Yoshitomi, H. | bioRxiv Science

bioRxiv2021

Tertiary lymphoid structures induced by CXCL13-producing CD4+ T cells increase tumor infiltrating CD8+ T cells and B cells in ovarian cancer

BackgroundTertiary lymphoid structures (TLSs) are transient ectopic lymphoid aggregates whose formation might be caused by chronic inflammation states, such as cancer. The presence of TLS is associated with a favorable prognosis in most solid malignancies. The recognition of the relevance of TLS to cancer has led to a growing interest in TLS as an immunomodulatory target to enhance tumor immunity, although how TLSs are induced in the tumor microenvironment (TME) and how they affect patient survival are not well understood. MethodsTLS distribution in relation to tumor infiltrating lymphocytes (TILs) and related gene expression were investigated in high grade serous ovarian cancer (HGSC) specimens. CXCL13 expression, which is strongly associated with TLS, and its localization in immune cells, were examined. We explored the tumor microenvironment for CXCL13 secretion by adding various inflammatory cytokines in vitro. The induction of TLS by CXCL13 was examined in a mouse model of ovarian cancer. ResultsCXCL13 gene expression correlated with TLS formation and the infiltration of T cells and B cells, and was a favorable prognostic factor for HGSC patients. The coexistence of CD8+ T cells and B-cell lineages in the TME was associated with a better prognosis of HGSC and was closely related to the presence of TLSs. CXCL13 expression was predominantly coincident with CD4+ T cells in TLSs and CD8+ T cells in TILs, and shifted from CD4+ T cells to CD21+ follicular dendritic cells as the TLS matured. Although TGF-{beta} was reported to stimulate CXCL13 production, our in vitro results revealed that CXCL13 secretion was promoted in CD4+ T cells under TGF-{beta} + IL-2-restricted conditions and in CD8+ T cells under TGF-{beta} + IL-12-rich conditions. In a mouse model of ovarian cancer, recombinant CXCL13 induced TLSs and enhanced survival by the infiltration of CD8+ T cells. ConclusionsTLS formation was promoted by CXCL13-producing CD4+ T cells and TLSs facilitated the coordinated antitumor responses of cellular and humoral immunity in ovarian cancer.

immunology↗

bioRxiv2019

Modeling the Human Segmentation Clock with Pluripotent Stem Cells

Pluripotent stem cells (PSCs) have increasingly been used to model different aspects of embryogenesis and organ formation1. Despite recent advances in the in vitro induction of major mesodermal lineages and mesoderm-derived cell types2,3, experimental model systems that can recapitulate more complex biological features of human mesoderm development and patterning are largely missing. Here, we utilized induced pluripotent stem cells (iPSCs) for the stepwise in vitro induction of presomitic mesoderm (PSM) and its derivatives to model distinct aspects of human somitogenesis. We focused initially on modeling the human segmentation clock, a major biological concept believed to underlie the rhythmic and controlled emergence of somites, which give rise to the segmental pattern of the vertebrate axial skeleton. We succeeded to observe oscillatory expression of core segmentation clock genes, including HES7 and DKK1, and identified novel oscillatory genes in human iPSC-derived PSM. We furthermore determined the period of the human segmentation clock to be around five hours and showed the presence of dynamic traveling wave-like gene expression within in vitro induced human PSM. Utilizing CRISPR/Cas9-based genome editing technology, we then targeted genes, for which mutations in patients with abnormal axial skeletal development such as spondylocostal dysostosis (SCD) (HES7, LFNG and DLL3) or spondylothoracic dysostosis (STD) (MESP2) have been reported. Subsequent analysis of patient-like iPSC knock-out lines as well as patient-derived iPSCs together with their genetically corrected isogenic controls revealed gene-specific alterations in oscillation, synchronization or differentiation properties, validating the overall utility of our model system, to recapitulate not only key features of human somitogenesis but also to provide novel insights into diseases associated with the formation and patterning of the human axial skeleton.

developmental biology↗