Nuclear Translocation Of HIF-1α Research Articles

Proximal tubular FHL2, a novel downstream target of hypoxia inducible factor 1, is a protector against ischemic acute kidney injury

Four-and-a-half LIM domains protein 2 (FHL2) is an adaptor protein that may interact with hypoxia inducible factor 1α (HIF-1α) or β-catenin, two pivotal protective signaling in acute kidney injury (AKI). However, little is known about the regulation and function of FHL2 during AKI. We found that FHL2 was induced in renal tubular cells in patients with acute tubular necrosis and mice model of ischemia-reperfusion injury (IRI). In cultured renal proximal tubular cells (PTCs), hypoxia induced FHL2 expression and promoted the binding of HIF-1 to FHL2 promoter. Compared with control littermates, mice with PTC-specific deletion of FHL2 gene displayed worse renal function, more severe morphologic lesion, more tubular cell death and less cell proliferation, accompanying by downregulation of AQP1 and Na, K-ATPase after IRI. Consistently, loss of FHL2 in PTCs restricted activation of HIF-1 and β-catenin signaling simultaneously, leading to attenuation of glycolysis, upregulation of apoptosis-related proteins and downregulation of proliferation-related proteins during IRI. In vitro, knockdown of FHL2 suppressed hypoxia-induced activation of HIF-1α and β-catenin signaling pathways. Overexpression of FHL2 induced physical interactions between FHL2 and HIF-1α, β-catenin, GSK-3β or p300, and the combination of these interactions favored the stabilization and nuclear translocation of HIF-1α and β-catenin, enhancing their mediated gene transcription. Collectively, these findings identify FHL2 as a direct downstream target gene of HIF-1 signaling and demonstrate that FHL2 could play a critical role in protecting against ischemic AKI by promoting the activation of HIF-1 and β-catenin signaling through the interactions with its multiple protein partners.

An E3 ligase TRIM1 promotes colorectal cancer progression via K63-linked ubiquitination and activation of HIF1α

Accumulating studies have shown that E3 ligases play crucial roles in regulating cellular biological processes and signaling pathways during carcinogenesis via ubiquitination. Tripartite-motif (TRIM) ubiquitin E3 ligases consist of over 70 members. However, the clinical significance and their contributions to tumorigenesis remain largely unknown. In this study, we analyzed the RNA-sequencing expression of TRIM E3 ligases in colorectal cancer (CRC) and identified 10 differentially expressed genes, among which TRIM1 expression predicted poor prognosis of CRC patients. We demonstrated that TRIM1 expression is positively associated with CRC pathological stages, and higher expression is positively correlated with infiltrating levels of immune cells and immunotherapy biomarkers. TRIM1 expression promotes the proliferation and migration of colorectal cancer cells in vitro and in vivo. Transcriptional analysis showed that TRIM1 is responsible for metabolism promotion and immune suppression. Mechanistically, we found that TRIM1 binds HIF1α and mediates its K63-linked ubiquitination, which is required for HIF1α nuclear translocation and subsequent activation. Ubiquitination occurs at Lys214 in the loop between the two PAS domains of HIF1α, and mutation of Lys214 severely disturbs the function of HIF1α. Besides, HIF1α ubiquitination enhances its binding with proteins involved in cellular trafficking and nucleocytoplasmic transport pathway. Collectively, our results indicate TRIM1’s role in predicting prognosis and reveal how TRIM1 functions to upregulate HIF1α expression and promote tumor cell proliferation.

