Converging cytokine and metabolite networks shape asymmetric T cell fate at the term human maternal-fetal interface.

Placentation presents immune conflict between mother and fetus, yet in normal pregnancy maternal immunity against infection is maintained without expense to fetal tolerance. This is believed to result from adaptations at the maternal-fetal interface (MFI) which affect T cell programming, but the identities (i.e., memory subsets and antigenic specificities) of T cells and the signals that mediate T cell fates and functions at the MFI remain poorly understood. We found intact recruitment programs as well as pro-inflammatory cytokine networks that can act on maternal T cells in an antigen-independent manner. These inflammatory signals elicit T cell expression of co-stimulatory receptors necessary for tissue retention, which can be engaged by local macrophages. Although pro-inflammatory molecules elicit T cell effector functions, we show that additional cytokine (TGF-ß1) and metabolite (kynurenine) networks may converge to tune T cell function to those of sentinels. Together, we demonstrate an additional facet of fetal tolerance, wherein T cells are broadly recruited and restrained in an antigen-independent, cytokine/metabolite-dependent manner. These mechanisms provide insight into antigen-nonspecific T cell regulation, especially in tissue microenvironments where they are enriched.

Hallmarks of tumor-experienced T cells are absent in multiple myeloma patients from diagnosis through maintenance therapy.

Dysregulation of the bone marrow (BM) niche in multiple myeloma (MM) alters the composition and state of resident immune cells, potentially impeding anti-tumor immunity. One common mechanism of immune inhibition in solid tumors is the induction of exhaustion in tumor-specific T cells. However, the extent of T cell tumor recognition and exhaustion is not well-characterized in MM. As the specific mechanisms of immune evasion are critical for devising effective therapeutic strategies, we deeply profiled the CD8+ T cell compartment of newly-diagnosed MM (NDMM) patients for evidence of tumor reactivity and T cell exhaustion. We applied single-cell multi-omic sequencing and antigen-specific mass cytometry to longitudinal BM and peripheral blood (PB) samples taken from timepoints spanning from diagnosis through induction therapy, autologous stem cell transplant (ASCT), and maintenance therapy. We identified an exhausted-like population that lacked several canonical exhaustion markers, was not significantly enriched in NDMM patients, and consisted of small, nonpersistent clones. We also observed an activated population with increased frequency in the PB of NDMM patients exhibiting phenotypic and clonal features consistent with homeostatic, antigen-nonspecific activation. However, there was no evidence of "tumor-experienced" T cells displaying hallmarks of terminal exhaustion and/or tumor-specific activation/expansion in NDMM patients at any timepoint.

Multi-omic profiling reveals the endogenous and neoplastic responses to immunotherapies in cutaneous T cell lymphoma.

Cutaneous T cell lymphomas (CTCLs) are skin cancers with poor survival rates and limited treatments. While immunotherapies have shown some efficacy, the immunological consequences of administering immune-activating agents to CTCL patients have not been systematically characterized. We apply a suite of high-dimensional technologies to investigate the local, cellular, and systemic responses in CTCL patients receiving either mono- or combination anti-PD-1 plus interferon-gamma (IFN-γ) therapy. Neoplastic T cells display no evidence of activation after immunotherapy. IFN-γ induces muted endogenous immunological responses, while anti-PD-1 elicits broader changes, including increased abundance of CLA+CD39+ T cells. We develop an unbiased multi-omic profiling approach enabling discovery of immune modules stratifying patients. We identify an enrichment of activated regulatory CLA+CD39+ T cells in non-responders and activated cytotoxic CLA+CD39+ T cells in leukemic patients. Our results provide insights into the effects of immunotherapy in CTCL patients and a generalizable framework for multi-omic analysis of clinical trials.

Characterization of neoantigen-specific T cells in cancer resistant to immune checkpoint therapies.

