Cholesteryl hemiazelate identified in CVD patients causes in vitro and in vivo inflammation.

Oxidation of PUFAs in LDLs trapped in the arterial intima plays a critical role in atherosclerosis. Though there have been many studies on the atherogenicity of oxidized derivatives of PUFA-esters of cholesterol, the effects of cholesteryl hemiesters (ChEs), the oxidation end products of these esters, have not been studied. Through lipidomics analyses, we identified and quantified two ChE types in the plasma of CVD patients and identified four ChE types in human endarterectomy specimens. Cholesteryl hemiazelate (ChA), the ChE of azelaic acid (n-nonane-1,9-dioic acid), was the most prevalent ChE identified in both cases. Importantly, human monocytes, monocyte-derived macrophages, and neutrophils exhibit inflammatory features when exposed to subtoxic concentrations of ChA in vitro. ChA increases the secretion of proinflammatory cytokines such as interleukin-1ß and interleukin-6 and modulates the surface-marker profile of monocytes and monocyte-derived macrophage. In vivo, when zebrafish larvae were fed with a ChA-enriched diet, they exhibited neutrophil and macrophage accumulation in the vasculature in a caspase 1- and cathepsin B-dependent manner. ChA also triggered lipid accumulation at the bifurcation sites of the vasculature of the zebrafish larvae and negatively impacted their life expectancy. We conclude that ChA behaves as an endogenous damage-associated molecular pattern with inflammatory and proatherogenic properties.

Phytophagous insects and natural enemies on Sapindus saponaria L. (Sapindales: Sapindaceae) plants fertilized with or without dehydrated sewage sludge.

Management programs and efficient techniques are necessary to recover degraded ecosystems. The sewage sludge is rich in nitrogen (N) and with the potential to fertilize Sapindus saponaria L. (Sapinales: Sapindaceae), used in the recovery of degraded areas; this can affect the insect fauna. The study's objective was to evaluate, for 24 months, the abundance of chewing insects, dipterans, pollinators, and predators on S. saponaria plants fertilized with or without dehydrated sewage sludge in a degraded area. The experimental design was completely randomized (with the same characteristics) with two treatments (with or without dehydrated sewage sludge) and 24 replicates, each with one plant. The abundance of Anastrepha sp. (Tephritidae), Cerotoma sp. (Chrysomelidae), Curculionidae, Musca domestica L. (Muscidae), Mantis religiosa L. (Mantodea: Mantidae), Oxyopidae, Salticidae, Tettigoniidae (Orthoptera), and Teudis sp. (Anyphaenidae) was higher on fertilized plants. The abundances of Teudis sp. and Tmarus sp. (Thomisidae) and M. religiosa and Teudis sp. were positively correlated with chewing insects and Diptera, respectively. The population increase of insects and spiders on S. saponaria plants fertilized with dehydrated sewage sludge (bigger crowns) has shown to be suitable for recovering degraded areas with a higher number of niches and better food quality, improving the ecological indices of the area.

Reconciling diagnostic traits in living and fossil taxa: The taxonomy and evolution of the genus Microceratina (Crustacea, Ostracoda, Cytheruridae).

The original diagnosis of the genus Microceratina Swanson is re-examined with the aim of elucidating the evolutionary history of this living and fossil genus. The different systematic placements of this genus in one of three alternative suprageneric taxa, family Bythocytheridae Sars, family Cytheruridae G.W. Müller (subfamily Eucytherurinae Puri, emend. Maddocks & Steineck) and/or family Loxoconchidae Sars are reviewed. We provide details of a special type of tiny pores, named Loophole Sieve-type Pore Canals, and show how their morphology and position on the valve differ from the typical Sieve-type Pore Canals present in other superfamily Cytheroidea groups, especially members of the Loxoconchidae. A comparative analysis of the valve ornamentation, especially the structure of the anterior peripheral area, and posterior margin morphology, between selected taxa of the three subfamilies of the Cytheruridae (Cytherurinae, Cytheropterinae, Eucytherurinae) demonstrates that Microceratina species belong to a special phylogenetic lineage of Eucytherurinae, which differs from another lineage represented by the genus Xylocythere Maddocks & Steineck. This proposal is strengthened by examination of the limb traits of Microceratina martensi Namiotko et al., currently the only living species of the genus for which both valves and limbs are described; the description is extended herein. Key diagnostic traits of the genus Microceratina are presented, a new Early Jurassic age species is described, and two new combinations are proposed. The homeomorphic valve shapes of M. andreui sp. nov., M. amfibola and ?S. rectum (Loxoconchidae) support the value of the approach adopted herein: analysis of subtle morphological details with high-resolution microscopy.

