{"database":"biostudies-arrayexpress","file_versions":[],"scores":null,"additional":{"omics_type":["Metabolomics","Unknown","Transcriptomics","Genomics","Proteomics"],"submitter":["Charles Girardot"],"instrument_platform":["NextSeq 2000"],"study_type":["RNA-seq of coding RNA from single cells"],"organism":["Homo sapiens"],"species":["Homo sapiens"],"full_dataset_link":["https://www.ebi.ac.uk/biostudies/studies/E-MTAB-14463"],"description":["This dataset consists of two individual sample-multiplexing (MULTI-seq) single-cell RNA sequencing experiments, MB10x01 and MB10x02. Single-cell RNA sequencing (10X Genomics) analyses were performed on a microfluidic 3D in vitro blood-brain-barrier model (containing primary human brain microvascular endothelial cells, brain vascular pericytes, and astrocytes) perfused with P. falciparum egress product (MB10x01) or P. falciparum-infected red blood cells (RBC) (MB10x02). Dataset MB10x01 included two samples multiplexed by MULTI-seq sample barcoding (TCCTCGAA for control RBC lysate, ATGCGATG for P. falciparum egress product). P. falciparum egress product was obtained by letting tightly synchronized P. falciparum-infected RBC egress in media used for perfusions (5x10^7 infected RBC/ml). 3D blood-brain-barrier models perfused with P. falciparum egress products were incubated for 24 hours and compared to a control perfused with uninfected red blood cell lysate. MULTI-seq barcoding (McGinnis et al.Ê2019) was used for sample-barcoding of these two conditions, and the dataset contains cDNA (transcriptome) and sample barcode read files. Dataset MB10x02 included three samples multiplexed by MULTI-seq sample barcoding (GCTATGCA for control RBC, CGATACTG for Trophozoite stage, TACGCAGT for Schizont stage). 3D blood-brain-barrier models were perfused for 30 minutes with P. falciparum-infected RBC in the Trophozoite stage (26-34 hours post invasion) or Schizont stage (42-48 hours post invasion) (5x10^7 infected RBC/ml). After a 20-minute wash, the 3D blood-brain-barrier models were incubated with the bound P. falciparum-infected RBC for 6 hours and compared to uninfected RBC perfused controls. MULTI-seq barcoding was used for sample-barcoding of the three conditions, and the dataset contains cDNA (transcriptome) and sample barcode read files."],"repository":["biostudies-arrayexpress"],"sample_protocol":["Growth Protocol - To obtain the P. falciparum egress media, P. falciparum-iRBC at 42-48 hours post-infection (hpi) were purified by gelaspan (40 mg/mL) and egress was inhibited for 5 hours using the reversible PKG inhibitor C2 (kindly donated by Michael Blackman, The Francis Crick Institute). After drug removal, parasites were resuspended in EGM-2MV media supplemented with 1% astrocyte and pericyte growth factors (ScienCell), gassed at a 5% O2 concentration and left in the incubator overnight on a shaker (50 rpm) to facilitate parasite egress. The concentration was adjusted at 5x10^7 ruptured P. falciparum-iRBC/mL before the suspension was centrifuged at 1000 rpm, aliquoted and flash frozen in liquid nitrogen. The same protocol was applied to uninfected erythrocytes to be used as a negative control medium. After 6 days in culture, 3D-BBB devices were perfused with 150 µL of P. falciparum-iRBC egress media or RBC-media by gravity-driven flow for 30 minutes, followed by a 20-minute wash. Control and P. falciparum stimulated microvessels were incubated for 24 hours at 37C. Reservoirs were refilled once after 12 hours. After incubation, microvessels were dissociated for scRNAseq.","Sequencing - paired-end sequencing on illumina NextSeq 2000","Library Construction - After obtaining the samples from the 10x Chromium Controller the samples were processed and the libraries were constructed with Chromium Next GEM Single Cell 3' - Reagent Kits v3.1 (10x Genomics) and single-indexed for sequencing according to the manufacturer's protocol.","Growth Protocol - P. falciparum-iRBC late-stage parasites