Synthetic Developmental Biology: Understanding Through Reconstitution.

Reconstitution is an experimental strategy that seeks to recapitulate biological events outside their natural contexts using a reduced set of components. Classically, biochemical reconstitution has been extensively applied to identify the minimal set of molecules sufficient for recreating the basic chemistry of life. By analogy, reconstitution approaches to developmental biology recapitulate aspects of developmental events outside an embryo, with the goal of revealing the basic genetic circuits or physical cues sufficient for recreating developmental decisions. The rapidly growing repertoire of genetic, molecular, microscopic, and bioengineering tools is expanding the complexity and precision of reconstitution experiments. We review the emerging field of synthetic developmental biology, with a focus on the ways in which reconstitution strategies and new biological tools have enhanced our modern understanding of fundamental questions in developmental biology.

Hematopoietic stem cell arrival triggers dynamic remodeling of the perivascular niche.

Hematopoietic stem and progenitor cells (HSPCs) can reconstitute and sustain the entire blood system. We generated a highly specific transgenic reporter of HSPCs in zebrafish. This allowed us to perform high-resolution live imaging on endogenous HSPCs not currently possible in mammalian bone marrow. Using this system, we have uncovered distinct interactions between single HSPCs and their niche. When an HSPC arrives in the perivascular niche, a group of endothelial cells remodel to form a surrounding pocket. This structure appears conserved in mouse fetal liver. Correlative light and electron microscopy revealed that endothelial cells surround a single HSPC attached to a single mesenchymal stromal cell. Live imaging showed that mesenchymal stromal cells anchor HSPCs and orient their divisions. A chemical genetic screen found that the compound lycorine promotes HSPC-niche interactions during development and ultimately expands the stem cell pool into adulthood. Our studies provide evidence for dynamic niche interactions upon stem cell colonization. PAPERFLICK:

Lineage regulators direct BMP and Wnt pathways to cell-specific programs during differentiation and regeneration.

BMP and Wnt signaling pathways control essential cellular responses through activation of the transcription factors SMAD (BMP) and TCF (Wnt). Here, we show that regeneration of hematopoietic lineages following acute injury depends on the activation of each of these signaling pathways to induce expression of key blood genes. Both SMAD1 and TCF7L2 co-occupy sites with master regulators adjacent to hematopoietic genes. In addition, both SMAD1 and TCF7L2 follow the binding of the predominant lineage regulator during differentiation from multipotent hematopoietic progenitor cells to erythroid cells. Furthermore, induction of the myeloid lineage regulator C/EBPα in erythroid cells shifts binding of SMAD1 to sites newly occupied by C/EBPα, whereas expression of the erythroid regulator GATA1 directs SMAD1 loss on nonerythroid targets. We conclude that the regenerative response mediated by BMP and Wnt signaling pathways is coupled with the lineage master regulators to control the gene programs defining cellular identity.

Author Correction: Epoxyeicosatrienoic acids enhance embryonic haematopoiesis and adult marrow engraftment.

An Amendment to this paper has been published and can be accessed via a link at the top of the paper.

LncRNA MIR210HG promotes phenotype switching of pulmonary arterial smooth muscle cells through autophagy-dependent ferroptosis pathway.

Hypoxic pulmonary hypertension (HPH) is a pathophysiological syndrome in which pulmonary vascular pressure increases under hypoxic stimulation and there is an urgent need to develop emerging therapies for the treatment of HPH. LncRNA MIR210HG is a long non-coding RNA closely related to hypoxia and has been widely reported in a variety of tumor diseases. But its mechanism in hypoxic pulmonary hypertension is not clear. In this study, we identified for the first time the potential effect of MIR210HG on disease progression in HPH. Furthermore, we investigated the underlying mechanism through which elevated levels of MIR210HG promotes the transition from a contractile phenotype to a synthetic phenotype in PASMCs under hypoxia via activation of autophagy-dependent ferroptosis pathway. While overexpression of HIF-2α in PASMCs under hypoxia significantly reversed the phenotypic changes induced by MIR210HG knockdown. We further investigated the potential positive regulatory relationship between STAT3 and the transcription of MIR210HG in PASMCs under hypoxic conditions. In addition, we established both in vivo and in vitro models of HPH to validate the differential expression of specific markers associated with hypoxia. Our findings suggest a potential mechanism of LncRNA MIR210HG in the progression of HPH and offer potential targets for disease intervention and treatment.