Circulating low density neutrophils of breast cancer patients are associated with their worse prognosis due to the impairment of T cell responses.

Neutrophils are prominent immune components of tumors, having either anti-tumor (N1) or pro-tumor activity (N2). Circulating neutrophils, divided into high density neutrophils (HDN) and low density neutrophils (LDN), functionally mirror those N1 and N2 cells, respectively. LDN are rare in non-pathological conditions, but frequent in cancer, exhibiting a pro-tumor phenotype. These findings have been mainly demonstrated in animal models, thus proper validation in humans is still imperative. Here, we observed that LDN were increased in the blood of breast cancer (BC) patients, particularly with metastatic disease. Within the population of non-metastatic patients, LDN were more prevalent in patients with poor response to neoadjuvant chemotherapy than patients with a good response. The higher incidence of LDN in BC patients with severe disease or resistance to treatment can be explained by their pro-tumor/immunosuppressive characteristics. Moreover, the percentage of LDN in BC patients' blood was negatively correlated with activated cytotoxic T lymphocytes and positively correlated with immunosuppressive regulatory T cells. The ability of LDN to spoil anti-tumor immune responses was further demonstrated ex vivo. Hence, this study reveals the potential of LDN as a biomarker of BC response to treatment and opens new avenues for developing new immunotherapies.

Expression of HLA-DR in Cytotoxic T Lymphocytes: A Validated Predictive Biomarker and a Potential Therapeutic Strategy in Breast Cancer.

Neoadjuvant chemotherapy (NACT) is common in breast cancer (BC) treatment, though more than half of the patients lack an effective response. Therefore, new predictive biomarkers and alternative therapies are crucial. Previously, we proposed HLA-DR-expressing cytotoxic T lymphocytes (CTLs) as a potential biomarker of the response to NACT. To validate this observation and further investigate these cells, 202 BC patients were enrolled. Flow cytometry analyses were performed in 61 biopsies and 41 blood samples pre-NACT and 100 non-NACT tumor samples. All the patients were followed up for 34 months. Blood-isolated immune cells were cultured with BC cell lines in a 3D system. We confirmed that HLA-DR level in CTLs is a highly sensitive, specific, and independent biomarker to predict response to NACT and developed a predictive probability model. This biomarker was also associated with progression-free survival, regardless of the treatment. The clinical observations are substantiated by the anti-tumor properties of HLA-DR-expressing CTLs. Intriguingly, HLA-DR level in CTLs can be modulated ex vivo, boosting their capacity to kill tumor cells synergistically with doxorubicin. Thus, HLA-DR expression in CTLs is a validated tool to select patients that will actually benefit from NACT, and its stimulation might be a novel therapeutic approach for BC.

Development of Dl1.72, a Novel Anti-DLL1 Antibody with Anti-Tumor Efficacy against Estrogen Receptor-Positive Breast Cancer.

The Notch-signaling ligand DLL1 has emerged as an important player and promising therapeutic target in breast cancer (BC). DLL1-induced Notch activation promotes tumor cell proliferation, survival, migration, angiogenesis and BC stem cell maintenance. In BC, DLL1 overexpression is associated with poor prognosis, particularly in estrogen receptor-positive (ER+) subtypes. Directed therapy in early and advanced BC has dramatically changed the natural course of ER+ BC; however, relapse is a major clinical issue, and new therapeutic strategies are needed. Here, we report the development and characterization of a novel monoclonal antibody specific to DLL1. Using phage display technology, we selected an anti-DLL1 antibody fragment, which was converted into a full human IgG1 (Dl1.72). The Dl1.72 antibody exhibited DLL1 specificity and affinity in the low nanomolar range and significantly impaired DLL1-Notch signaling and expression of Notch target genes in ER+ BC cells. Functionally, in vitro treatment with Dl1.72 reduced MCF-7 cell proliferation, migration, mammosphere formation and endothelial tube formation. In vivo, Dl1.72 significantly inhibited tumor growth, reducing both tumor cell proliferation and liver metastases in a xenograft mouse model, without apparent toxicity. These findings suggest that anti-DLL1 Dl1.72 could be an attractive agent against ER+ BC, warranting further preclinical investigation.

Immunohistochemical analysis of BRCA1 and acetyl-histone H3 in squamous cell carcinoma of the mobile tongue.

