ScRNA-seq of Anopheles gambiae hemocytes after blood-feeding, Plasmodium berghei infection, or sugar feeding
ABSTRACT: Anopheline mosquitoes transmit Plasmodium parasites to humans, and are responsible for an estimated 219 million cases of malaria, leading to over 400,000 deaths annually. The mosquito’s immune system limits Plasmodium infection in several ways, and hemocytes, the insect white blood cells, are key players in these defense responses. However, the full functional diversity of mosquito hemocytes and their developmental trajectories have not been established. We use single cell RNA sequencing (scRNA-seq) to analyze the transcriptional profiles of individual mosquito hemocytes in response to blood feeding or infection with Plasmodium. Circulating hemocytes were collected from adult A. gambiae M form (A. coluzzii) females that were either kept on a sugar meal or fed on a healthy or a Plasmodium berghei-infected mouse. Transcriptomes from 5,383 cells (collected 1, 3, and 7 days after feeding) revealed nine major cell clusters.
Project description:We aimed at characterizing the diverse hemocyte populations present in the hemolymph of the Drosophila larvae. The hemocytes were collected from wandering larvae infested by wasp (WI) or not infested (NI). The hemocytes were then sequenced using 10x genomics technology.
Project description:Plasmodium-specific CD4+ T cells from mice infected with Plasmodium chabaudi chabaudi AS parasites were recovered at Days 0, 7, and 28 to undergo processing and generate scRNA-seq dataset. At Day 28, mice were administered with either saline or artesunate (intermittent artesunate therapy - IAT). scRNA-seq dataset was analysed to investigate transcriptome dynamics of CD4+ T cells from effector to memory states.
Project description:Anopheline mosquitoes transmit Plasmodium parasites to humans, and are responsible for an estimated 219 million cases of malaria, leading to over 400,000 deaths annually. The mosquito’s immune system limits Plasmodium infection in several ways, and hemocytes, the insect white blood cells, are key players in these defense responses. However, the full functional diversity of mosquito hemocytes and their developmental trajectories have not been established. We use bulk RNA sequencing (scRNA-seq) to analyze the transcriptional profiles of hemocytes, of guts, and of carcasses of mosquito hemocytes in response to blood feeding or infection with Plasmodium. Data from three independent biological replicates for each condition and time-point (day 0, 1, 2, 3, and 7 after sugar-feeding, blood-feeding or P. berghei infection).
Project description:Spermatogenesis in mammals is a complex and highly orchestrated process, which involves the differentiation of diploid (2n-DNA content) spermatogonia into haploid (n) sperm. This process begins with spermatogonia, a niche of stem cells, which drive this process. Spermatogonial stem cells (SSCs) undergo mitotic self-renewal to maintain this niche, while sub-populations of spermatogonia progressively undergo differentiation and later gain competence to initiate meiosis and form sperm. SSCs undergo extensive chromatin remodelling and major morphological changes, while maintaining genome integrity and parental imprints. To study this in greater detail we performed single-cell RNA sequencing of spermatogenic cells derived from the testis of the non-human primate Macaca fascicularis.