Hematopoietic stem cell development is dependent on blood flow.

During vertebrate embryogenesis, hematopoietic stem cells (HSCs) arise in the aorta-gonads-mesonephros (AGM) region. We report here that blood flow is a conserved regulator of HSC formation. In zebrafish, chemical blood flow modulators regulated HSC development, and silent heart (sih) embryos, lacking a heartbeat and blood circulation, exhibited severely reduced HSCs. Flow-modifying compounds primarily affected HSC induction after the onset of heartbeat; however, nitric oxide (NO) donors regulated HSC number even when treatment occurred before the initiation of circulation, and rescued HSCs in sih mutants. Morpholino knockdown of nos1 (nnos/enos) blocked HSC development, and its requirement was shown to be cell autonomous. In the mouse, Nos3 (eNos) was expressed in HSCs in the AGM. Intrauterine Nos inhibition or embryonic Nos3 deficiency resulted in a reduction of hematopoietic clusters and transplantable murine HSCs. This work links blood flow to AGM hematopoiesis and identifies NO as a conserved downstream regulator of HSC development.

Communication codes in developmental signaling pathways.

A handful of core intercellular signaling pathways play pivotal roles in a broad variety of developmental processes. It has remained puzzling how so few pathways can provide the precision and specificity of cell-cell communication required for multicellular development. Solving this requires us to quantitatively understand how developmentally relevant signaling information is actively sensed, transformed and spatially distributed by signaling pathways. Recently, single cell analysis and cell-based reconstitution, among other approaches, have begun to reveal the 'communication codes' through which information is represented in the identities, concentrations, combinations and dynamics of extracellular ligands. They have also revealed how signaling pathways decipher these features and control the spatial distribution of signaling in multicellular contexts. Here, we review recent work reporting the discovery and analysis of communication codes and discuss their implications for diverse developmental processes.

Nivolumab monotherapy or combination therapy with ipilimumab for lung cancer: a systemic review and meta-analysis.

BACKGROUND: The high incidence and mortality of lung cancer have seriously affected human life and health. Nivolumab is a monoclonal antibody that can inhibit programmed death 1 (PD-1) and Ipilimumab is a monoclonal antibody against CTLA-4(cytotoxic T lymphocyte-associated antigen 4), both of which can prevent the immune escape of tumor cells. Our goal was to synthesize evidence from published randomized controlled trials involving the safety and efficacy of either Nivolumab alone or in combination for the treatment of unresectable lung cancer. METHODS: We searched the following electronic databases: PubMed, Embase, and Cochrane libraries, and screened the retrieved records for eligibility. We used the Stata16 software for the analyses. The results of the analysis are expressed as hazard ratios (HRs) or risk ratios (RRs) and their corresponding 95% confidence intervals (CI). RESULTS: The final analysis included seven trials involving 3817 patients. Among patients with advanced lung cancer, patients using immunotherapy had better overall survival (OS), progression-free survival (PFS), and an objective response rate (ORR) than patients receiving chemotherapy. The HR of Nivolumab monotherapy or combination therapy with OS was compared with that of chemotherapy (HR: 0.73, 95% CI 0.64-0.83; HR: 0.67, 95% CI 0.55-0.81), and the HR of PFS was (HR: 0.81, 95% CI 0.69-0.94; HR: 0.67, 95% CI 0.55-0.82). CONCLUSIONS: Immunotherapy has been shown to have more clinically meaningful survival benefits for patients with lung cancer, whether monotherapy or combination immunotherapy. CRD42020213440.

Epoxyeicosatrienoic acids enhance embryonic haematopoiesis and adult marrow engraftment.

