bims-kishpe 2024-11-24 papers

bims-kishpe

Biomed News

on HSP70 role in hypoxia and metabolism in ECs

Issue of 2024–11–24
seven papers selected by
Alia Ablieh, Universität Heidelberg

ARF6 as a Novel Activator of HIF-2α in Pulmonary Arterial Hypertension.
A novel hypoxia-induced lncRNA, SZT2-AS1, boosts HCC progression by mediating HIF heterodimerization and histone trimethylation under a hypoxic microenvironment.
Exploring the Endothelin-1 pathway in chronic thromboembolic pulmonary hypertension microvasculopathy.
Knockdown of the long noncoding RNA VSIG2-1:1 promotes the angiogenic ability of human pulmonary microvascular endothelial cells by activating the VEGF/PI3K/AKT pathway.
Catalpol regulates apoptosis and proliferation of endothelial cell via activating HIF-1α/VEGF signaling pathway.
Unanticipated mechanisms of covalent inhibitor and synthetic ligand cobinding to PPARγ.
ETV2 transcriptionally activates Rig1 gene expression and promotes reprogramming of the endothelial lineage.

Am J Respir Cell Mol Biol. 2024 Nov 18.

ARF6 as a Novel Activator of HIF-2α in Pulmonary Arterial Hypertension.

Adam L Fellows, Chien-Nien Chen, Chongyang Xie, Nayana Iyer, Lukas Schmidt, Xiaoke Yin, Luke A Yates, Manuel Mayr, Andrew Cowburn, Lan Zhao, Beata Wojciak-Stothard.

  ADP-ribosylation factor 6 (ARF6), a GTPase associated with cancer metastasis, is activated in the lung endothelium in pulmonary arterial hypertension (PAH). To identify ARF6-regulated pathways relevant to PAH, we performed a state-of-the-art proteomic analysis of human pulmonary artery endothelial cells (HPAECs) overexpressing the wildtype, constitutively active, fast-cycling and dominant negative mutants of ARF6. The analysis revealed a novel link of ARF6 with hypoxia-inducible factor (HIF), in addition to endocytotic vesicle trafficking, cell proliferation, angiogenesis, oxidative stress and lipid metabolism. Active ARF6 markedly increased expression and activity of HIF-2, critical in PAH, with HIF-1 relatively unaffected. Hypoxic ARF6 activation was a prerequisite for HIF-2 activation and HIF-dependent gene expression in HPAECs, PAH blood-derived late outgrowth endothelial colony forming cells (ECFCs) and hypoxic mouse lungs in vivo. A novel ARF6 inhibitor, chlortetracycline (CTC), reduced hypoxia-induced HIF-2 activation, proliferation and angiogenesis in HPAECs and reduced HIF-2 expression in lung and heart tissues of hypoxic mice. PAH ECFCs showed elevated expression and activity of ARF6 and HIF2, which was attenuated by CTC, and oral CTC attenuated development of PH in chronically hypoxic mice. We identify epidermal growth factor receptor (EGFR) as a direct interactor of ARF6, and EGFR signalling as a crucial mechanism linking ARF6 and HIF activation. In conclusion, we are first to demonstrate a key role of ARF6 in the regulation of HIF-2α activation in vitro and in vivo and show that HIF-2α, a master-regulator of vascular remodelling in PAH, can be targeted by a clinically approved antibiotic chlortetracycline.

Keywords:  Endothelium; Hypoxia; Pulmonary Arterial Hypertension

DOI:  https://doi.org/10.1165/rcmb.2024-0149OC
Cell Death Differ. 2024 Nov 22.

A novel hypoxia-induced lncRNA, SZT2-AS1, boosts HCC progression by mediating HIF heterodimerization and histone trimethylation under a hypoxic microenvironment.

Runkun Liu, Yixian Guo, Liang Wang, Guozhi Yin, Hang Tuo, Yifeng Zhu, Wei Yang, Qingguang Liu, Yufeng Wang.

