Cholinergic receptor pathways involved in apoptosis, cell proliferation and neuronal differentiation

Acetylcholine (ACh) has been shown to modulate neuronal differentiation during early development. Both muscarinic and nicotinic acetylcholine receptors (AChRs) regulate a wide variety of physiological responses, including apoptosis, cellular proliferation and neuronal differentiation. However, the intracellular mechanisms underlying these effects of AChR signaling are not fully understood. It is known that activation of AChRs increase cellular proliferation and neurogenesis and that regulation of intracellular calcium through AChRs may underlie the many functions of ACh. Intriguingly, activation of diverse signaling molecules such as Ras-mitogen-activated protein kinase, phosphatidylinositol 3-kinase-Akt, protein kinase C and c-Src is modulated by AChRs. Here we discuss the roles of ACh in neuronal differentiation, cell proliferation and apoptosis. We also discuss the pathways involved in these processes, as well as the effects of novel endogenous AChRs agonists and strategies to enhance neuronal-differentiation of stem and neural progenitor cells. Further understanding of the intracellular mechanisms underlying AChR signaling may provide insights for novel therapeutic strategies, as abnormal AChR activity is present in many diseases

Longitudinal characterization of the tumoral microbiome during radiotherapy in HPV-associated oropharynx cancer

Purpose: To describe the baseline and serial tumor microbiome in HPV-associated oropharynx cancer (OPC) over the course of radiotherapy (RT). Methods: Patients with newly diagnosed HPV-associated OPC treated with definitive radiotherapy +/- concurrent chemotherapy were enrolled in this prospective study. Using 16S rRNA gene sequencing, dynamic changes in the tumor site microbiome during RT were investigated. Surface tumor samples were obtained before RT and at week 1, 3 and 5 of RT. Radiological primary tumor response at mid-treatment was categorized as complete (CR) or partial (PR). Results: Ten patients were enrolled, but 9 patients were included in the final analysis. Mean age was 62 years (range: 51-71). As per AJCC 8th Ed, 56%, 22% and 22% of patients had stage I, II and III, respectively. At 4-weeks, 6 patients had CR and 3 patients had PR; at follow-up imaging post treatment, all patients had CR. The baseline diversity of the tumoral versus buccal microbiome was not statistically different. For the entire cohort, alpha diversity was significantly decreased over the course of treatment (p = 0.04). There was a significant alteration in the bacterial community within the first week of radiation. Baseline tumor alpha diversity of patients with CR was significantly higher than those with PR (p = 0.03). While patients with CR had significant reduction in diversity over the course of radiation (p = 0.01), the diversity remained unchanged in patients with PR. Patients with history of smoking had significantly increased abundance of  Kingella (0.05) and lower abundance of Stomatobaculum (p = 0.03) compared to never smokers. Conclusions: The tumor microbiome of HPV-associated OPC exhibits reduced alpha diversity and altered taxa abundance over the course of radiotherapy. The baseline bacterial profiles of smokers vs. non-smokers were inherently different. Baseline tumor alpha diversity of patients with CR was higher than patients with PR, suggesting that the microbiome deserves further investigation as a biomarker of radiation response

Circulating neutrophils and tumor-associated myeloid cells function as a powerful biomarker for response to chemoradiation in locally advanced cervical cancer

Purpose: The immune system’s role in mediating the cytotoxic effects of chemoradiotherapy remains not completely understood. The integration of immunotherapies into treatment will require insight into features and timing of the immune microenvironment associated with treatment response. Here, we investigated the role of circulating neutrophils and tumor-associated myeloid cells (TSAMs) as potential agents and biomarkers for disease-related outcomes in locally advanced cervical cancer (LACC). Material and Methods: Hematologic parameters for two LACC patient cohorts, a retrospective clinical and a prospective translational cohort, were obtained at baseline, weekly during chemoradiotherapy for the retrospective cohort, biweekly during chemoradiotherapy for the prospective cohort, and at the first follow-up visit for both cohorts (mean 14.7 weeks, range 8.1–25.1 weeks for the prospective cohort and 5.3 weeks with a range of 2.7–9.0 weeks for the retrospective cohort). In both cohorts, baseline as well as mean and lowest on-treatment values for platelets, hemoglobin, absolute neutrophil count (ANC), and absolute lymphocyte count (ALC) were analyzed for correlations with disease-related outcomes. In the prospective cohort, circulating myeloid cells were isolated from peripheral blood mononuclear cells (PBMCs), and TSAMs were isolated from tumor tissue via a novel serial cytobrush sampling assay. The samples were analyzed by flow cytometry. Results: In both cohorts, the only hematologic parameter significantly associated with survival was elevated on-treatment mean ANC (mANC), which was associated with lower local failure-free and overall survival rates in the retrospective and prospective cohorts, respectively. mANC was not associated with a difference in distant metastases. CD11b+CD11c- TSAMs, which act as a surrogate marker for intratumoral neutrophils, steadily decreased during the course of chemoRT and nadier’d at week 5 of treatment. Conversely, circulating myeloid cells identified from PBMCs steadily increased through week 5 of treatment. Regression analysis confirmed an inverse relationship between circulating myeloid cells and TSAMs at this time point. Conclusions: These findings identify on-treatment mean neutrophil count as a predictor of disease-related outcomes, suggest that neutrophils contribute to chemoradiation treatment resistance, and demonstrate the importance of techniques to measure intratumoral immune activity

Ascitic fluid cell CDR3β motifs associated with prognosis.

(A) Examples of CD4+ T cell receptor (TCR) motifs associated with different prognoses. (B) Examples of CD8+ TCR motifs associated with different prognoses. The figures show the logo and phylogenetic tree of the clustered CDR3β peptides for each motif. The phylogenetic tree also includes peptides found as the best hit from McPAS; the complete set of motifs associated with excellent or poor/worst prognosis is shown in S7A Table. (C) Association networks constructed to form the cluster of motifs associated with response to platinum therapy. Nodes in the networks indicate individual patients; the color of the node indicates prognosis. Edges in the networks indicate an association between a pair of samples if they share GLIPH specificity groups associated with prognosis. The networks validated the statistical algorithm used to identify prognosis-associated specificity groups.</p

Gating strategy for the detection of different immune cell subsets from malignant ascites of ovarian cancer patients.

Cells collected from ascites were stained with a 9-color flow cytometry panel and analyzed in an X-20 Fortessa flow cytometer. (A) The gating strategy is depicted starting with the detection of lymphocytes, followed by live cells and CD3+ T cells that are separated according to CD4 and CD8 expression. (B) Fluorescence-minus-one (FMO) staining is shown for the different markers analyzed on CD4+ and CD8+ T cells. (PDF)</p

Increased T cell receptor productive rearrangements in ascites of patients with <i>BRCA1/2</i> tumor mutations.

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Difference in T cell receptor (TCR) characteristics between ascites of patients grouped in Cluster A and ascites of patients grouped in the remaining clusters (others).

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