scholarly journals Isolation of Primary Mouse Lung Endothelial Cells

10.3791/63253 ◽  
2021 ◽  
Author(s):  
Erika Wong ◽  
Nina Nguyen ◽  
Judith Hellman
Keyword(s):  
Endothelial Cells ◽  
Mouse Lung ◽  
Primary Mouse ◽  
Lung Endothelial Cells

Abstract 436: Mfn2 is a Nodal Point Regulatory Step That Coordinates Angiogenic Phenotype in the Vascular Endothelium

2016 ◽  
Vol 119 (suppl_1) ◽  
Author(s):  
Gladys A Nchaw ◽  
Kyriakos N Papanicolaou ◽  
Jennifer L Parker-Duffen ◽  
Ramon G Bonegio ◽  
Kenneth Walsh
Keyword(s):  
Endothelial Cells ◽  
Mitochondrial Dynamics ◽  
Nodal Point ◽  
Mouse Lung ◽  
Published Data ◽  
Dependent Manner ◽  
Wild Type ◽  
Primary Mouse ◽  
Enos Phosphorylation ◽  
Sprout Formation

Recent published data suggests that endothelial metabolism regulates angiogenesis. The key regulators of mitochondrial dynamics, mitofusins (Mfn1 and Mfn2) have been linked to cellular metabolic profile in tissues with high energetic requirements but their role in endothelial cells, were mitochondria function as signaling organelles is unknown. Here, we found that Mfn2 modulates postnatal neovascularization. Endothelial deletion of Mfn2 (Mfn2 ecko ) impaired, whereas, endothelial Mfn1 deletion (Mfn1 ecko ) did not affect clinical recovery, limb blood flow reperfusion, and capillary density compared with wild-type mice. These effects were associated with reduced eNOS activation in the ischemic leg of Mfn2 ecko mice compared with wild-type mice. Transplantation of bone marrow cells from WT to Mfn2ecko mice did not significantly improve blood perfusion in the ischemic limb of Mfn2 ecko mice. In agreement with these results, salient features of angiogenesis such as eNOS phosphorylation, cell migration, proliferation and capillary sprout formation were significantly inhibited in Mfn2 null primary mouse lung endothelial cells (Mfn2 -/- MLECs), whereas ablation of Mfn1 did not affect angiogenic signaling following VEGF stimulation. Because Mfn2 modulates the unfolded protein response (UPR) and the UPR machinery has been shown to generate angiomodulatory signals, we ascertained the effects of Mfn2 loss on UPR-mediated VEGF angiogenic signal transduction as potential mechanism. Mfn2 ablation sensitized Mfn2 -/- MLECs to proapototic UPR activation following VEGF treatment according to CHOP levels. Augmented CHOP levels corresponded with reduced eNOS phosphorylation. Inhibition of the UPR PERK signaling axis with PERK siRNA or reconstitution of Mfn2 (AdMfn2) reduced CHOP levels and significantly improved eNOS activation, EC proliferation and migration in Mfn2 -/- MLECs. Moreover, PERK siRNA or reconstitution of Mfn2 rescued the defect in capillary sprout formation in Mfn2 ecko mice. These data suggest that Mfn2 regulate angiogenic signaling in a UPR PERK axis-dependent manner and extend the role of Mfn2 beyond mitochondrial dynamics, metabolism and ER stress to angiogenesis and reparative vascular remodeling.

scholarly journals A simple protocol for isolating mouse lung endothelial cells

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Jinping Wang ◽  
Niu Niu ◽  
Suowen Xu ◽  
Zheng Gen Jin
Keyword(s):  
Endothelial Cells ◽  
Mouse Lung ◽  
Simple Protocol ◽  
Lung Endothelial Cells

HISTONE ACETYLATION MODULATES DNA DAMAGE IN BACTERIAL ENDOTOXIN-TREATED MOUSE LUNG ENDOTHELIAL CELLS

2004 ◽  
Vol 13 (3) ◽  
pp. 150
Author(s):  
Jane Rose ◽  
Scott Wray ◽  
Hong Huang ◽  
Rostislav Likhotvorik ◽  
Dale Hoyt
Keyword(s):  
Dna Damage ◽  
Endothelial Cells ◽  
Histone Acetylation ◽  
Treated Mouse ◽  
Bacterial Endotoxin ◽  
Mouse Lung ◽  
Lung Endothelial Cells

scholarly journals Targeted Delivery of Oligodeoxynucleotides to Mouse Lung Endothelial Cells in Vitro and in Vivo