Haematopoietic stem and progenitor cell (HSPC) transplant is a widely used treatment for life-threatening conditions such as leukaemia; however, the molecular mechanisms regulating HSPC engraftment of the recipient niche remain incompletely understood. Here we develop a competitive HSPC transplant method in adult zebrafish, using in vivo imaging as a non-invasive readout. We use this system to conduct a chemical screen, and identify epoxyeicosatrienoic acids (EETs) as a family of lipids that enhance HSPC engraftment. The pro-haematopoietic effects of EETs were conserved in the developing zebrafish embryo, where 11,12-EET promoted HSPC specification by activating a unique activator protein 1 (AP-1) and runx1 transcription program autonomous to the haemogenic endothelium. This effect required the activation of the phosphatidylinositol-3-OH kinase (PI(3)K) pathway, specifically PI(3)Kγ. In adult HSPCs, 11,12-EET induced transcriptional programs, including AP-1 activation, which modulate several cellular processes, such as migration, to promote engraftment. Furthermore, we demonstrate that the EET effects on enhancing HSPC homing and engraftment are conserved in mammals. Our study establishes a new method to explore the molecular mechanisms of HSPC engraftment, and discovers a previously unrecognized, evolutionarily conserved pathway regulating multiple haematopoietic generation and regeneration processes. EETs may have clinical application in marrow or cord blood transplantation.

Integrated analysis of the molecular mechanisms in idiopathic pulmonary fibrosis.

Rationale: Idiopathic pulmonary fibrosis (IPF) is one of the most aggressive forms of idiopathic interstitial pneumonia. Some miRNAs may be associated with IPF and may affect the occurrence and development of IPF in various pathways. Many miRNAs and genes that may be involved in the development of IPF have been discovered using chip and high throughput technologies. Methods: We analyzed one miRNA and four mRNA databases. We identified hub genes and pathways related to IPF using GO, KEGG enrichment analysis, gene set variation analysis (GSVA), PPI network construction, and hub gene analysis. A comprehensive analysis of differentially expressed miRNAs (DEMs), predicted miRNA target genes, and differentially expressed genes (DEGs) led to the creation of a miRNA-mRNA regulatory network in IPF. Results: We found 203 DEGs and 165 DEMs that were associated with IPF. The findings of enrichment analyses showed that these DEGs were mainly involved in antimicrobial humoral response, antimicrobial humoral immune response mediated by antimicrobial peptide, extracellular matrix organization, cell killing, and organ or tissue specific immune response. The VEGFA, CDH5, and WNT3A genes overlapped between hub genes and the miRNA-mRNA regulatory network. The miRNAs including miR-199b-5p, miR-140-5p, miR-199a-5p, miR-125A-5p, and miR-107 that we predicted would regulate the VEGFA, CDH5, and WNT3A genes, which were also associated with IPF or other fibrosis-related diseases. GSVA indicated that metabolic processes of UTP and IMP, immune response, regulation of Th2 cell cytokine production, and positive regulation of NK cell-mediated immunity are associated with the pathogenesis and treatment of IPF. These pathways also interact with VEGFA, CDH5, and WNT3A. Conclusion: These findings provide a new research direction for the diagnosis and treatment of IPF.

Specific oxylipins enhance vertebrate hematopoiesis via the receptor GPR132.

Epoxyeicosatrienoic acids (EETs) are lipid-derived signaling molecules with cardioprotective and vasodilatory actions. We recently showed that 11,12-EET enhances hematopoietic induction and engraftment in mice and zebrafish. EETs are known to signal via G protein-coupled receptors, with evidence supporting the existence of a specific high-affinity receptor. Identification of a hematopoietic-specific EET receptor would enable genetic interrogation of EET signaling pathways, and perhaps clinical use of this molecule. We developed a bioinformatic approach to identify an EET receptor based on the expression of G protein-coupled receptors in cell lines with differential responses to EETs. We found 10 candidate EET receptors that are expressed in three EET-responsive cell lines, but not expressed in an EET-unresponsive line. Of these, only recombinant GPR132 showed EET-responsiveness in vitro, using a luminescence-based ß-arrestin recruitment assay. Knockdown of zebrafish gpr132b prevented EET-induced hematopoiesis, and marrow from GPR132 knockout mice showed decreased long-term engraftment capability. In contrast to high-affinity EET receptors, GPR132 is reported to respond to additional hydroxy-fatty acids in vitro, and we found that these same hydroxy-fatty acids enhance hematopoiesis in the zebrafish. We conducted structure-activity relationship analyses using both cell culture and zebrafish assays on diverse medium-chain fatty acids. Certain oxygenated, unsaturated free fatty acids showed high activation of GPR132, whereas unoxygenated or saturated fatty acids had lower activity. Absence of the carbon-1 position carboxylic acid prevented activity, suggesting that this moiety is required for receptor activation. GPR132 responds to a select panel of oxygenated polyunsaturated fatty acids to enhance both embryonic and adult hematopoiesis.

