Divergent expression of Neurl3 from hemogenic endothelial cells to hematopoietic stem progenitor cells during development.

Prior to the generation of hematopoietic stem cells (HSCs) from the hemogenic endothelial cells (HECs) mainly in the dorsal aorta in midgestational mouse embryos, multiple hematopoietic progenitors including erythro-myeloid progenitors and lymphoid progenitors are generated from yolk sac HECs. These HSC-independent hematopoietic progenitors have recently been identified as major contributors to functional blood cell production until birth. However, little is known about yolk sac HECs. Here, combining integrative analyses of multiple single-cell RNA-sequencing datasets and functional assays, we reveal that Neurl3-EGFP, in addition to marking the continuum throughout the ontogeny of HSCs from HECs, can also serve as a single enrichment marker for yolk sac HECs. Moreover, while yolk sac HECs have much weaker arterial characteristics than either arterial endothelial cells in the yolk sac or HECs within the embryo proper, the lymphoid potential of yolk sac HECs is largely confined to the arterial-biased subpopulation featured by the Unc5b expression. Interestingly, the B lymphoid potential of hematopoietic progenitors, but not for myeloid potentials, is exclusively detected in Neurl3-negative subpopulations in midgestational embryos. Taken together, these findings enhance our understanding of blood birth from yolk sac HECs and provide theoretical basis and candidate reporters for monitoring step-wise hematopoietic differentiation.

Nupr1 Negatively Regulates Endothelial to Hematopoietic Transition in the Aorta-Gonad-Mesonephros Region.

In the aorta of mid-gestational mouse embryos, a specialized endothelial subpopulation termed hemogenic endothelial cells (HECs) develops into hematopoietic stem and progenitor cells (HSPCs), through a conserved process of endothelial-to-hematopoietic transition (EHT). EHT is tightly controlled by multiple intrinsic and extrinsic mechanisms. Nevertheless, the molecular regulators restraining this process remain poorly understood. Here, it is uncovered that, one of the previously identified HEC signature genes, Nupr1, negatively regulates the EHT process. Nupr1 deletion in endothelial cells results in increased HSPC generation in the aorta-gonad-mesonephros region. Furthermore, single-cell transcriptomics combined with serial functional assays reveals that loss of Nupr1 promotes the EHT process by promoting the specification of hematopoiesis-primed functional HECs and strengthening their subsequent hematopoietic differentiation potential toward HSPCs. This study further finds that the proinflammatory cytokine, tumor necrosis factor α (TNF-α), is significantly upregulated in Nupr1-deficient HECs, and the use of a specific TNF-α neutralizing antibody partially reduces excessive HSPC generation in the explant cultures from Nupr1-deficient embryos. This study identifies a novel negative regulator of EHT and the findings indicate that Nupr1 is a new potential target for future hematopoietic stem cell regeneration research.

Glioblastoma stem cell-specific histamine secretion drives pro-angiogenic tumor microenvironment remodeling.

The communication between glioblastoma stem cells (GSCs) and the surrounding microenvironment is a prominent feature accounting for the aggressive biology of glioblastoma multiforme (GBM). However, the mechanisms by which GSCs proactively drive interactions with microenvironment is not well understood. In this study, we interrogated metabolites that are preferentially secreted from GSCs and found that GSCs produce and secrete histamine to shape a pro-angiogenic tumor microenvironment. This histamine-producing ability is attributed to H3K4me3 modification-activated histidine decarboxylase (HDC) transcription via MYC. Notably, HDC is highly expressed in GBM, which is associated with poor survival of these patients. GSC-secreted histamine activates endothelial cells by triggering a histamine H1 receptor (H1R)-Ca2+-NF-κB axis, thereby promoting angiogenesis and GBM progression. Importantly, pharmacological blockage of H1R using antihistamines impedes the growth of GBM xenografts in mice. Our findings establish that GSC-specific metabolite secretion remodels the tumor microenvironment and highlight histamine targeting as a potential strategy for GBM therapy.

MBD3 promotes hepatocellular carcinoma progression and metastasis through negative regulation of tumour suppressor TFPI2.