Chrysin inhibits ferroptosis of cerebral ischemia/reperfusion injury via regulating HIF-1α/CP loop

Chrysin is a natural flavonoid with powerful neuroprotective capacity. Cerebral ischemia/reperfusion injury (CIRI) is associated with oxidative stress and ferroptosis. Hypoxia-inducible factor 1α (HIF-1α) and ceruloplasmin (CP) are the critical targets for oxidation reactions and iron transport. But the regulatory mechanism between them is still unclear. Transient middle cerebral artery occlusion (tMCAO) model in rats and oxygen and glucose deprivation/re-oxygenation (OGD/R) model in PC12 cells were applied. Pathological tissue staining and biochemical kit were used to evaluate the effect of chrysin. The relationship between HIF-1α and CP was verified by transcriptomics, qRT-PCR and Western blot. In CIRI, HIF-1α/CP loop was discovered to be the regulatory pathway of ferroptosis. CIRI led to activation and nuclear translocation of HIF-1α, which promoted CP transcription and translation, and downstream ferroptosis. Inhibition of HIF-1α had opposite effect on CP and ferroptosis regulation. Overexpression of CP increased the expression of HIF-1α, nevertheless, inhibited the nuclear translocation of HIF-1α and alleviated CIRI. Silencing CP promoted HIF-1α elevation in nucleus and aggravated CIRI. Mechanistically, chrysin restrained HIF-1α nuclear translocation, thereby inhibiting CP transcription and translation, which in turn reduced downstream HIF-1α expression and mitigated ferroptosis in CIRI. Our results highlight chrysin restrains ferroptosis in CIRI through HIF-1α/CP loop.

Morusin suppresses the stemness characteristics of gastric cancer cells induced by hypoxic microenvironment through inhibition of HIF-1α accumulation

Gastric cancer (GC) is a common, life-threatening malignancy that contributes to the global burden of cancer-related mortality, as conventional therapeutic modalities show limited effects on GC. Hence, it is critical to develop novel agents for GC therapy. Morusin, a typical prenylated flavonoid, possesses antitumor effects against various cancers. The present study aimed to demonstrate the inhibitory effect and mechanism of morusin on the stemness characteristics of human GC in vitro under hypoxia and to explore the potential molecular mechanisms. The effects of morusin on cell proliferation and cancer stem cell-like properties of the human GC cell lines SNU-1 and AGS were assessed by MTT assay, colony formation test, qRT-PCR, flow cytometry analysis, and sphere formation test under hypoxia or normoxia condition through in vitro assays. The potential molecular mechanisms underlying the effects of morusin on the stem-cell-like properties of human GC cells in vitro were investigated by qRT-PCR, western blotting assay, and immunofluorescence assay by evaluating the nuclear translocation and expression level of hypoxia-inducible factor-1α (HIF-1α). The results showed that morusin exerted growth inhibitory effects on SNU-1 and AGS cells under hypoxia in vitro. Moreover, the proportions of CD44+/CD24– cells and the sphere formation ability of SNU-1 and AGS reduced in a dose-dependent manner following morusin treatment. The expression levels of stem cell-related genes, namely Nanog, OCT4, SOX2, and HIF-1α, gradually decreased, and the nuclear translocation of the HIF-1α protein was apparently attenuated. HIF-1α overexpression partially reversed the abovementioned effects of morusin. Taken together, morusin could restrain stemness characteristics of GC cells by inhibiting HIF-1α accumulation and nuclear translocation and could serve as a promising compound for GC treatment.