Ubiquitous transgene expression and Cre-based recombination driven by the ubiquitin promoter in zebrafish.

Molecular genetics approaches in zebrafish research are hampered by the lack of a ubiquitous transgene driver element that is active at all developmental stages. Here, we report the isolation and characterization of the zebrafish ubiquitin (ubi) promoter, which drives constitutive transgene expression during all developmental stages and analyzed adult organs. Notably, ubi expresses in all blood cell lineages, and we demonstrate the application of ubi-driven fluorophore transgenics in hematopoietic transplantation experiments to assess true multilineage potential of engrafted cells. We further generated transgenic zebrafish that express ubiquitous 4-hydroxytamoxifen-controlled Cre recombinase activity from a ubi:cre(ERt2) transgene, as well as ubi:loxP-EGFP-loxP-mCherry (ubi:Switch) transgenics and show their use as a constitutive fluorescent lineage tracing reagent. The ubi promoter and the transgenic lines presented here thus provide a broad resource and important advancement for transgenic applications in zebrafish.

Assessing the benefits and safety profile of incorporating poly ADP-ribose polymerase (PARP) inhibitors in the treatment of advanced lung cancer: a thorough systematic review and meta-analysis.

Background: Poly (ADP-Ribose) Polymerase (PARP) inhibitors represent a novel class of drugs that hinder DNA repair mechanisms in tumor cells, leading to cell death. This systematic review aims to evaluate the effectiveness, safety, and potential adverse effects of PARP inhibitors (PARPi) in the management of patients with advanced lung cancer. Materials and Methods: We conducted a comprehensive search for relevant studies in PubMed, Embase, Cochrane, and ClinicalTrials.gov. We extracted primary and secondary outcome measures, including progression-free survival (PFS), overall survival (OS), and adverse events (AEs), from the identified literature for subsequent meta-analysis and systematic review. Results: This study encompassed twelve randomized controlled trials, involving 3,132 patients with advanced lung cancer. In comparison to non-PARPi treatments, the administration of PARPi significantly extended OS (hazard ratio (HR) = 0.90, 95% CI = 0.83-0.97, p = 0.006). However, the difference in PFS did not reach statistical significance. Conclusion: In summary, therapies incorporating PARPi provide a degree of benefit by extending OS in patients with advanced lung cancer. Nonetheless, further trials are necessary to furnish additional evidence regarding the efficacy and safety of PARPi in the treatment of lung cancer.Systematic Review Registration: https://www.crd.york.ac.uk/PROSPERO/, identifier number: CRD42023424673.

Diffusion barriers imposed by tissue topology shape morphogen gradients.

Animals use a small number of morphogens to pattern tissues, but it is unclear how evolution modulates morphogen signaling range to match tissues of varying sizes. Here, we used single molecule imaging in reconstituted morphogen gradients and in tissue explants to determine that Hedgehog diffused extra-cellularly as a monomer, and rapidly transitioned between membrane-confined and -unconfined states. Unexpectedly, the vertebrate-specific protein SCUBE1 expanded Hedgehog gradients by accelerating the transition rates between states without affecting the relative abundance of molecules in each state. This observation could not be explained under existing models of morphogen diffusion. Instead, we developed a topology-limited diffusion model in which cell-cell gaps create diffusion barriers, and morphogens can only overcome the barrier by passing through a membrane-unconfined state. Under this model, SCUBE1 promotes Hedgehog secretion and diffusion by allowing it to transiently overcome diffusion barriers. This multiscale understanding of morphogen gradient formation unified prior models and discovered novel knobs that nature can use to tune morphogen gradient sizes across tissues and organisms.

