exclusively found in hospitalized patients. IL-15, IL-12p70, IL-6 constituted a hallmark of critical illness in our study. A significant inverse association was found between IL-6, IL-8 and PaO2 in critical patients. Conclusions While infection with the nvH1N1 induces a typical innate response in both mild and severe patients, severe disease with respiratory involvement is characterized by early secretion of Th17 and Th1 cytokines usually associated with cell mediated immunity but also commonly linked to the pathogenesis of autoimmune/inflammatory diseases. The exact role of Th1 and Th17 mediators in the evolution of nvH1N1 mild and severe disease merits further investigation as to the detrimental or beneficial role these cytokines play in severe illness

SMG-1 and mTORC1 Act Antagonistically to Regulate Response to Injury and Growth in Planarians

Planarian flatworms are able to both regenerate their whole bodies and continuously adapt their size to nutrient status. Tight control of stem cell proliferation and differentiation during these processes is the key feature of planarian biology. Here we show that the planarian homolog of the phosphoinositide 3-kinase-related kinase (PIKK) family member SMG-1 and mTOR complex 1 components are required for this tight control. Loss of smg-1 results in a hyper-responsiveness to injury and growth and the formation of regenerative blastemas that remain undifferentiated and that lead to lethal ectopic outgrowths. Invasive stem cell hyper-proliferation, hyperplasia, hypertrophy, and differentiation defects are hallmarks of this uncontrolled growth. These data imply a previously unappreciated and novel physiological function for this PIKK family member. In contrast we found that planarian members of the mTOR complex 1, tor and raptor, are required for the initial response to injury and blastema formation. Double smg-1 RNAi experiments with tor or raptor show that abnormal growth requires mTOR signalling. We also found that the macrolide rapamycin, a natural compound inhibitor of mTORC1, is able to increase the survival rate of smg-1 RNAi animals by decreasing cell proliferation. Our findings support a model where Smg-1 acts as a novel regulator of both the response to injury and growth control mechanisms. Our data suggest the possibility that this may be by suppressing mTOR signalling. Characterisation of both the planarian mTORC1 signalling components and another PIKK family member as key regulators of regeneration and growth will influence future work on regeneration, growth control, and the development of anti-cancer therapies that target mTOR signalling

Total and Subtypes of Dietary Fat Intake and Its Association with Components of the Metabolic Syndrome in a Mediterranean Population at High Cardiovascular Risk

Background: The effect of dietary fat intake on the metabolic syndrome (MetS) and in turn on cardiovascular disease (CVD) remains unclear in individuals at high CVD risk. Objective: To assess the association between fat intake and MetS components in an adult Mediterranean population at high CVD risk. Design: Baseline assessment of nutritional adequacy in participants (n = 6560, men and women, 55-75 years old, with overweight/obesity and MetS) in the PREvención con DIeta MEDiterránea (PREDIMED)-Plus randomized trial. Methods: Assessment of fat intake (total fat, monounsatured fatty acids: MUFA, polyunsaturated fatty acids: PUFA, saturated fatty acids: SFA, trans-fatty acids: trans-FA, linoleic acid, α-linolenic acid, and ω-3 FA) using a validated food frequency questionnaire, and diet quality using 17-item Mediterranean dietary questionnaire and fat quality index (FQI). Results: Participants in the highest quintile of total dietary fat intake showed lower intake of energy, carbohydrates, protein and fiber, but higher intake of PUFA, MUFA, SFA, TFA, LA, ALA and ω-3 FA. Differences in MetS components were found according to fat intake. Odds (5th vs. 1st quintile): hyperglycemia: 1.3-1.6 times higher for total fat, MUFA, SFA and ω-3 FA intake; low high-density lipoprotein cholesterol (HDL-c): 1.2 higher for LA; hypertriglyceridemia: 0.7 lower for SFA and ω-3 FA intake. Conclusions: Dietary fats played different role on MetS components of high CVD risk patients. Dietary fat intake was associated with higher risk of hyperglycemia

Efficacy and Safety of Ixekizumab in the Treatment of Radiographic Axial Spondyloarthritis:Sixteen-Week Results From a Phase III Randomized, Double-Blind, Placebo-Controlled Trial in Patients With Prior Inadequate Response to or Intolerance of Tumor Necrosis Factor Inhibitors

Objective: To investigate the efficacy and safety of ixekizumab in patients with active radiographic axial spondyloarthritis (SpA) and prior inadequate response to or intolerance of 1 or 2 tumor necrosis factor inhibitors (TNFi). Methods: In this phase III randomized, double-blind, placebo-controlled trial, adult patients with an inadequate response to or intolerance of 1 or 2 TNFi and an established diagnosis of axial SpA (according to the Assessment of SpondyloArthritis international Society [ASAS] criteria for radiographic axial SpA, with radiographic sacroiliitis defined according to the modified New York criteria and ≥1 feature of SpA) were recruited and randomized 1:1:1 to receive placebo or 80-mg subcutaneous ixekizumab every 2 weeks (IXEQ2W) or 4 weeks (IXEQ4W), with an 80-mg or 160-mg starting dose. The primary end point was 40% improvement in disease activity according to the ASAS criteria (ASAS40) at week 16. Secondary outcomes and safety were also assessed. Results: A total of 316 patients were randomized to receive placebo (n = 104), IXEQ2W (n = 98), or IXEQ4W (n = 114). At week 16, significantly higher proportions of IXEQ2W patients (n = 30 [30.6%]; P = 0.003) or IXEQ4W patients (n = 29 [25.4%]; P = 0.017) had achieved an ASAS40 response versus the placebo group (n = 13 [12.5%]), with statistically significant differences reported as early as week 1 with ixekizumab treatment. Statistically significant improvements in disease activity, function, quality of life, and spinal magnetic resonance imaging–evident inflammation were observed after 16 weeks of ixekizumab treatment versus placebo. Treatment-emergent adverse events (AEs) with ixekizumab treatment were more frequent than with placebo. Serious AEs were similar across treatment arms. One death was reported (IXEQ2W group). Conclusion: Ixekizumab treatment for 16 weeks in patients with active radiographic axial SpA and previous inadequate response to or intolerance of 1 or 2 TNFi yields rapid and significant improvements in the signs and symptoms of radiographic axial SpA versus placebo