Hypoxic microenvironment plays a critical role in solid tumor growth, metastasis and angiogenesis. Hypoxia-inducible factors (HIFs), which are canonical transcription factors in response to hypoxia, are stabilized under hypoxia and coordinate the process of hypoxia-induced gene expression, leading to cancer progression. Increasing evidence has uncovered that long noncoding RNAs (lncRNAs), which are closely associated with cancer, play crucial roles in hypoxia-mediated HCC progression, while the mechanisms are largely unknown. Here, we identified SZT2-AS1 as a novel lncRNA in HCC, which was induced by hypoxia in a HIF-1-dependent manner and promoted HCC growth, metastasis and angiogenesis both in vitro and in vivo. And SZT2-AS1 also mediated the hypoxia-induced HCC progression. Clinical data indicated that SZT2-AS1 level was substantially increased in HCC and closely associated with poor clinical outcomes, acting as an independent prognostic predictor. Mechanistically, SZT2-AS1 recruited HIF-1α and HIF-1β to form the HIF-1 heterodimer, and it was required for the occupancy of HIF-1 to hypoxia response elements (HREs) and HIF target gene transcription. In addition, SZT2-AS1 was required for hypoxia-induced histone trimethylation (H3K4me3 and H3K36me3) at HREs. Through recruiting methyltransferase SMYD2, SZT2-AS1 promoted trimethylation of H3K4 and H3K36 in HCC cells. Taken together, our results uncovered a lncRNA-involved positive feedback mechanism under hypoxia and established the clinical value of SZT2-AS1 in prognosis and as a potential therapeutic target in HCC.

DOI: https://doi.org/10.1038/s41418-024-01419-x
Sci Rep. 2024 11 16. 14(1): 28308

Exploring the Endothelin-1 pathway in chronic thromboembolic pulmonary hypertension microvasculopathy.

Benchenouf Feriel, Cuomo Alessandra, Gorth J Deborah, Normand Corinne, Thuillet Raphaël, Ottaviani Mina, Akamkam Ali, Menager Jean-Baptiste, Fadel Guillaume, Grynblat Julien, Ghigna Maria-Rosa, Fadel Elie, Savale Laurent, Mercier Olaf, Tu Ly, Humbert Marc, Guignabert Christophe.

  Targeted vasopeptide therapies have significantly advanced the management of pulmonary arterial hypertension (PAH). However, due to insufficient preclinical evidence regarding the involvement of the endothelin-1 (ET-1) pathway in chronic thromboembolic pulmonary hypertension (CTEPH) pathophysiology, the potential of ET-1 receptor antagonism in treating CTEPH remains uncertain. In this study, we investigated the role of the ET-1 pathway in CTEPH microvasculopathy using a multifaceted approach. Plasma ET-1 levels were measured in a cohort of 59 CTEPH patients and 41 healthy controls. Additionally, we evaluated the expression of key ET-1 pathway members in pulmonary explants from CTEPH, idiopathic PAH, and control patients. We used an in vitro system to test the hypothesis that the turbulent flow, observed near the vascular obstructions pathognomonic of CTEPH, enhances ET-1 expression. Our findings were further validated in vivo using a CTEPH piglet model that contains distinct regions representing pre- and post-thrombus lung territories. We found a twofold increase in circulating ET-1 levels in CTEPH patients compared to healthy subjects. Pulmonary explants from CTEPH patients exhibited pronounced overexpression of ET-1, endothelin receptor A (ETA), and phosphorylated myosin light chain (p-MLC) in muscularized pulmonary microvessels, suggesting heightened vascular contraction. In vitro experiments showed that turbulent flow facilitates ET-1 secretion compared to laminar flow regions. Additionally, in the CTEPH piglet model, elevated plasma ET-1 levels were observed compared to controls. Immunofluorescence and confocal microscopy analyses confirmed increased ETA and p-MLC in remodeled arteries from both pulmonary territories. However, ET-1 protein elevation was exclusively observed in the obstructed territory. These findings collectively indicate impaired vascular tone in microvessels of CTEPH patients and the CTEPH piglet model. Furthermore, our data implicates the ET-1 pathway in microvasculopathy, with turbulent flow playing a pathological role. These insights underscore the potential utility of ET-1 receptor antagonists as a promising therapeutic approach for CTEPH treatment.