Safety and efficacy of Paxlovid in the treatment of adults with mild to moderate COVID-19 during the omicron epidemic: a multicentre study from China.

BACKGROUND: Since December 2022, the Omicron variant has led to a widespread pandemic in China. The study was to explore the safety and effectiveness of Paxlovid for the treatment of coronavirus disease 2019 (COVID-19). RESEARCH DESIGN AND METHODS: We included patients at risk of developing severe COVID-19, all of whom exhibited mild to moderate symptoms and were admitted to three hospital centers. Patients were divided into two groups: one received Paxlovid alongside standard care, while the other was given only standard care. We compared clinical characteristics, hospital stay duration, and clinical outcomes between two groups. Multi-factor analysis determined the independent risk factors influencing the duration of hospitalization and disease progression. RESULTS: In the study, those treated with Paxlovid shorter hospital stays than those in the control group (p < 0.001). Multivariate analysis indicated that the absence of Paxlovid treatment was a distinct risk factor for hospitalizations lasting over 7 days (OR: 4.983, 95% CI: 3.828-6.486, p < 0.001) and 14 days (OR: 2.940, 95% CI: 2.402-3.597, p < 0.001). CONCLUSION: Amid the Omicron outbreak, Paxlovid has proven to be a safe and effective treatment for reducing hospitalization durations for patients with mild to moderate COVID-19.

Novel clinical radiomic nomogram method for differentiating malignant from non-malignant pleural effusions.

Objectives: To establish a clinical radiomics nomogram that differentiates malignant and non-malignant pleural effusions. Methods: A total of 146 patients with malignant pleural effusion (MPE) and 93 patients with non-MPE (NMPE) were included. The ROI image features of chest lesions were extracted using CT. Univariate analysis was performed, and least absolute shrinkage selection operator and multivariate logistic analysis were used to screen radiomics features and calculate the radiomics score. A nomogram was constructed by combining clinical factors with radiomics scores. ROC curve and decision curve analysis (DCA) were used to evaluate the prediction effect. Results: After screening, 19 radiomics features and 2 clinical factors were selected as optimal predictors to establish a combined model and construct a nomogram. The AUC of the combined model was 0.968 (95% confidence interval [CI] = 0.944-0.986) in the training cohort and 0.873 (95% CI = 0.796-0.940) in the validation cohort. The AUC value of the combined model was significantly higher than those of the clinical and radiomics models (0.968 vs. 0.874 vs. 0.878, respectively). This was similar in the validation cohort (0.873, 0.764, and 0.808, respectively). DCA confirmed the clinical utility of the radiomics nomogram. Conclusion: CT-based radiomics showed better diagnostic accuracy and model fit than clinical and radiological features in distinguishing MPE from NMPE. The combination of both achieved better diagnostic performance. These findings support the clinical application of the nomogram in diagnosing MPE using chest CT.

Hypoxia-induced long non-coding RNA plasmacytoma variant translocation 1 upregulation aggravates pulmonary arterial smooth muscle cell proliferation by regulating autophagy via miR-186/Srf/Ctgf and miR-26b/Ctgf signaling pathways.

