Determination of patulin in apple juice by HPLC using a simple and fast sample preparation method

The goal of this work was to develop a simple and rapid preparation method for patulin analysis in apple juice without previous clean-up. This method combined sonication and liquid extraction techniques and was used for determination of patulin in 37 commercial apple juices available on the market in the South of Brazil. The method performance characteristics were determined using a sample obtained in a local market fortified at five concentration levels of patulin and done in triplicates. The coefficient of variation for repeatability at the fortification level of 20.70µg.L-1 was 3.53 % and the recovery 94.63 %, respectively. The correlation coefficient was 0.9996 and agrees with the requirements for a linear analytical method value. The detection limit was 0.21µg.L-1 and the quantification limit 0.70 µg.L-1. Only three of the analyzed samples were upper the allowed level of 50.00 µg.L-1 recommended for the World Health Organization.O objetivo deste trabalho foi desenvolver um método simples e rápido para preparação de amostras para análise de patulina em sucos de maçã sem "clean-up" prévio. Este método combina os métodos de extração líquida com o ultrassom e foi utilizado para a determinação de patulina em 37 amostras de suco de maçã disponíveis no mercado do sul do Brasil. Os parâmetros estatísticos para validação do método foram obtidos através de fortificação de suco de maçã, obtido no mercado local, em cinco níveis de concentração de patulina e realizadas em triplicatas. O coeficiente de variação para a repetibilidade no nível de fortificação de 20,70 µg.L-1 foi de 3,53 % e a recuperação de 94,63 %. O coeficiente de correlação foi de 0,9996 e este valor está de acordo com os requisitos de linearidade de um método analítico. O limite de detecção obtido foi de 0,21 µg.L-1 e o limite de quantificação de 0,7 µg.L-1. Somente três das amostras analisadas tiveram valores superiores ao limite de 50,00 µg.L-1 recomendado pela Organização Mundial da Saúde.FATE

Microfluidic tools for enhanced characterization of therapeutic stem cells and prediction of their potential antimicrobial secretome

Antibiotic resistance is creating enormous attention on the development of new antibiotic-free therapy strategies for bacterial diseases. Mesenchymal stromal stem cells (MSCs) are the most promising candidates in current clinical trials and included in several cell-therapy protocols. Together with the well-known immunomodulatory and regenerative potential of the MSC secretome, these cells have shown direct and indirect anti-bacterial effects. However, the low reproducibility and standardization of MSCs from different sources are the current limitations prior to the purification of cell-free secreted antimicrobial peptides and exosomes. In order to improve MSC characterization, novel label-free functional tests, evaluating the biophysical properties of the cells, will be advan-tageous for their cell profiling, population sorting, and quality control. We discuss the potential of emerging microfluidic technologies providing new insights into density, shape, and size of live cells, starting from heterogeneous or 3D cultured samples. The prospective application of these technologies to studying MSC populations may contribute to developing new biopharmaceutical strategies with a view to naturally overcoming bacterial defense mechanisms

Physical characterization of colorectal cancer spheroids and evaluation of NK cell Infiltration through a flow-based analysis

To improve pathogenetic studies in cancer development and reliable preclinical testing of anti-cancer treatments, three-dimensional (3D) cultures, including spheroids, have been widely recognized as more physiologically relevant in vitro models of in vivo tumor behavior. Currently, the generation of uniformly sized spheroids is still challenging: different 3D cell culture methods produce heterogeneous populations in dimensions and morphology, that may strongly influence readouts reliability correlated to tumor growth rate or antitumor natural killer (NK) cell-mediated cytotoxicity. In this context, an increasing consensus claims the integration of microfluidic technologies within 3D cell culture, as the physical characterization of tumor spheroids is unavoidably demanded to standardize protocols and assays for in vitro testing. In this paper, we employed a flow-based method specifically conceived to measure weight, size and focused onto mass density values of tumor spheroids. These measurements are combined with confocal and digital imaging of such samples. We tested the spheroids of four colorectal cancer (CRC) cell lines that exhibit statistically relevant differences in their physical characteristics, even though starting from the same cell seeding density. These variations are seemingly cell line-dependent and associated with the number of growing cells and the degree of spheroid compaction as well, supported by different adenosine-triphosphate contents. We also showed that this technology can estimate the NK cell killing efficacy by measuring the weight loss and diameter shrinkage of tumor spheroids, alongside with the commonly used cell viability in vitro test. As the activity of NK cells relies on their infiltration rate, the in vitro sensitivity of CRC spheroids proved to be exposure time- and cell line-dependent with direct correlation to the cell viability reduction. All these functional aspects can be measured by the system and are documented by digital image analysis. In conclusion, this flow-based method potentially paves the way towards standardization of 3D cell cultures and its early adoption in cancer research to test antitumor immune response and set up new immunotherapy strategies

Magnesium Deprivation Potentiates Human Mesenchymal Stem Cell Transcriptional Remodeling

Magnesium plays a pivotal role in energy metabolism and in the control of cell growth. While magnesium deprivation clearly shapes the behavior of normal and neoplastic cells, little is known on the role of this element in cell differentiation. Here we show that magnesium deficiency increases the transcription of multipotency markers and tissue-specific transcription factors in human adipose-derived mesenchymal stem cells exposed to a mixture of natural molecules, i.e., hyaluronic, butyric and retinoid acids, which tunes differentiation. We also demonstrate that magnesium deficiency accelerates the osteogenic differentiation of human bone marrow-derived mesenchymal stem cells. We argue that magnesium deprivation generates a stressful condition that modulates stem cell plasticity and differentiation potential. These studies indicate that it is possible to remodel transcription in mesenchymal stem cells by lowering extracellular magnesium without the need for genetic manipulation, thus offering new hints for regenerative medicine applications