at the desired stage of the life cycle (26-34 hpi for trophozoites or 42-48 hpi for schizonts) were purified by magnetic separation (MACS) to above 80% enrichment. After 6 days in culture, 3D-BBB devices were perfused with 150 µL of P. falciparum-iRBC at 5x10^7 iRBC/mL concentration under gravity-driven flow for 30 minutes followed by a 20-minute wash to remove unbound cells. As a control, the same volume and concentration of uninfected erythrocytes was used for perfusion. Devices were then incubated for 6 hours at 37C. After incubation, microvessels were dissociated for scRNAseq.","Growth Protocol - Primary human brain astrocytes (HA) (ScienCell) and brain vascular pericytes (ScienCell) were added to the 7.5 mg/mL collagen solution in a 7:3 ratio, using a concentration of 7.5x10^5 HA/mL(collagen) : 3.2x10^5 HBVP/mL(collagen). A multi-step process combining soft lithography and injection molding was used to imprint a microfluidic network in the collagen bulk, as previously reported (Zheng et al. 2012; Piatti et al. 2022). Primary human brain microvascular endothelial cells (HBMEC) (CellSystems) were seeded into the microfluidic network at a concentration of 7x10^6 cells/mL under gravity-driven flow until full coverage of the microchannels was achieved. 3D-BBB devices were cultured for 6 days and EGM-2MV medium supplemented with 1% astrocyte and pericyte growth factors (ScienCell) was changed every 12 hours by gravity-driven flow, as previously described (Bernabeu et al. 2019)."],"figure_sub":["Organization","MINSEQE Score","Assays and Data","MAGE-TAB Files"],"pubmed_authors":["Alina Batzilla","Charles Girardot","Maria Bernabeu"],"additional_accession":[]},"is_claimable":false,"name":"Single-cell RNA sequencing of a 3D blood-brain-barrier model perfused with P. falciparum-infected red blood cells and egress products","description":"This dataset consists of two individual sample-multiplexing (MULTI-seq) single-cell RNA sequencing experiments, MB10x01 and MB10x02. Single-cell RNA sequencing (10X Genomics) analyses were performed on a microfluidic 3D in vitro blood-brain-barrier model (containing primary human brain microvascular endothelial cells, brain vascular pericytes, and astrocytes) perfused with P. falciparum egress product (MB10x01) or P. falciparum-infected red blood cells (RBC) (MB10x02). Dataset MB10x01 included two samples multiplexed by MULTI-seq sample barcoding (TCCTCGAA for control RBC lysate, ATGCGATG for P. falciparum egress product). P. falciparum egress product was obtained by letting tightly synchronized P. falciparum-infected RBC egress in media used for perfusions (5x10^7 infected RBC/ml). 3D blood-brain-barrier models perfused with P. falciparum egress products were incubated for 24 hours and compared to a control perfused with uninfected red blood cell lysate. MULTI-seq barcoding (McGinnis et al.Ê2019) was used for sample-barcoding of these two conditions, and the dataset contains cDNA (transcriptome) and sample barcode read files. Dataset MB10x02 included three samples multiplexed by MULTI-seq sample barcoding (GCTATGCA for control RBC, CGATACTG for Trophozoite stage, TACGCAGT for Schizont stage). 3D blood-brain-barrier models were perfused for 30 minutes with P. falciparum-infected RBC in the Trophozoite stage (26-34 hours post invasion) or Schizont stage (42-48 hours post invasion) (5x10^7 infected RBC/ml). After a 20-minute wash, the 3D blood-brain-barrier models were incubated with the bound P. falciparum-infected RBC for 6 hours and compared to uninfected RBC perfused controls. MULTI-seq barcoding was used for sample-barcoding of the three conditions, and the dataset contains cDNA (transcriptome) and sample barcode read files.","dates":{"release":"2025-06-25T00:00:00Z","modification":"2025-07-08T14:01:41.294Z","creation":"2024-09-19T09:53:59.575Z"},"accession":"E-MTAB-14463","cross_references":{"ENA":["ERP164332"],"EFO":["EFO_0004170","EFO_0005684","EFO_0005518","EFO_0004184"]}}