2005 ◽  
Vol 12 (3) ◽  
pp. 510-518 ◽  
Author(s):  
Annette Wilson ◽  
Wen Zhou ◽  
Hunter C. Champion ◽  
Sean Alber ◽  
Zhi-Lue Tang ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Targeted Delivery ◽  
Mouse Lung ◽  
Lung Endothelial Cells

scholarly journals Intermittent Hypoxia Activates Duration-Dependent Protective and Injurious Mechanisms in Mouse Lung Endothelial Cells

2018 ◽  
Vol 9 ◽  
Author(s):  
Peter Wohlrab ◽  
Lourdes Soto-Gonzales ◽  
Thomas Benesch ◽  
Max Paul Winter ◽  
Irene Marthe Lang ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Intermittent Hypoxia ◽  
Mouse Lung ◽  
Lung Endothelial Cells

scholarly journals Diminished metastasis in tetraspanin CD151–knockout mice

Blood ◽  
2011 ◽  
Vol 118 (2) ◽  
pp. 464-472 ◽  
Author(s):  
Yoshito Takeda ◽  
Qinglin Li ◽  
Alexander R. Kazarov ◽  
Mathieu Epardaud ◽  
Kutlu Elpek ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Endothelial Cells ◽  
Endothelial Cell ◽  
Cell Activation ◽  
Immune Cell ◽  
Critical Role ◽  
Mouse Lung ◽  
Null Mouse ◽  
Host Animal ◽  
Lung Endothelial Cells

Abstract Tetraspanin protein CD151 on tumor cells supports invasion and metastasis. In the present study, we show that host animal CD151 also plays a critical role. CD151-null mice showed markedly diminished experimental lung metastasis after injection of Lewis lung carcinoma or B16F10 melanoma cells. Diminished tumor cell residence in the lungs was evident 6-24 hours after injection. Consistent with an endothelial cell deficiency, isolated CD151-null mouse lung endothelial cells showed diminished support for B16F10 adhesion and transendothelial migration, diminished B16F10-induced permeability, and diminished B16F10 adhesion to extracellular matrix deposited by CD151-null mouse lung endothelial cells. However, CD151 deletion did not affect the size of metastatic foci or subcutaneous primary B16F10 tumors, tumor aggregation, tumor clearance from the blood, or tumor-induced immune cell activation and recruitment. Therefore, the effects of host CD151 on metastasis do not involve altered local tumor growth or immune surveillance. VEGF-induced endothelial cell signaling through Src and Akt was diminished in CD151-null endothelial cells. However, deficient signaling was not accompanied by reduced endothelial permeability either in vitro (monolayer permeability assay) or in vivo (VEGF-stimulated Miles assay). In summary, diminished metastasis in CD151-null host animals may be due to impaired tumor-endothelial interactions, with underlying defects in mouse lung endothelial cell extracellular matrix production.

Pulmonary Irradiation Fibrosis Is Preceded By Increased Endothelial Cell Gene Expression

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 5569-5569
Author(s):  
Ronny Kalash ◽  
Jeremiah Au ◽  
Michael W. Epperly ◽  
Julie P. Goff ◽  
Tracy M. Dixon ◽  
...  
Keyword(s):  
Gene Expression ◽  
Endothelial Cells ◽  
Endothelial Cell ◽  
Conflicts Of Interest ◽  
The Body ◽  
Mouse Lung ◽  
Alveolar Cells ◽  
Cell Gene Expression ◽  
Lung Endothelial Cells ◽  
Endothelial Genes