A preventative role of nitrate for hypoxia-induced intestinal injury

BackgroundStudying effective interventions for hypoxia-induced injury is crucial, particularly in high-altitude areas. Symptoms stemming from intestinal injuries have a significant impact on the health of individuals transitioning from plains to plateau regions. This research explores the effects and mechanisms of nitrate supplementation in preventing hypoxia-induced intestinal injury. MethodsA hypoxia survival mouse model was established using 7% O2 conditions. The intervention with 4 mM sodium nitrate (NaNO3) in drinking water commenced 7 days prior to hypoxia exposure. Weight monitoring, hematoxylin and eosin (HE) staining, transmission electron microscopy (TEM), and intestinal permeability assays were employed for physiological, histological, and functional analyses. Quantitative PCR (qPCR), Western blot, and immunofluorescence were utilized to analyze the levels of tight junction (TJ) proteins and hypoxia-inducible factor 1α (Hif 1α). RNA sequencing (RNA-seq) identified nitrate's target, and chromatin immunoprecipitation (ChIP) verified the transcriptional impact of Hif 1α on TJ proteins. Villin-cre mice infected with AAV9-FLEX-EGFP-Hif 1α were used for mechanism validation. ResultsThe results demonstrated that nitrate supplementation significantly alleviated small intestinal epithelial cell necrosis, intestinal permeability, disruption of TJs, and weight loss under hypoxia. Moreover, the nitrate-triggered enhancement of TJs is mediated by Hif 1α nuclear translocation and its subsequent transcriptional function. The effect of nitrate supplementation on TJs was largely attributed to the stimulation of the EGFR/PI3K/AKT/mTOR/Hif 1α signaling pathways. ConclusionNitrate serves as a novel approach in preventing hypoxia-induced intestinal injury, acting through Hif 1α activation to promote the transcription of TJ proteins. Furthermore, our study provides new and compelling evidence for the protective effects of nitrate in hypoxic conditions, especially at high altitudes.

Hsa_circ_0081065 exacerbates IH-induced EndMT via regulating miR-665/HIF-1α signal axis and HIF-1α nuclear translocation

CircRNAs play an important role in various physiological and pathological biological processes. Despite their widespread involvement, the function of circRNAs in intermittent hypoxia (IH) remain incompletely understood. This study aims to clarify the molecular mechanism of it in IH. Differentially expressed circRNAs were identified by transcriptome sequencing analysis in intermittent hypoxia (IH) model. GO and KEGG enrichment analys were performed on the identified differentially expressed circRNAs. The circular characteristics of hsa_circ_0081065 in human umbilical vein endothelial cells (HUVECs) were detected by RT-qPCR. The sublocalization of hsa_circ_0081065 was examined by FISH. The effect of hsa_circ_0081065 on endothelial to mesenchymal transition (EndMT) was estimated by detecting the expression of EndMT related markers. Various techniques, including RNA-pull down, RIP, EMSA, dual-luciferase reporter assay and immunofluorescence staining were used to investigate the relationship among hsa_circ_0081065, miR-665 and HIF-1α. A total of 13,304 circRNAs were identified in HUVECs treatment with IH, among which 73 were differentially expressed, including 24 upregulated circRNAs and 49 downregulated circRNAs. Notably, hsa_circ_0081065 demonstrated a significantly upregulation. Hsa_circ_0081065 exhibited the circular characteristics of circRNA and was predominantly localized in the cytoplasm. Knockdown of hsa_circ_0081065 inhibited EndMT. Mechanically, we demonstrated that hsa_circ_0081065 acts as a sponge for miR-665 to up-regulate HIF-1α and exacerbate HIF-1α nuclear translocation in HUVECs. We have demonstrated that hsa_circ_0081065 is significantly upregulated in HUVECs treated with IH. Our findings indicate that hsa_circ_0081065 exacerbates IH-induced EndMT through the regulation of the miR-665/HIF-1α signal axis and facilitating HIF-1α nuclear translocation. These results provide a theoretical basis for considering of EndMT as a potential therapeutic target for OSAHS intervention.

Coix seed oil alleviates synovial angiogenesis through suppressing HIF-1α/VEGF-A signaling pathways via SIRT1 in collagen-induced arthritis rats

