RNA-seq analysis of pulmonary arterial smooth muscle cells(PASMCs)stimulated with BMP9/10
Ontology highlight
ABSTRACT: Inactivation of BMP9/10 in mice leads to attenuated contractility of smooth muscle cells and decreased blood pressure.Treatment of de-differentiated PASMCs with BMP9/10 resulted in a strong increase of ACTA2 expression as measured by immunofluorescence and RT-PCR, indicating a switch from the de-differentiated, synthetic to a differentiated, contractile state. To further study the role of BMP9/10 in smooth muscle cells, we isolate PSAMCs and stimulate cells with or without BMP9/10 and perform the RNA-seq analysis.
Project description:Aorta was isolated from ANF-Cre positive/BMP10 loxP/loxP/BMP9-/- (BMP9/10 dKO) mice. ANF-Cre negative BMP10 loxP/loxP mice were used as controls. Transcritional profiling was performed to understand the impact of BMP9/10 expression on aorta function.
Project description:The liver is a largest solid organ in the body and is majorly composed of HCs, ECs, KCs, and HSCs, which spatially interact and cooperate each other to maintain liver homeostasis. However, the complexity and molecular mechanisms underlying the crosstalk between these different cell types remain to be revealed. Here, we generated mice with conditional deletion of Bmp9/10 in different liver cell types and demonstrated that HSCs was the major source of BMP9 and BMP10 in the liver. Using transgenic ALK1 (receptor for BMP9/10) reporter mice, we found that ALK1 is expressed on KCs and ECs other than HCs and HSCs. KCs from Bmp9/10HSC-KO (conditional deletion of Bmp9/10 from HSCs) mice lost their signature genes expression, such as ID1/3, CLEC4F, VSIG4 and CLEC2, and were replaced by monocyte-derived macrophages. ECs from Bmp9/10HSC-KO mice also lost their identity and were transdifferentiated to continuous ECs, ultimately leading to collagen IV deposition and liver fibrosis. Hepatic ECs expressed several angiocrine factors, such as BMP2, BMP6, Wnt2 and Rspo3, to regulate liver iron metabolism and metabolic zonation. We found that these angiocrine factors were significantly decreased in ECs from Bmp9/10HSC-KO mice, which further resulted in liver iron overload and disruption of HCs zonation. In addition, focal fatty liver spontaneously occurred in Bmp9/10HSC-KO mice at the age of 28W. In summary, we demonstrated that HSCs play a central role in mediating liver cell-cell crosstalk via production of BMP9/10 to maintain liver health.
Project description:Aorta was isolated from ANF-Cre positive / BMP10 loxP/loxP / BMP9-/- (BMP9/10 dKO) mice. ANF-Cre negative BMP10 loxP/loxP mice were used as controls. Transcriptional profiling was performed to understand the importance of BMP9/10 expression on aorta function.
Project description:This experiment was desinged to investigate the difference in global gene expression induced by the circulating form of BMP9, pro-BMP9, and BMP9 in complex with soluble endoglin (sENG:BMP9).
Project description:As the critical step of pathogenesis during hypoxic pulmonary arterial hypertension (PAH) vascular remodeling is closely associated with pulmonary arterial smooth muscle cell (PASMC) alterations induced by hypoxia that include persistent vasoconstriction, abnormal proliferation and PASMC resistance to apoptosis. Quercetin is a flavonoid compound extracted from green plants that inhibits proliferation, induces apoptosis, arrests the cell cycle, and rescues the constriction of PASMCs, but the underlying mechanisms remain poorly understood. In this study, we used a commercial Agilent Whole Rat Genome Oligo Microarray to determine the overall transcriptional response of PASMCs in response to exposure to hypoxia and the optimal concentration of quercetin. Hypoxia induced the upregulation of 1694 genes and the downregulation of 2091 genes compared with the normoxia group. Quercetin treatment resulted in 1790 upregulated genes and 1450 downregulated genes. Quercetin is known to cause differential expression of several of these genes that are known to promote proliferation, induce apoptosis (Cycs, Ppp3ca, Prkar2b, Akt3, Ppp3cc, Il1rap, Ntrk1), arrest the cell cycle (Chek2, Cdkn1c, Gadd45b, Stag2, Anapc7, Orc1, Ccne1, Myc3, Skp1, Espl1, Cdc45, Mcm4), and rescue PASMC constriction (Ramp1, Ramp3, Adcy5, Gnas, Prkcd, Itpr3, Adra1d, Calm1, Npr1, Avpr1a, Ednra, Adcy8). Real-time quantitative RT-PCR was performed to verify the microarray results. In conclusion, quercetin altered the expression profile of many genes regulated by hypoxia in PASMCs, which helps to further explore the mechanism of the effects of quercetin treatment on hypoxic PAH.
Project description:This experiment was performed in order to investigate the difference in the gene expression pattern between differentiated airway/vascular smooth muscle cells and progenitors for alveolar myofibroblasts in embryonic lung. To answer this question, we genetically labeled Smooth muscle actin-expressing cells using Sma-Cre-ERT2 crossed with Tomato Cre-reporter line. This line allows to label both airway/vascular smooth muscle cells (high SMA expression) and progenitors for alveolar myofibroblasts (low SMA expression). Mice were treated with tamoxifen at embryonic day E15.5 and cells from embryonic lungs were collected at E18.5. Mice used in this study were offspring from RjOrl:SWISS (CD1) [mother] x STOCK Tg (Acta2-cre/ERT2)12Pcn Gt(ROSA)26Sortm9(CAG-tdTomato)Hze/sbel (Acta2-CreERT2; tdTomatoflox) [father]
Project description:Smooth muscle cells exist in many different locations within the body, including blood vessels and airways. The heterogeneity of smooth muscle cells has been related to their embryological origins and could have implications in many diseases, including atherosclerosis, pulmonary hypertension, and asthma. Here we aimed to study the heterogeneity of Acta2+ cells in the mouse lung and to identify specific markers for Acta2+ sub-populations. We used a mouse line containing an Acta2hrGFP/Cspg4dsRed reporter and performed single cell RNA sequencing of Acta2+ cells isolated from adult mouse lungs. We identified 3 main distinct clusters, corresponding to airway smooth muscle (ASM), vascular smooth muscle (VSM) and mesenchymal alveolar niche cells (MANCs), and extracted specific gene signatures for each of these cell types. Our dataset and gene signatures help address the heterogeneity of Acta2+ lineages in the lung and will be of use for future studies focusing on smooth muscle cells in the lung and in other organs.