BACKGROUND: The mechanism of recurrence and metastasis of hepatocellular carcinoma (HCC) is complex and challenging. Methyl-CpG binding domain protein 3 (MBD3) is a key epigenetic regulator involved in the progression and metastasis of several cancers, but its role in HCC remains unknown. METHODS: MBD3 expression in HCC was detected by immunohistochemistry and its association with clinicopathological features and patient's survival was analysed. The effects of MBD3 on hepatoma cells growth and metastasis were investigated, and the mechanism was explored. RESULTS: MBD3 is significantly highly expressed in HCC, associated with the advanced tumour stage and poor prognosis in HCC patients. MBD3 promotes the growth, angiogenesis and metastasis of HCC cells by inhibiting the tumour suppressor tissue factor pathway inhibitor 2 (TFPI2). Mechanistically, MBD3 can inhibit the TFPI2 transcription via the Nucleosome Remodeling and Deacetylase (NuRD) complex-mediated deacetylation, thus reactivating the activity of matrix metalloproteinases (MMPs) and PI3K/AKT signaling pathway, leading to the progression and metastasis of HCC CONCLUSIONS: Our results unravel the novel regulatory function of MBD3 in the progression and metastasis of HCC and identify MBD3 as an independent unfavourable prognostic factor for HCC patients, suggesting its potential as a promising therapeutic target as well.

Tumor evolution selectively inactivates the core microRNA machinery for immune evasion.

Cancer cells acquire genetic heterogeneity to escape from immune surveillance during tumor evolution, but a systematic approach to distinguish driver from passenger mutations is lacking. Here we investigate the impact of different immune pressure on tumor clonal dynamics and immune evasion mechanism, by combining massive parallel sequencing of immune edited tumors and CRISPR library screens in syngeneic mouse tumor model and co-culture system. We find that the core microRNA (miRNA) biogenesis and targeting machinery maintains the sensitivity of cancer cells to PD-1-independent T cell-mediated cytotoxicity. Genetic inactivation of the machinery or re-introduction of ANKRD52 frequent patient mutations dampens the JAK-STAT-interferon-γ signaling and antigen presentation in cancer cells, largely by abolishing miR-155-targeted silencing of suppressor of cytokine signaling 1 (SOCS1). Expression of each miRNA machinery component strongly correlates with intratumoral T cell infiltration in nearly all human cancer types. Our data indicate that the evolutionarily conserved miRNA pathway can be exploited by cancer cells to escape from T cell-mediated elimination and immunotherapy.

Mitochondrial Ca2+ and cell cycle regulation.

It has been demonstrated for more than 40 years that intracellular calcium (Ca2+) controls a variety of cellular functions, including mitochondrial metabolism and cell proliferation. Cytosolic Ca2+ fluctuation during key stages of the cell cycle can lead to mitochondrial Ca2+ uptake and subsequent activation of mitochondrial oxidative phosphorylation and a range of signaling. However, the relationship between mitochondrial Ca2+ and cell cycle progression has long been neglected because the molecule responsible for Ca2+ uptake has been unknown. Recently, the identification of the mitochondrial Ca2+ uniporter (MCU) has led to key advances. With improved Ca2+ imaging and detection, effects of MCU-mediated mitochondrial Ca2+ have been observed at different stages of the cell cycle. Elevated Ca2+ signaling boosts ATP and ROS production, remodels cytosolic Ca2+ pathways and reprograms cell fate-determining networks. These findings suggest that manipulating mitochondrial Ca2+ signaling may serve as a potential strategy in the control of many crucial biological events, such as tumor development and cell division in hematopoietic stem cells (HSCs). In this review, we summarize the current understanding of the role of mitochondrial Ca2+ signaling during different stages of the cell cycle and highlight the potential physiological and pathological significance of mitochondrial Ca2+ signaling.

AMPK-mediated activation of MCU stimulates mitochondrial Ca2+ entry to promote mitotic progression.