BackgroundRheumatoid arthritis (RA) is a chronic autoimmune disorder characterized by symmetric arthritis. Coix Seed Oil (CSO) has been shown to reduce inflammation in collagen induced arthritis (CIA) rats. However, the effect of CSO on synovial angiogenesis in RA is unknown. In this study, we aimed to explore whether CSO could inhibit RA synovial angiogenesis and elucidate the underlying mechanisms.MethodsCIA rat models were established and subjected to different doses of CSO treatments for four weeks in vivo. Arthritis index, paw swelling, and weight were recorded to assess clinical symptoms. Hematoxylin and Eosin staining, Safarnin O fast green staining, Micro-CT, Immunohistochemical, and Immunofluorescence (IF) staining were performed to examined changes in synovial and joint tissues. The serum HIF-1α and VEGF-A levels were evaluated through enzyme-linked immunosorbent assay. Fibroblast-like synoviocytes (FLS) of rats was stimulated with tumor necrosis factor-α (TNF-α) for developing inflammatory model in vitro. Optimal concentrations of CSO and TNF-α for stimulation were measured through Cell Counting Kit-8 test. Wound healing and Transwell migration experiments were employed to determine FLS migratory ability. IF staining was performed to assess HIF-1α nuclear translocation in FLS. Protein levels of SIRT1, HIF-1α, VEGF-A, and CD31 were assessed through Western blot. The isolated aortic rings were induced with recombinant rat VEGF-A 165 (VEGF-A165) to observe the CSO inhibitory impact on angiogenesis ex vivo.ResultsCSO attenuated the progression of arthritis in CIA rats, mitigated histopathological deterioration in synovial and joint tissues, significantly inhibited immature vessels labeled with CD31+/αSMA−, and reduced the micro-vessels in VEGF-A165 induced aortic rings. Moreover, it upregulated SIRT1 protein levels in CIA rats and TNF-α induced FLS, but decreased HIF-1α and VEGF-A protein levels. Furthermore, CSO inhibited the migration ability and HIF-1α nuclear translocation of TNF-α induced FLS. Finally, suppressing SIRT1 levels in TNF-α induced FLS enhanced their migration ability, HIF-1α nuclear translocation, and the protein levels of HIF-1α, VEGF-A, and CD31, whereas the inhibitory effect of CSO on TNF-α induced FLS was severely constrained.ConclusionsThis study indicates that CSO can alleviate synovial angiogenesis through suppressing HIF-1α/VEGF-A signaling pathways via SIRT1 in CIA rats.

Shikonin induces hepatocellular carcinoma cell apoptosis by suppressing PKM2/PHD3/HIF-1α signaling pathway

To investigate the mechanism of shikonin-induced death of human hepatocellular carcinoma SMMC-7721 cells. Cultured SMMC-7721 cells and normal hepatocytes (L-02 cells) were treated with 4, 8, or 16 μmol/L shikonin, and the changes in cell viability was assessed using MTT assay. The levels of ATP and lactic acid in the cell cultures were detected using commercial kits. Co-immunoprecipitation and immunofluorescence staining were used to determine the relationship among pyruvate kinase M2 (PKM2), prolyl hydroxylase 3 (PHD3), and hypoxia-inducible factor-1α (HIF-1α). The expressions of PHD3, PKM2, HIF-1α, Bax, cleaved caspase-3, and Bcl-2 in SMMC-7721 cells were detected with Western blotting, and cell apoptosis was analyzed with annexin V-FITC/PI staining. The effects of RNA interference of PKM2 on PHD3 and HIF-1α expressions in SMMC-7721 cells were detected using Western blotting. The IC50 of shikonin against SMMC-7721 and L-02 cells was 8.041 μmol/L and 31.75 μmol/L, respectively. Treatment with shikonin significantly inhibited the protein expressions of PKM2, HIF-1α and PHD3 and nuclear translocation of PKM2 and HIF-1α in SMMC-7721 cells. Coimmunoprecipitation and immunofluorescence staining confirmed that shikonin inhibited the formation of PKM2/PHD3/HIF-1α complex and significantly reduced the contents of lactic acid and ATP in SMMC-7721 cells (P < 0.05). The expressions of PHD3 and HIF-1α decreased significantly after PKM2 knockdown (P < 0.05). Shikonin treatment significantly increased the apoptosis rate, enhanced the expressions of Bax and cleaved caspase-3, and decreased Bcl-2 expression in SMMC-7721 cells (P < 0.05). Shikonin induces apoptosis of SMMC-7721 cells possibly by inhibiting aerobic glycolysis through the PKM2/PHD3/HIF-1α signaling pathway to cause energy supply dysfunction in the cells.