BACKGROUND: The lncRNA PVT1 reportedly functions as a competing endogenous RNA (ceRNA) of miR-186 and miR-26b in different tissue types. In this study, we investigated the possible involvement of the miR-186/Srf/Ctgf and miR-26b/Ctgf signaling pathways in the pathogenesis of hypoxia-induced PAH. METHODS: Expression of PVT1, miR-186, miR-26b, and Srf and Ctgf mRNAs were evaluated by real-time polymerase chain reaction. Protein expression of SRF, CTGF, LC3B-I, LC3B-II, and Beclin-I was evaluated using western blotting. The regulatory relationship between the lncRNA, miRNAs, and target mRNAs was explored using luciferase assays. Immunohistochemistry was used to evaluate the expression of SRF and CTGF in situ. MTT assay was performed to assess the proliferation of PASMCs. RESULTS: Exposure to hypoxia markedly altered the expression of PVT1, Srf, Ctgf, miR-186, and miR-26b in a rat model. MiR-186 binding sites in the sequences of Srf mRNA and PVT1 were confirmed by luciferase assays, indicating that miR-186 may interact with both PVT1 and Srf mRNA. Additionally, miR-26b binding sites were identified in the sequences of Ctgf mRNA and PVT1, suggesting that miR-26b may interact with both PVT1 and Ctgf mRNA. In line with this, we found that overexpression of PVT1 reduced expression of miR-26b and miR-186 but activated expression of Srf, Ctgf, LC3B-II, and Beclin-I. CONCLUSIONS: Upregulation of PVT1 by exposure to hypoxia promoted the expression of CTGF, leading to deregulation of autophagy and abnormal proliferation of PASMCs. Dysregulation of the miR-186/Srf/Ctgf and miR-26b/Ctgf signaling pathways may be involved in the pathogenesis of hypoxia-induced PASMCs.

An optical platform for cell tracking in adult zebrafish.

Adult zebrafish are being increasingly used as a model in cancer and stem cell research. Here we describe an integrated optical system that combines a laser scanning confocal microscope (LSCM) and an in vivo flow cytometer (IVFC) for simultaneous visualization and cell quantification. The system is set up specifically for non-invasive tracking of both stationary and circulating cells in adult zebrafish (casper) that have been engineered to be optically transparent. Confocal imaging in this instrument serves the dual purpose of visualizing fish tissue microstructure and an imaging-based guide to locate a suitable vessel for quantitative analysis of circulating cells by IVFC. We demonstrate initial testing of this novel instrument by imaging the transparent adult zebrafish casper vasculature and tracking circulating cells in CD41-GFP/Gata1-DsRed transgenic fish whose thrombocytes/erythrocytes express the green and red fluorescent proteins. In vivo measurements allow cells to be tracked under physiological conditions in the same fish over time, without drawing blood samples or sacrificing animals. We also discuss the potential applications of this instrument in biomedical research.

Mortality and risk factors for COVID-19 in hemodialysis patients: A systematic review and meta-analysis.

Introduction: The present study systematically reviewed the clinical features and risk factors in patients undergoing maintenance hemodialysis (MHD) who also acquired coronavirus disease 2019 (COVID-19). More specifically, clinical manifestations, prognosis, and risk factors for death among this population were explored. Method: A literature search using the PubMed, Web of Science, and Embase databases, for articles involving patients with laboratory-confirmed COVID-19 and undergoing MHD published between January 1, 2020, and March 13, 2022, was performed. Random-effects meta-analyses were performed to calculate the weighted mean prevalence and corresponding 95% confidence interval (CI) or weighted means and 95% CI. Heterogeneity among studies was assessed using I2 statistics. Results: Twenty-two studies including 13,191 patients with COVID-19 undergoing MHD were selected. The most common symptoms included fever (53% [95% CI 41%-65%]) and cough (54% [95% CI 48%-60%]); however, 17% (95% CI 11%-22%) of the cases were asymptomatic. In subgroup analysis, the proportion of male patients (65% [95% CI 58%-71%]), and patients with coronary artery disease (30% [95% CI 17%-44%) and chronic obstructive pulmonary disease (9% [95% CI 4%-15%]) was greater in the non-survivor group compared with the survivor group. Furthermore, patients undergoing MHD, who were also positive for COVID-19, exhibited a high mortality rate (24% [95% CI 19%-28%]). Conclusions: MHD patients with COVID-19 may initially present as asymptomatic or with mild symptoms; nevertheless, in this study, these patients exhibited a higher risk for death compared with COVID-19 patients not undergoing MHD. Moreover, male sex and underlying cardiovascular and respiratory diseases increased the mortality risk.