Neoantigen-specific T cells are strongly implicated as being critical for effective immune checkpoint blockade treatment (ICB) (e.g., anti-PD-1 and anti-CTLA-4) and are being targeted for vaccination-based therapies. However, ICB treatments show uneven responses between patients, and neoantigen vaccination efficiency has yet to be established. Here, we characterize neoantigen-specific CD8+ T cells in a tumor that is resistant to ICB and neoantigen vaccination. Leveraging the use of mass cytometry combined with multiplex major histocompatibility complex (MHC) class I tetramer staining, we screened and identified tumor neoantigen-specific CD8+ T cells in the Lewis Lung carcinoma (LLC) tumor model (mRiok1). We observed an expansion of mRiok1-specific CD8+ tumor-infiltrating lymphocytes (TILs) after ICB targeting PD-1 or CTLA-4 with no sign of tumor regression. The expanded neoantigen-specific CD8+ TILs remained phenotypically and functionally exhausted but displayed cytotoxic characteristics. When combining both ICB treatments, mRiok1-specific CD8+ TILs showed a stem-like phenotype and a higher capacity to produce cytokines, but tumors did not show signs of regression. Furthermore, combining both ICB treatments with neoantigen vaccination did not induce tumor regression either despite neoantigen-specific CD8+ TIL expansion. Overall, this work provides a model for studying neoantigens in an immunotherapy nonresponder model. We showed that a robust neoantigen-specific T-cell response in the LLC tumor model could fail in tumor response to ICB, which will have important implications in designing future immunotherapeutic strategies.

High-Throughput Screening Enhances Kidney Organoid Differentiation from Human Pluripotent Stem Cells and Enables Automated Multidimensional Phenotyping.

Organoids derived from human pluripotent stem cells are a potentially powerful tool for high-throughput screening (HTS), but the complexity of organoid cultures poses a significant challenge for miniaturization and automation. Here, we present a fully automated, HTS-compatible platform for enhanced differentiation and phenotyping of human kidney organoids. The entire 21-day protocol, from plating to differentiation to analysis, can be performed automatically by liquid-handling robots, or alternatively by manual pipetting. High-content imaging analysis reveals both dose-dependent and threshold effects during organoid differentiation. Immunofluorescence and single-cell RNA sequencing identify previously undetected parietal, interstitial, and partially differentiated compartments within organoids and define conditions that greatly expand the vascular endothelium. Chemical modulation of toxicity and disease phenotypes can be quantified for safety and efficacy prediction. Screening in gene-edited organoids in this system reveals an unexpected role for myosin in polycystic kidney disease. Organoids in HTS formats thus establish an attractive platform for multidimensional phenotypic screening.

Androgens Exert a Cysticidal Effect upon Taenia crassiceps by Disrupting Flame Cell Morphology and Function.

The effects of testosterone (T4) and dihydrotestosterone (DHT) on the survival of the helminth cestode parasite Taenia crassiceps, as well as their effects on actin, tubulin and myosin expression and their assembly into the excretory system of flame cells are described in this paper. In vitro evaluations on parasite viability, flow cytometry, confocal microscopy, video-microscopy of live flame cells, and docking experiments of androgens interacting with actin, tubulin, and myosin were conducted. Our results show that T4 and DHT reduce T. crassiceps viability in a dose- and time-dependent fashion, reaching 90% of mortality at the highest dose used (40 ng/ml) and time exposed (10 days) in culture. Androgen treatment does not induce differences in the specific expression pattern of actin, tubulin, and myosin isoforms as compared with control parasites. Confocal microscopy demonstrated a strong disruption of the parasite tegument, with reduced assembly, shape, and motion of flame cells. Docking experiments show that androgens are capable of affecting parasite survival and flame cell morphology by directly interacting with actin, tubulin and myosin without altering their protein expression pattern. We show that both T4 and DHT are able to bind actin, tubulin, and myosin affecting their assembly and causing parasite intoxication due to impairment of flame cell function. Live flame cell video microscopy showing a reduced motion as well changes in the shape of flame cells are also shown. In summary, T4 and DHT directly act on T. crassiceps cysticerci through altering parasite survival as well as the assembly and function of flame cells.