Targeting fatty acid synthase modulates sensitivity of hepatocellular carcinoma to sorafenib via ferroptosis

BackgroundSorafenib resistance is a key impediment to successful treatment of patients with advanced hepatocellular carcinoma (HCC) and recent studies have reported reversal of drug resistance by targeting ferroptosis. The present study aimed to explore the association of fatty acid synthase (FASN) with sorafenib resistance via regulation of ferroptosis and provide a novel treatment strategy to overcome the sorafenib resistance of HCC patients.MethodsIntracellular levels of lipid peroxides, glutathione, malondialdehyde, and Fe2+ were measured as indicators of ferroptosis status. Biological information analyses, immunofluorescence assays, western blot assays, and co-immunoprecipitation analyses were conducted to elucidate the functions of FASN in HCC. Both in vitro and in vivo studies were conducted to examine the antitumor effects of the combination of orlistat and sorafenib and CalcuSyn software was used to calculate the combination index.ResultsSolute carrier family 7 member 11 (SLC7A11) was found to play an important role in mediating sorafenib resistance. The up-regulation of FASN antagonize of SLC7A11-mediated ferroptosis and thereby promoted sorafenib resistance. Mechanistically, FASN enhanced sorafenib-induced ferroptosis resistance by binding to hypoxia-inducible factor 1-alpha (HIF1α), promoting HIF1α nuclear translocation, inhibiting ubiquitination and proteasomal degradation of HIF1α, and subsequently enhancing transcription of SLC7A11. Orlistat, an inhibitor of FASN, with sorafenib had significant synergistic antitumor effects and reversed sorafenib resistance both in vitro and in vivo.ConclusionTargeting the FASN/HIF1α/SLC7A11 pathway resensitized HCC cells to sorafenib. The combination of orlistat and sorafenib had superior synergistic antitumor effects in sorafenib-resistant HCC cells.

Oxygen Gradient Induced in Microfluidic Chips Can Be Used as a Model for Liver Zonation

Availability of oxygen plays an important role in tissue organization and cell-type specific metabolism. It is, however, difficult to analyze hypoxia-related adaptations in vitro because of inherent limitations of experimental model systems. In this study, we establish a microfluidic tissue culture protocol to generate hypoxic gradients in vitro, mimicking the conditions found in the liver acinus. To accomplish this, four microfluidic chips, each containing two chambers, were serially connected to obtain eight interconnected chambers. HepG2 hepatocytes were uniformly seeded in each chamber and cultivated under a constant media flow of 50 µL/h for 72 h. HepG2 oxygen consumption under flowing media conditions established a normoxia to hypoxia gradient within the chambers, which was confirmed by oxygen sensors located at the inlet and outlet of the connected microfluidic chips. Expression of Hif1α mRNA and protein was used to indicate hypoxic conditions in the cells and albumin mRNA and protein expression served as a marker for liver acinus-like zonation. Oxygen measurements performed over 72 h showed a change from 17.5% to 15.9% of atmospheric oxygen, which corresponded with a 9.2% oxygen reduction in the medium between chamber1 (inlet) and 8 (outlet) in the connected microfluidic chips after 72 h. Analysis of Hif1α expression and nuclear translocation in HepG2 cells additionally confirmed the hypoxic gradient from chamber1 to chamber8. Moreover, albumin mRNA and protein levels were significantly reduced from chamber1 to chamber8, indicating liver acinus zonation along the oxygen gradient. Taken together, microfluidic cultivation in interconnected chambers provides a new model for analyzing cells in a normoxic to hypoxic gradient in vitro. By using a well-characterized cancer cell line as a homogenous hepatocyte population, we also demonstrate that an approximate 10% reduction in oxygen triggers translocation of Hif1α to the nucleus and reduces albumin production.