Keywords:  Chronic thromboembolic pulmonary hypertension; Endothelin receptor antagonist; Endothelin-1; Microvasculopathy; Pulmonary vasculature; Therapeutic target

DOI:  https://doi.org/10.1038/s41598-024-79623-5
Respir Res. 2024 Nov 20. 25(1): 412

Knockdown of the long noncoding RNA VSIG2-1:1 promotes the angiogenic ability of human pulmonary microvascular endothelial cells by activating the VEGF/PI3K/AKT pathway.

Xiaoya Hu, Yihui Zheng, Mingchu Fang, Zhongjie Liang, Chao Wen, Jing Lin, Zhenlang Lin, Shangqin Chen.

   BACKGROUND: Abnormal pulmonary vascular development poses significant clinical challenges for infants with bronchopulmonary dysplasia (BPD). Although numerous factors have been suggested to control the development of pulmonary blood vessels, the mechanisms underlying the role of long noncoding RNAs (lncRNAs) in this process remain unclear.
METHODS: A lncRNA array was used to measure the differential expression of lncRNAs in premature infants with and without BPD. The expression of lncRNA-VSIG2-1:1 in patients with BPD and hyperoxia-induced human pulmonary microvascular endothelial cells (HPMECs) was assessed using quantitative reverse transcription-polymerase chain reaction (qRT-PCR). Fluorescence in situ hybridization (FISH) assay was performed to detect the subcellular localization of lncRNA-VSIG2-1:1. Pulmonary microvascular endothelial cells were stably transfected with adenoviral vectors to silence or overexpress lncRNA-VSIG2-1:1. The effects of lncRNA-VSIG2-1:1 on the proliferation, migration, and tube formation abilities of HPMECs subjected to hyperoxia were examined by performing Cell Counting Kit-8 (CCK-8), cell migration, and tubule formation assays. RNA sequencing (RNA-seq) was performed to determine the correlation between lncRNA-VSIG2-1:1 and phosphoinositide-3-kinase (PI3K)/protein kinase B (AKT). The protein levels of vascular endothelial growth factor (VEGF), p-PI3K, PI3K, p-AKT, and AKT were determined using western blotting.
RESULTS: The expression of lncRNA-VSIG2-1:1 was upregulated in patients with BPD and hyperoxia-treated HPMECs. Inhibiting lncRNA-VSIG2-1:1 expression promoted the proliferation, migration, and tube-formation abilities of HPMECs, while significantly increasing VEGF, p-PI3K, and p-AKT levels.
CONCLUSION: Our findings reveal that the suppression of lncRNA-VSIG2-1:1 expression stimulates angiogenesis in vitro by inducing the initiation of the VEGF/PI3K/AKT signaling pathway. This observation may aid the development of novel therapeutic targets for treating BPD.

Keywords:  Angiogenesis; Bronchopulmonary dysplasia; Long noncoding RNA; Pulmonary microvascular endothelial cells; VEGF/PI3K/AKT pathway

DOI:  https://doi.org/10.1186/s12931-024-03039-y
Sci Rep. 2024 11 16. 14(1): 28327

Catalpol regulates apoptosis and proliferation of endothelial cell via activating HIF-1α/VEGF signaling pathway.

Jinrong Ni, Qunhu Zhang, Luetao Jiang, Haihu Wang, Chengji Zhang, Jielin Deng.

  Burn injuries, especially severe ones, causes microcirculation disorders in local wounds and distant tissues, leading to ischemia and hypoxia of body tissues and organs. The key to prevent and treat complications and improve prognosis after burns is to improve the state of ischemia and hypoxia of tissue and restore the blood supply of organs. Catalpol is an iridoid glycoside compound isolated from Rehmannia radix, which has been widely reported to have various of functions, including antioxidative stress, anti-inflammation, anti-apoptosis, and neuroprotection. However, the pharmacologic action and underlying mechanism of Catalpol in angiogenesis after burn injury remains unclear. The study investigated whether Catalpol regulates apoptosis and proliferation following vascular injury induced by burns using an in vitro model of oxygen-glucose deprivation (OGD) with a human umbilical vein endothelial (HUVE) cell line. The results showed that treatment with Catalpol reduces the level of apoptosis and promotes proliferation of endothelial cell. Mechanistically, Catalpol increases the expression of vascular endothelial growth factor (VEGF) by activating Hypoxia-inducible factor-1α (Hif-1α), resulting in increased expression of related downstream effector molecules. The current study suggested that Catalpol is a promising compound for endothelial protection in burns. It may be an efficient Hif-1α activator for endothelial cell deprived of oxygen and glucose.