Abstract Irradiation of the C57BL/6NHsd mouse lung results in pulmonary fibrosis. An acute phase (days 1 to 28) following irradiation recognized by an inflammatory response is followed by a latent phase (day 28 to day 125) where lung appears normal. Fibrosis occurs from day 125 till death characterized by bone marrow stromal cell migration to and proliferation in lung and increased collagen deposition. To investigate the role of endothelial cells, C57BL/6NHsd female mice were irradiated to 20 Gy to the pulmonary cavity shielding the rest of the body. Lung was excised at serial times, endothelial cells separated and tested for expression of endothelial genes vWF, VEGF, FGF1, CCL13 and CTGF; inflammatory genes IL-6; IGFbp7 and genes associated with fibrosis including MnSOD, Nrf2, NfkB, TLR4 and TGF-B. Endothelial cells were separated by using antibodies to CD45, or PECAM and sorting using flow cytometry. RNA was extracted using Triazol and gene expression determined using Real Time Polymerase Chain Reaction (RT-PCR). In irradiated total lung there was significant increase in endothelial cell markers at all times, compared to nonirradiated lung. Transcripts for vWF increased by 474.1 ± 388.5% (p = 0.017) by 2 days after irradiation, 520 ± 55.4% (p < 0.0001) at day 50, and 484.8 ± 26.8% (p < 0.0001) by day 200. VEGF and CTGF also remained elevated. In contrast, total lung MnSOD gene expression increased by 140.6 ± 46.1% at day 2 (p = 0.0185), decreased at day 50 to 17.9 ± 11.2% (p = 0.2808), and increased to 31.5 ± 11.4% by day 200 (p = 0.023). However, purified endothelial cells compared to alveolar cells, showed increased expression of not only the endothelial cell genes but also MnSOD, Nrf2, NfkB, and TGF-B. MnSOD expression in endothelial compared to alveolar cells at day 28 after irradiation was higher at 244.5 ± 7.8 vs 80.5 ± 4.9 (p = 0.0016), at day 150 expression was 140.1 ± 25.0 vs 5.4 ± 1.2 (p < 0.0001) and day 200 expression was 368.5 ± 6.4 vs 51.8 ± 10.3 (p = 0.0007). Lung endothelial cells display critical biomarkers of irradiation injury leading to fibrosis. Disclosures: No relevant conflicts of interest to declare.

scholarly journals MKK3 regulates mitochondrial biogenesis and mitophagy in sepsis-induced lung injury

2014 ◽  
Vol 306 (7) ◽  
pp. L604-L619 ◽  
Author(s):  
Praveen Mannam ◽  
Amanda S. Shinn ◽  
Anup Srivastava ◽  
Radu F. Neamu ◽  
Wendy E. Walker ◽  
...  
Keyword(s):  
Mitochondrial Biogenesis ◽  
Mitochondrial Function ◽  
Mononuclear Cells ◽  
Critical Role ◽  
Mitochondrial Pathway ◽  
Mouse Lung ◽  
Peripheral Blood Mononuclear ◽  
Primary Mouse ◽  
Effective Therapeutic Strategy ◽  
Lung Endothelial Cells

Sepsis is a systemic inflammatory response to infection and a major cause of death worldwide. Because specific therapies to treat sepsis are limited, and underlying pathogenesis is unclear, current medical care remains purely supportive. Therefore targeted therapies to treat sepsis need to be developed. Although an important mediator of sepsis is thought to be mitochondrial dysfunction, the underlying molecular mechanism is unclear. Modulation of mitochondrial processes may be an effective therapeutic strategy in sepsis. Here, we investigated the role of the kinase MKK3 in regulation of mitochondrial function in sepsis. Using clinically relevant animal models, we examined mitochondrial function in primary mouse lung endothelial cells exposed to LPS. MKK3 deficiency reduces lethality of sepsis in mice and by lowering levels of lung and mitochondrial injury as well as reactive oxygen species. Furthermore, MKK3 deficiency appeared to simultaneously increase mitochondrial biogenesis and mitophagy through the actions of Sirt1, Pink1, and Parkin. This led to a more robust mitochondrial network, which we propose provides protection against sepsis. We also detected higher MKK3 activation in isolated peripheral blood mononuclear cells from septic patients compared with nonseptic controls. Our findings demonstrate a critical role for mitochondria in the pathogenesis of sepsis that involves a previously unrecognized function of MKK3 in mitochondrial quality control. This mitochondrial pathway may help reveal new diagnostic markers and therapeutic targets against sepsis.

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