Bone Marrow Macrophages Induce Inflammation by Efferocytosis of Apoptotic Prostate Cancer Cells via HIF-1α Stabilization.

The clearance of apoptotic cancer cells by macrophages, known as efferocytosis, fuels the bone-metastatic growth of prostate cancer cells via pro-inflammatory and immunosuppressive processes. However, the exact molecular mechanisms remain unclear. In this study, single-cell transcriptomics of bone marrow (BM) macrophages undergoing efferocytosis of apoptotic prostate cancer cells revealed a significant enrichment in their cellular response to hypoxia. Here, we show that BM macrophage efferocytosis increased hypoxia inducible factor-1alpha (HIF-1α) and STAT3 phosphorylation (p-STAT3 at Tyr705) under normoxic conditions, while inhibitors of p-STAT3 reduced HIF-1α. Efferocytosis promoted HIF-1α stabilization, reduced its ubiquitination, and induced HIF-1α and p-STAT3 nuclear translocation. HIF-1α stabilization in efferocytic BM macrophages resulted in enhanced expression of pro-inflammatory cytokine MIF, whereas BM macrophages with inactive HIF-1α reduced MIF expression upon efferocytosis. Stabilization of HIF-1α using the HIF-prolyl-hydroxylase inhibitor, Roxadustat, enhanced MIF expression in BM macrophages. Furthermore, BM macrophages treated with recombinant MIF protein activated NF-κB (p65) signaling and increased the expression of pro-inflammatory cytokines. Altogether, these findings suggest that the clearance of apoptotic cancer cells by BM macrophages triggers p-STAT3/HIF-1α/MIF signaling to promote further inflammation in the bone tumor microenvironment where a significant number of apoptotic cancer cells are present.

Hypoxia-inducible factor-1α nuclear accumulation via a MAPK/ERK-dependent manner partially explains the accelerated glycogen metabolism in yak longissimus dorsi postmortem under oxidative stress

Although exhaustive studies have established that oxidative stress is a key contributor to accelerated postmortem metabolism, the specific reasons remain fully undefined. We hypothesized that nuclear accumulation of hypoxia-inducible factor-1α (HIF-1α), a significant transcription factor for energy metabolism, regulates postmortem glycolysis under oxidative stress. Postmortem yak longissimus dorsi (LD) muscles were incubated with saline, hydrogen peroxide (H2O2), H2O2 plus MAPK/ERK inhibitor (U0126), or H2O2 plus U0126 and HIF-1α activator, dimethyloxalylglycine (DMOG), respectively. Changes in HIF-1α, MAPK/ERK signaling pathway, and postmortem metabolism levels were measured in muscles collected from all time points. Oxidative stress significantly activated MAPK/ERK pathway and increased HIF-1α expression and nuclear translocation (P < 0.05) although its mRNA levels were hardly affected (P > 0.05). Additionally, oxidative stress enhanced early postmortem muscle metabolism (P < 0.05). These effects were substantially weakened by U0126 treatment, which prevented oxidative stress-induced HIF-1α accumulation and glycolysis. Interestingly, this attenuation could be bypassed by DMOG (P < 0.05). Altogether, MAPK/ERK-dependent maintenance and enhancement of HIF-1α nuclear accumulation promotes postmortem metabolism during early postmortem under oxidative stress. These findings provide valuable information on postmortem metabolism that HIF-1α regulates glycolysis under oxidative stress and suggest that HIF-1α can serve as a target for controlling postmortem glycolysis.

Anti-angiogenic potential of novel 31kDa protein of Zanthoxylum rhesta is mediated by inhibition of HIF-1α nuclear translocation invivo

A novel anticancer protein was isolated and purified from the pericarp of Zanthoxylum rhesta, a plant of culinary and ethnomedicinal importance from the Asian subcontinent. The partial purification process was followed by the fast protein liquid chromatography, gel permeation chromatography. This purified protein was obtained as a single band in the SDS-PAGE with molecular mass of 31 kDa and was named as ZRP. It showed anti-angiogenic activity in vivo in Swiss albino mice inoculated with EAC cells indicated by the reduced peritoneal angiogenesis. In ovo CAM assay also showed reduced neoangiogenesis. ZRP accompanied in VEGF reduction due to hindered nuclear translocation of HIF-1α which is a clear evidence of anti-angiogenic potential of the ZRP protein.