Keywords:  Apoptosis; Catalpol; Endothelial cell; Hif-1α; Proliferation; VEGF

DOI:  https://doi.org/10.1038/s41598-024-78126-7
Elife. 2024 Nov 18. pii: RP99782. [Epub ahead of print]13

Unanticipated mechanisms of covalent inhibitor and synthetic ligand cobinding to PPARγ.

Jinsai Shang, Douglas J Kojetin.

  Peroxisome proliferator-activated receptor gamma (PPARγ) is a nuclear receptor transcription factor that regulates gene expression programs in response to ligand binding. Endogenous and synthetic ligands, including covalent antagonist inhibitors GW9662 and T0070907, are thought to compete for the orthosteric pocket in the ligand-binding domain (LBD). However, we previously showed that synthetic PPARγ ligands can cooperatively cobind with and reposition a bound endogenous orthosteric ligand to an alternate site, synergistically regulating PPARγ structure and function (Shang et al., 2018). Here, we reveal the structural mechanism of cobinding between a synthetic covalent antagonist inhibitor with other synthetic ligands. Biochemical and NMR data show that covalent inhibitors weaken-but do not prevent-the binding of other ligands via an allosteric mechanism, rather than direct ligand clashing, by shifting the LBD ensemble toward a transcriptionally repressive conformation, which structurally clashes with orthosteric ligand binding. Crystal structures reveal different cobinding mechanisms including alternate site binding to unexpectedly adopting an orthosteric binding mode by altering the covalent inhibitor binding pose. Our findings highlight the significant flexibility of the PPARγ orthosteric pocket, its ability to accommodate multiple ligands, and demonstrate that GW9662 and T0070907 should not be used as chemical tools to inhibit ligand binding to PPARγ.

Keywords:  NMR spectroscopy; covalent inhibitor; crystal structure; molecular biophysics; none; nuclear receptors; pharmacology; structural biology

DOI:  https://doi.org/10.7554/eLife.99782
Sci Rep. 2024 Nov 19. 14(1): 28688

ETV2 transcriptionally activates Rig1 gene expression and promotes reprogramming of the endothelial lineage.

Young Geun Choi, Xiao Ma, Satyabrata Das, Javier E Sierra-Pagan, Thijs Larson, Wuming Gong, Hesham A Sadek, Jianyi Jay Zhang, Mary G Garry, Daniel J Garry.

  ETV2 is an essential transcription factor as Etv2 null murine embryos lack all vasculature, blood and are lethal early during embryogenesis. Previous studies have established that ETV2 functions as a pioneer factor and directly reprograms fibroblasts to endothelial cells. However, the underlying molecular mechanisms regulating this reprogramming process remain incompletely defined. In the present study, we examined the ETV2-RIG1 cascade as regulators that govern ETV2-mediated reprogramming. Mouse embryonic fibroblasts (MEFs) harboring an inducible ETV2 expression system were used to overexpress ETV2 and reprogram these somatic cells to the endothelial lineage. Single-cell RNA-seq from reprogrammed fibroblasts defined the induction of the transcriptional network involved in Rig1-like receptor signaling pathways. Studies using ChIP-seq, electrophoretic mobility shift assays, and transcriptional assays demonstrated that ETV2 was a direct upstream activator of Rig1 gene expression. We further demonstrated that the knockdown of Rig1 and separately, Nfκb1 using shRNA significantly reduced the efficiency of endothelial cell reprogramming. These results highlight that ETV2 reprograms fibroblasts to endothelial cells by directly activating RIG1. These findings extend our current understanding of the molecular mechanisms underlying ETV2-mediated reprogramming and will be important in the design of revascularization strategies for the treatment of ischemic tissues such as ischemic heart disease.

Keywords:  ETV2; RIG1; Reprogramming

DOI:  https://doi.org/10.1038/s41598-024-78115-w