Autoantibodies against chemokines post-SARS-CoV-2 infection correlate with disease course

Infection with severe acute respiratory syndrome coronavirus 2 associates with diverse symptoms, which can persist for months. While antiviral antibodies are protective, those targeting interferons and other immune factors are associated with adverse coronavirus disease 2019 (COVID-19) outcomes. Here we discovered that antibodies against specific chemokines were omnipresent post-COVID-19, were associated with favorable disease outcome and negatively correlated with the development of long COVID at 1 yr post-infection. Chemokine antibodies were also present in HIV-1 infection and autoimmune disorders, but they targeted different chemokines compared with COVID-19. Monoclonal antibodies derived from COVID-19 convalescents that bound to the chemokine N-loop impaired cell migration. Given the role of chemokines in orchestrating immune cell trafficking, naturally arising chemokine antibodies may modulate the inflammatory response and thus bear therapeutic potential

Anti-chemokine antibodies after SARS-CoV-2 infection correlate with favorable disease course.

Infection by SARS-CoV-2 leads to diverse symptoms, which can persist for months. While antiviral antibodies are protective, those targeting interferons and other immune factors are associated with adverse COVID-19 outcomes. Instead, we discovered that antibodies against specific chemokines are omnipresent after COVID-19, associated with favorable disease, and predictive of lack of long COVID symptoms at one year post infection. Anti-chemokine antibodies are present also in HIV-1 and autoimmune disorders, but they target different chemokines than those in COVID-19. Finally, monoclonal antibodies derived from COVID- 19 convalescents that bind to the chemokine N-loop impair cell migration. Given the role of chemokines in orchestrating immune cell trafficking, naturally arising anti-chemokine antibodies associated with favorable COVID-19 may be beneficial by modulating the inflammatory response and thus bear therapeutic potential. One-Sentence Summary Naturally arising anti-chemokine antibodies associate with favorable COVID-19 and are predictive of lack of long COVID

Autoantibodies against chemokines post-SARS-CoV-2 infection correlate with disease course

Infection with severe acute respiratory syndrome coronavirus 2 associates with diverse symptoms, which can persist for months. While antiviral antibodies are protective, those targeting interferons and other immune factors are associated with adverse coronavirus disease 2019 (COVID-19) outcomes. Here we discovered that antibodies against specific chemokines were omnipresent post-COVID-19, were associated with favorable disease outcome and negatively correlated with the development of long COVID at 1 yr post-infection. Chemokine antibodies were also present in HIV-1 infection and autoimmune disorders, but they targeted different chemokines compared with COVID-19. Monoclonal antibodies derived from COVID-19 convalescents that bound to the chemokine N-loop impaired cell migration. Given the role of chemokines in orchestrating immune cell trafficking, naturally arising chemokine antibodies may modulate the inflammatory response and thus bear therapeutic potential

Colorectal Cancer Stage at Diagnosis Before vs During the COVID-19 Pandemic in Italy

IMPORTANCE Delays in screening programs and the reluctance of patients to seek medical attention because of the outbreak of SARS-CoV-2 could be associated with the risk of more advanced colorectal cancers at diagnosis. OBJECTIVE To evaluate whether the SARS-CoV-2 pandemic was associated with more advanced oncologic stage and change in clinical presentation for patients with colorectal cancer. DESIGN, SETTING, AND PARTICIPANTS This retrospective, multicenter cohort study included all 17 938 adult patients who underwent surgery for colorectal cancer from March 1, 2020, to December 31, 2021 (pandemic period), and from January 1, 2018, to February 29, 2020 (prepandemic period), in 81 participating centers in Italy, including tertiary centers and community hospitals. Follow-up was 30 days from surgery. EXPOSURES Any type of surgical procedure for colorectal cancer, including explorative surgery, palliative procedures, and atypical or segmental resections. MAIN OUTCOMES AND MEASURES The primary outcome was advanced stage of colorectal cancer at diagnosis. Secondary outcomes were distant metastasis, T4 stage, aggressive biology (defined as cancer with at least 1 of the following characteristics: signet ring cells, mucinous tumor, budding, lymphovascular invasion, perineural invasion, and lymphangitis), stenotic lesion, emergency surgery, and palliative surgery. The independent association between the pandemic period and the outcomes was assessed using multivariate random-effects logistic regression, with hospital as the cluster variable. RESULTS A total of 17 938 patients (10 007 men [55.8%]; mean [SD] age, 70.6 [12.2] years) underwent surgery for colorectal cancer: 7796 (43.5%) during the pandemic period and 10 142 (56.5%) during the prepandemic period. Logistic regression indicated that the pandemic period was significantly associated with an increased rate of advanced-stage colorectal cancer (odds ratio [OR], 1.07; 95%CI, 1.01-1.13; P = .03), aggressive biology (OR, 1.32; 95%CI, 1.15-1.53; P < .001), and stenotic lesions (OR, 1.15; 95%CI, 1.01-1.31; P = .03). CONCLUSIONS AND RELEVANCE This cohort study suggests a significant association between the SARS-CoV-2 pandemic and the risk of a more advanced oncologic stage at diagnosis among patients undergoing surgery for colorectal cancer and might indicate a potential reduction of survival for these patients