Oestradiol and progesterone differentially alter cytoskeletal protein expression and flame cell morphology in Taenia crassiceps.

We examined the effects of oestradiol (E2) and progesterone (P4) on cytoskeletal protein expression in the helminth Taenia crassiceps - specifically actin, tubulin and myosin. These proteins assemble into flame cells, which constitute the parasite excretory system. Total protein extracts were obtained from E2- and P4-treated T. crassiceps cysticerci and untreated controls, and analysed by one- and two-dimensional protein electrophoresis, flow cytometry, immunofluorescence and videomicroscopy. Exposure of T. crassiceps cysticerci to E2 and P4 induced differential protein expression patterns compared with untreated controls. Changes in actin, tubulin and myosin expression were confirmed by flow cytometry of parasite cells and immunofluorescence. In addition, parasite morphology was altered in response to E2 and P4 versus controls. Flame cells were primarily affected at the level of the ciliary tuft, in association with the changes in actin, tubulin and myosin. We conclude that oestradiol and progesterone act directly on T. crassiceps cysticerci, altering actin, tubulin and myosin expression and thus affecting the assembly and function of flame cells. Our results increase our understanding of several aspects of the molecular crosstalk between host and parasite, which might be useful in designing anthelmintic drugs that exclusively impair parasitic proteins which mediate cell signaling and pathogenic reproduction and establishment.

Analysis of the expression of cytoskeletal proteins of Taenia crassiceps ORF strain cysticerci (Cestoda).

The Taenia crassiceps ORF strain is used to generate a murine model of cysticercosis, which is used for diagnosis, evaluation of drugs, and vaccination. This particular strain only exists as cysticerci, is easily maintained under in vivo and in vitro conditions, and offers an excellent model for studying the cytoskeletons of cestodes. In this study, several experimental approaches were used to determine the tissue expression of its cytoskeletal proteins. The techniques used were microscopy (video, confocal, and transmission electron), one-dimensional (1D) and two-dimensional (2D) electrophoresis, immunochemistry, and mass spectrometry. The tissue expression of actin, tubulin, and paramyosin was assessed using microscopy, and their protein isoforms were determined with 1D and 2D electrophoresis and immunochemistry. Nineteen spots were excised from a proteomic gel and identified by liquid chromatography-tandem mass spectrometry and immunochemistry. The proteins identified were classic cytoskeletal proteins, metabolic enzymes, and proteins with diverse biological functions, but mainly involved in detoxification activities. Research suggests that most noncytoskeletal proteins interact with actin or tubulin, and the results of the present study suggest that the proteins identified may be involved in supporting the dynamics and plasticity of the cytoskeleton of T. crassiceps cysticerci. These results contribute to our knowledge of the cellular biology and physiology of cestodes.

Effect of chronic pesticide exposure in farm workers of a Mexico community.

Pesticides are frequently used substances worldwide, even when the use of some of them is forbidden due to the recognized adverse effect they have on the health of not only the people who apply the pesticides, but also of those that consume the contaminated products. The objectives of this study were to know the health issues of farm workers chronically exposed to pesticides, to evaluate possible damage at genetic level, as well as to explore some hepatic, renal, and hematological alterations. A transversal comparative study was performed between 2 groups, one composed of 25 farm workers engaged in pesticide spraying, and a control group of 21 workers not exposed to pesticides; both groups belonged to the Nextipac community in Jalisco, Mexico. Each member of both groups underwent a full medical history. Blood samples were taken from all farm workers in order to obtain a complete blood count and chemistry, clinical chemistry, lipid profile, liver and kidney function tests, erythrocyte cholinesterase quantification, lipid peroxidation profile, and free DNA fragment quantification. For the information analysis, central tendency and dispersion measurements were registered. In order to know the differences between groups, a cluster multivariate method was used, as well as prevalence reasons. The most used pesticides were mainly organophosphates, triazines and organochlorine compounds. The exposed group showed acute poisoning (20% of the cases) and diverse alterations of the digestive, neurological, respiratory, circulatory, dermatological, renal, and reproductive system probably associated to pesticide exposure. More importantly, they presented free DNA fragments in plasma (90.8 vs 49.05 ng/mL) as well as a higher level of lipid peroxidation (41.85 vs. 31.91 nmol/mL) in comparison with those data from unexposed farm workers. These results suggest that there exist health hazards for those farm workers exposed to pesticides, at organic and cellular levels.