Project description:Background: The mosquito Anopheles gambiae is a major vector of human malaria. Increasing evidence indicates that blood cells (hemocytes) comprise an essential arm of the mosquito innate immune response against both bacteria and malaria parasites. To further characterize the role of hemocytes in mosquito immunity, we undertook the first genome-wide transcriptomic analyses of adult female An. gambiae hemocytes following infection by two species of bacteria and a malaria parasite. Results: We identified 4047 genes expressed in hemocytes, using An. gambiae genome-wide microarrays. While 279 transcripts were significantly enriched in hemocytes relative to whole adult female mosquitoes, 959 transcripts exhibited immune challenge-related regulation. The global transcriptomic responses of hemocytes to challenge with different species of bacteria and/or different stages of malaria parasite infection revealed discrete, minimally overlapping, pathogen-specific signatures of infection-responsive gene expression; 105 of these represented putative immunity-related genes including anti-Plasmodium factors. Of particular interest was the specific co-regulation of various members of the Imd and JNK immune signaling pathways during malaria parasite invasion of the mosquito midgut epithelium. Conclusion: Our genome-wide transcriptomic analysis of adult mosquito hemocytes reveals pathogen-specific signatures of gene regulation and identifies several novel candidate genes for future functional studies. In order to identify hemocyte-specific and immune-responsive transcripts, we first compared transcripts expressed in hemocytes from one day old sugar-fed mosquitoes to transcripts detected in whole mosquitoes of the same age and feeding status. This resulted in identification of the hemocyte-enriched transcriptome. We then compared hemocytes from 1 day old mosquitoes, 1 hour after immune challenge with heat-killed Escherichia coli or Micrococcus luteus, to control female mosquitoes injected with sterile PBS to determine the bacteria challenge responsive transcriptomes. We used heat-killed bacteria in these assays, because our primary interest was in identifying the bacterial responsive transcriptome and to avoid the potentially confounding effects of altered gene expression due to the lethal effects of a systemic infection associated with injection of living bacteria. Lastly, we compared hemocytes from mosquitoes at 24 hours and 19 days after ingestion of a blood meal infected with Plasmodium berghei to mosquitoes of the same age fed a non-infected blood meal to determine the ookinete and sporozoite infection responsive transcriptomes, respectively. This design resulted in a total of five experimental treatments. The following samples are not included in this submission: Hemo E coli vs. hemo unchallenged A Hemo E coli vs. hemo unchallenged B Hemo m luteus vs. hemo unchallenged A Hemo m luteus vs. hemo unchallenged B
Project description:Here, we performed single cell RNA sequencing (scRNA-seq) of human fetal duodenal tissue samples from 8 individual biological specimens across 7-21 weeks of gestation. The data set is composed of over 37,000 cells (between 3,000-9,000 cells per time point) from diverse intestinal lineages. Lineages captured include epithelium, mesenchyme, immune, neurons, and endothelium. These data were used to specifically interrogate the development and emergence of mesenchymal lineages during human development.
Project description:Underdeveloped lungs are a primary cause of morbidity and mortality in premature infants, but our ability to help these patients by speeding up lung development are hindered by a lack of understanding of human lung developmental biology. Here, we performed single cell RNA sequencing of the human fetal lung from samples spanning from 11.5 weeks gestation to 21 weeks gestation from the distal lung, middle airways, and the tracheal epithelium. The primary goal of this experiment was to define fetal cell states to serve as a gold standard for pluripotent stem cell-derived lung cells and tissues, and to identify potential signaling pathways that drive differentiation of lung progenitor cells to mature cell types. Additionally, we generated bud tip progenitor organoids from 12 week human fetal lung bud tip progenitors. We show that treatment of bud tip progenitor organoids with a short pulse of dual SMAD activation (BMP4+TGFb1) led to the upregulation of lung basal cell markers, a cell type that serves as a critical stem cell for the adult airway, and that further treatment with dual SMAD inhibition leads to the generation of airway-like organoids containing differentiated cell types of the adult airway, including basal stem cells.
Project description:The chemokines CXCL13 and CXCL12 are reported to be important for the germinal center reaction. Since CXCL12-deficient mice are embryonically lethal, here we took advantage of the Cxcl13-Cre/TdTomato mouse models to genetically ablate CXCL12 from B cell-interacting reticular cells and examine the molecular consequence on germinal center B cells. Spatial segregation of follicular dendritic cells, germinal center B cells and follicular helper T cells is impaired in Cxcl13-Cre/TdTomato Cxcl12fl/fl mice. Single cell transcriptomic analysis revealed that all germinal center B cell subsets (corresponding to distinct stages of the germinal center response) are present in draining lymph nodes of immunized CXCL12-conditionally deficient mice. While most transcriptional regulators of the germinal center response are unperturbed by the genetic perturbation of CXCL12, Bach2 levels were elevated in germinal center B cells from lymph nodes of Cxcl12fl/fl mice. Moreover, single cell B cell receptor sequencing revealed that germinal center B cells in Cxcl13-Cre/TdTomato Cxcl12fl/fl mice harbour a lower mutational burden when compared to germinal center B cells isolated from immunized control mice. Gene expression profiles were validated by flow cytometry and suggest that the provision of CXCL12 by reticular cells governs efficient germinal center responses.
Project description:In dogs, a species for which markers of cell populations are often limiting, we sought to evaluate in an unbiased way the heterogeneity of cell subpopulations in the bronchoalveolar lavage fluid of healthy dogs, by single-cell RNA-sequencing.