The capacity of cells to alter bioenergetics in response to the demands of various biological processes is essential for normal physiology. The coordination of energy sensing and production with highly energy-demanding cellular processes, such as cell division, is poorly understood. Here, we show that a cell cycle-dependent mitochondrial Ca2+ transient connects energy sensing to mitochondrial activity for mitotic progression. The mitochondrial Ca2+ uniporter (MCU) mediates a rapid mitochondrial Ca2+ transient during mitosis. Inhibition of mitochondrial Ca2+ transients via MCU depletion causes spindle checkpoint-dependent mitotic delay. Cellular ATP levels drop during early mitosis, and the mitochondrial Ca2+ transients boost mitochondrial respiration to restore energy homeostasis. This is achieved through mitosis-specific MCU phosphorylation and activation by the mitochondrial translocation of energy sensor AMP-activated protein kinase (AMPK). Our results establish a critical role for AMPK- and MCU-dependent mitochondrial Ca2+ signalling in mitosis and reveal a mechanism of mitochondrial metabolic adaptation to acute cellular energy stress.

FLI1 Exonic Circular RNAs as a Novel Oncogenic Driver to Promote Tumor Metastasis in Small Cell Lung Cancer.

PURPOSE: The aberrantly upregulated Friend leukemia virus integration 1 (FLI1) is closely correlated with the malignant phenotype of small cell lung cancer (SCLC). It is interesting to note that the CRISPR gene knockout by Cas9 gRNAs that target the FLI1 coding region and the posttranscriptional knockdown by shRNAs that target the 3' region of FLI1 mRNA yielded distinct antimetastasis effects in SCLC cells. This study attempts to examine if FLI1 exonic circular RNAs (FECR) function as a new malignant driver that determines the metastatic phenotype in SCLC. EXPERIMENTAL DESIGN: The clinical relevance of FECRs was examined in 56 primary SCLC tissues and 50 non-small cell lung cancer (NSCLC) tissues. The prognostic value of FECRs was examined by measuring serum exosomal FECRs in a longitudinal cohort of patients with SCLC. The oncogenic activity of FECRs was investigated in both SCLC cell lines and animal xenograft studies. Finally, we explored the molecular mechanisms underlying these noncoding RNAs as a malignant driver. RESULTS: Therapeutic comparison of CRISPR Cas9 knockout and shRNA knockdown of FLI1 identified FECRs as a new noncanonical malignant driver in SCLC. Using RNA FISH and quantitative PCR, we found that FECR1 (exons 4-2-3) and FECR2 (exons 5-2-3-4) were aberrantly upregulated in SCLC tissues (P < 0.0001), and was positively associated with lymph node metastasis (P < 0.01). Notably, serum exosomal FECR1 was associated with poor survival (P = 0.038) and clinical response to chemotherapy. Silencing of FECRs significantly inhibited the migration in two highly aggressive SCLC cell lines and reduced tumor metastasis in vivo. Mechanistically, we uncovered that FECRs sequestered and subsequently inactivated tumor suppressor miR584-3p, leading to the activation of the Rho Associated Coiled-Coil Containing Protein Kinase 1 gene (ROCK1). CONCLUSIONS: This study identifies FLI1 exonic circular RNAs as a new oncogenic driver that promotes tumor metastasis through the miR584-ROCK1 pathway. Importantly, serum exosomal FECR1 may serve as a promising biomarker to track disease progression of SCLC.

The structural and biological properties of hydroxyapatite-modified titanate nanowire scaffolds.

Hydroxyapatite-modified titanate nanowire scaffolds as alternative materials for tissue engineering have been developed via a titanate nanowire matrix assisted electrochemical deposition method. The macroporous titanate nanowire matrix on Ti metal was fabricated by a hydrothermal method, and then followed by an electrochemical synthesis of hydroxyapatite nanoparticles on titanate nanowire. The incorporation of titanate nanowire matrix with high oriented hydroxyapatite nanoparticles generates hierarchical scaffolds with highly osteogenic, structural integrity and excellent mechanical performance. As-prepared porous three dimensional interconnected hydroxyapatite-modified titanate nanowire scaffolds, mimicking the nature's extracellular matrix, could provide a suitable microenvironment for tissue cell ingrowth and differentiation. The ceramic titanate nanowire core with HA nanoparticle sheath structure displays superhydrophilicity, which facilitates the cell attachment and proliferation, and induces the in vitro tissue-engineered bone. Human osteoblast-like MG63 cells were cultured on the hydroxyapatite-modified titanate nanowire scaffolds, and the results showed that the scaffolds highly promote the bioactivity, osteoconductivity and osteoblast differentiation.