OBJECTIVE: The objective of this study was to evaluate the immunoexpression profiles of breast cancer 1 (BRCA1) and acetyl-histone H3 (AcH3) in squamous cell carcinoma of the mobile tongue (SCC-MT) and their correlation with epidemiologic data and the histopathological grade of tumors. STUDY DESIGN: Incisional biopsies of 43 SCC-MT were submitted to immunohistochemistry for AcH3 and BRCA1. Samples were microscopically graded as well differentiated (n = 21) or poorly differentiated (n = 22). Both groups were submitted to statistical analysis (P < .05) regarding the percentage of positive cells. RESULTS: Thirty-nine cases were positive for AcH3 (91%), but no difference was observed for the histologic grading (P = .27). Positivity for BRCA1 was observed in all samples regardless of their cellular locations. Most cases in the poorly differentiated group presented with less than 10% nuclear staining (P < .01) and a predominance of cytoplasmic staining (P = .034). The well-differentiated group showed nuclear staining in most of the cases, with more than 50% of cells staining positive (P < .01). CONCLUSION: AcH3 and BRCA1 were expressed in all samples. There was a significant decrease in cytoplasmic BRCA1 expression in the poorly differentiated group, suggesting BRCA1 as a possible prognostic marker for SCC-MT.

"Having a Baby Can Wait": Experiences of a Sexual and Reproductive Health Promotion Program in the Context of Homelessness among Asian American, Native Hawaiian, and Other Pacific Islander Youth Captured Through PhotoVoice.

Nearly half of female youth experiencing homelessness (YEH) become pregnant due to myriad individual, family, community, and structural factors. In response, our team developed and tested Wahine ("woman") Talk, a multilevel, trauma-informed sexual and reproductive health intervention created with and for female YEH aged 14 to 22. After Wahine Talk, youth were invited to participate in a participatory action PhotoVoice project regarding experiences of the program, waiting to start or expand their families, and homelessness. Photographs were taken and captioned by youth, and then further examined through Thematic Analysis. Final project themes include (a) Youth-Driven Birth Timing Decisions; (b) A Sense of Place: Finding Safe Spaces; and (c) Glimpsing Hope. Because YEH live under society's radar, it is critical to understand their experiences from their own perspectives to improve interventions at multiple levels. Implications for meeting the needs of YEH in the areas of reproductive justice, financial stability, and affordable housing are discussed.

Establishment of a 3D Co-culture With MDA-MB-231 Breast Cancer Cell Line and Patient-Derived Immune Cells for Application in the Development of Immunotherapies.

3D cell culture including different cell types, such as immune cells, is a representative platform that mimics the tumor microenvironment. Here we disclose an easy-to-handle 3D co-culture protocol using a scaffold-free technique with the breast cancer cell line MDA-MB-231 and breast cancer patient-derived immune cells from peripheral blood. The method presented is simple, less time-consuming and less expensive when compared to other 3D techniques. Additionally, this is an optimized protocol for the establishment of a 3D system for this cell line, which is normally seen as challenging to spontaneously form spheroids. The addition of patient-derived immune cells to the cancer cells' spheroid allows the study of the crosstalk between both cell types, as well as the assessment of individual therapeutic approaches to intensify the antitumor immune response. In fact, with this model, we observed that patients' immune cells exhibit a wide range of antitumor responses and we further demonstrated that it is possible to manipulate the less effective ones with a canonical stimulus, as a proof-of-concept, in order to improve their ability to lower the viability of tumor cells. Therefore, this platform could be applied for a personalized immune-based drug screening, with results after a maximum of 10 days of culture, in order to develop more tailored breast cancer treatments and ameliorate patients' survival rate.

Evaluating SARS-CoV-2 Seroconversion Following Relieve of Confinement Measures.

Seroprevalence studies are crucial both for estimating the prevalence of SARS-CoV-2 exposure and to provide a measure for the efficiency of the confinement measures. Portuguese universities were closed on March 16th 2020, when Portugal only registered 62 SARS-CoV-2 infection cases per million. We have validated a SARS-CoV-2 ELISA assay to a stabilized full-length spike protein using 216 pre-pandemic and 19 molecularly diagnosed SARS-CoV-2 positive individual's samples. At NOVA University of Lisbon, presential work was partially resumed on May 25th with staggered schedules. From June 15th to 30th, 3-4 weeks after the easing of confinement measures, we screened 1,636 collaborators of NOVA university of Lisbon for the presence of SARS-CoV-2 spike specific IgA and IgG antibodies. We found that spike-specific IgG in 50 of 1,636 participants (3.0%), none of which had anti-spike IgA antibodies. As participants self-reported as asymptomatic or paucisymptomatic, our study also provides a measurement of the prevalence of asymptomatic/paucisymptomatic SARS-CoV-2 infections. Our study suggests that essential workers have a 2-fold increase in viral exposure, when compared to non-essential workers that observed confinement. Additional serological surveys in different population subgroups will paint a broader picture of the effect of the confinement measures in the broader community.