Visualization and 3D reconstruction of flame cells of Taenia solium (cestoda).

BACKGROUND: Flame cells are the terminal cells of protonephridial systems, which are part of the excretory systems of invertebrates. Although the knowledge of their biological role is incomplete, there is a consensus that these cells perform excretion/secretion activities. It has been suggested that the flame cells participate in the maintenance of the osmotic environment that the cestodes require to live inside their hosts. In live Platyhelminthes, by light microscopy, the cells appear beating their flames rapidly and, at the ultrastructural, the cells have a large body enclosing a tuft of cilia. Few studies have been performed to define the localization of the cytoskeletal proteins of these cells, and it is unclear how these proteins are involved in cell function. METHODOLOGY/PRINCIPAL FINDINGS: Parasites of two different developmental stages of T. solium were used: cysticerci recovered from naturally infected pigs and intestinal adults obtained from immunosuppressed and experimentally infected golden hamsters. Hamsters were fed viable cysticerci to recover adult parasites after one month of infection. In the present studies focusing on flame cells of cysticerci tissues was performed. Using several methods such as video, confocal and electron microscopy, in addition to computational analysis for reconstruction and modeling, we have provided a 3D visual rendition of the cytoskeletal architecture of Taenia solium flame cells. CONCLUSIONS/SIGNIFICANCE: We consider that visual representations of cells open a new way for understanding the role of these cells in the excretory systems of Platyhelminths. After reconstruction, the observation of high resolution 3D images allowed for virtual observation of the interior composition of cells. A combination of microscopic images, computational reconstructions and 3D modeling of cells appears to be useful for inferring the cellular dynamics of the flame cell cytoskeleton.

Expression of NK cells activation receptors after occupational exposure to toxics: a preliminary study.

The expression of NK cells activation receptors was assessed by comparative study of two groups of women workers at a chemical reagents factory, located in Zapopan, Jalisco, Mexico. Twenty of them were exposed to environmental toxics identified and quantified by gas chromatography, and 20 women unexposed to toxic substances. The expression of the surface markers CD56+ and CD3+, and of the activation receptors and co-receptors on NK cells was quantified by flow cytometry. To assess the cellular damage produced by chronic exposure to the toxics, the thiobarbituric acid reacting substances (TBARS) generated and the total plasma antioxidizing capacity (TPAC) were quantified in both groups. The exposed women had been exposed at least to 12 volatile toxic compounds, benzene, benz(a)pyrene, ethylbenzene, dimethylbenz(a)anthracene, xylene, toluene, styrene, chloroform, formaldehyde, iodine, chlorine and fluorine. Significant difference between the two groups was in the proportion of CD3 lymphocytes, 72.7+/-10.3% in the unexposed women versus 66.8+/-7.9% in the exposed group (p<0.05). The density of expression of NKG2D and NKp30 receptors was significantly higher in the unexposed women compared to the exposed group: NKG2D were 31.3+/-6.3 and NKp30 were 9.5+/-5.2 in the unexposed women and 5.14+/-2.9 (p<0.01) and 4.6+/-1.9 (p<0.05), respectively in the exposed women. No statistically significant differences were found in the expression of NKp80, NKp46 and 2B4 receptors. The concentration of TBARS was lower in women from the unexposed group than the corresponding data from women of the exposed group. However, no significant difference was observed in TPAC between the two groups studied. The results of this preliminary study suggest that from the five activation receptors and co-receptors of NK cells evaluated (NKp30, NKp46, NKp80, NKG2D and 2B4), only NKp30 and NKG2D receptor expression was diminished in women exposed to toxics when compared with data from unexposed women. These results suggest that the occupational exposure to mixture of toxics is one of the important factors in the diminution of the NK cell receptor expression.

[Application of gender perspective in papers published between 2000 and 2003 from four National Institutes of Health journals]. / Aplicación de la perspectiva de género en artículos publicados en cuatro revistas nacionales de salud, México, 2000-2003.

OBJECTIVE: The objective of this study was to determine the application of gender perspective in public health papers in four journals published by the Mexican National Health Institutes. MATERIAL AND METHODS: A total of 999 papers published in the four journals between 2000 and 2003 was reviewed. Two levels of analysis were considered: (a) data presented by sex, providing description of differences between women and men, and (b) the analysis of these differences from gender perspective. RESULTS: One quarter (25.4%) of the articles described results by sex. The largest percentage was published in Salud Pública de México (48.8%) and the smallest in Revista de Investigación Clínica (16.1%). Gender perspective was used in only 4.2% of papers; of these, drug addictions, health behaviour and violence were the topics that occurred most frequently. CONCLUSIONS: This is the first study to assess the application of gender perspective in Mexican health articles. Similarly to other countries, a very small percentage of authors analyzed sex differences, and a smaller proportion applied gender perspective. Results demonstrate the importance of promoting interdisciplinary research that may promote the elucidation of health inequalities between men and women.

Aplicación de la perspectiva de género en artículos publicados en cuatro revistas nacionales de salud, México, 2000-2003 / Application of gender perspective in papers published between 2000 and 2003 from four National Institutes of Health journals

OBJETIVO: Elaborar un diagnóstico sobre la aplicación de la perspectiva de género en el campo de la investigación en salud en cuatro revistas mexicanas de los institutos nacionales de salud. MATERIAL Y MÉTODOS: Se revisaron 999 artículos publicados en cuatro revistas científicas (2000-2003), con dos niveles de análisis: a) desagregación de datos por sexo, que permite describir las diferencias entre mujeres y hombres; y b) análisis de estas diferencias desde la perspectiva de género. RESULTADOS: De los artículos revisados, 25.4 por ciento desagregó sus resultados por sexo, de los cuales el mayor porcentaje se publicó en Salud Pública de México (48.8 por ciento) y el menor en la Revista de Investigación Clínica (16.1 por ciento). Solamente 4.2 por ciento de los artículos aplicó la perspectiva de género, y los temas mayormente abordados fueron las adicciones, el comportamiento sexual y la violencia. CONCLUSIONES: Este es el primer estudio en México que evalúa la aplicación de la perspectiva de género en la investigación en salud. Al igual que en otros países, es baja la proporción de artículos que analizan las diferencias por sexo y aplican la perspectiva de género. Esto destaca la importancia de promover la investigación interdisciplinaria que ayude a comprender los orígenes biológicos y sociales, o la combinación de ambos, que determinan la salud desigual entre mujeres y hombres.

Parejas y Disparejas: para conocer mejor nuestra sexualidad / Couples and uneven: knowing best our sexuality

"Parejas y Disparejas" es un juego dirigido a conocer mejor la sexualidad de cada individuo, que ayuda a pensar sobre su ser y busca ser útil en el difícil proceso de definir que tipo de persona se quiere ser. También es un material de utilidad para padres y madres de familia, maestros y maestras, promotores juveniles, educadores de la salud o simplemente para grupos de amigas o amigos. Contenido: 1) Presentación. 2) Instructivo. 3) Cuerpo. 4) Sensaciones (erotismo). 5) Ser hombre/ser mujer (papeles sexuales). 6) Afectos. 7) Prevención de riesgos. 8) Actividades. 9) Lecturas recomendadas