RETRACTED ARTICLE: Overexpressed transient receptor potential vanilloid 1 (TRPV1) in lung adenocarcinoma harbours a new opportunity for therapeutic targeting

The specific biological function of transient receptor potential vanilloid 1 (TRPV1) in pathogenesis of lung adenocarcinoma (LUAD) remains unclear. In this study, TRPV1 expression in tumor tissues, primary cells and cell lines of LUAD, as well as the mechanism mediating its hyperexpression were systematically studied. Multiple models and techniques were adopted to elucidate the relationship between TRPV1 hyperexpression and tumor recurrence and metastasis. Results showed that TRPV1 expression was increased in tumor tissues and primary tumor cells of LUAD patients. The increased expression was associated with worse overall survival outcome and raised HIF1α levels. TRPV1 expression in A549 and NCI-H292 cells was increased after pretreatment with cigarette smoke extract or spermine NONOate. Moreover, A549 cells with TRPV1 overexpression has enhanced tumor growth rates in subcutaneous grafted tumor models, and increased intrapulmonary metastasis after tail vein infusion in nude BALB/c nude mice. Mechanistically, TRPV1 overexpression in A549 cells promoted HIF1α expression and nuclear translocation by promoting CREB phosphorylation and activation of NOS1-NO pathway, ultimately leading to accelerated cell proliferation and stronger invasiveness. In addition, based on photothermal effects, CuS-TRPV1 mAb effectively targeted and induced apoptosis of TRPV1-A549 cells both in vivo and in vitro, thereby mitigating tumor growth and metastasis induced by xenotransplantation of TRPV1-A549 cells. In conclusion, TRPV1 hyperexpression in LUAD is a risk factor for tumor progression and is involved in proliferation and migration of tumor cells through activation of HIF1α. Our study also attempted a new strategy inhibiting the recurrence and metastasis of LUAD: by CuS-TRPV1 mAb precisely kill TRPV1 hyperexpression cells through photothermal effects.

COMMD10 inhibits HIF1α/CP loop to enhance ferroptosis and radiosensitivity by disrupting Cu-Fe balance in hepatocellular carcinoma

Copper (Cu) is an essential trace element whose serum levels have been reported to act as an effective indicator of the efficacy of radiotherapy. However, little is known about the role of Cu in radiotherapy. In this study we aimed to determine this role and investigate the precise mechanism by which Cu or Cu-related proteins regulate the radiosensitivity of hepatocellular carcinoma (HCC). The expression and function of Cu and copper metabolism MURR1 domain 10 (COMMD10) were assessed via a Cu detection assay, immunostaining, real-time PCR, western blot, a radiation clonogenic assay and a 5-ethynyl-2'-deoxyuridine assay. Ferroptosis was determined by detecting glutathione, lipid peroxidation, malondialdehyde and ferrous ion (Fe) levels. The invivo effects of Cu and COMMD10 were examined with Cu/Cu chelator treatment or lentivirus modification of COMMD10 expression in radiated mouse models. We identified a novel role of Cu in promoting the radioresistance of HCC cells. Ionizing radiation (IR) induced a reduction of COMMD10, which increased intracellular Cu and led to radioresistance of HCC. COMMD10 enhanced ferroptosis and radiosensitivity invitro and invivo. Mechanistically, low expression of COMMD10 induced by IR inhibited the ubiquitin degradation of HIF1α (by inducing Cu accumulation) and simultaneously impaired its combination with HIF1α, promoting HIF1α nuclear translocation and the transcription of ceruloplasmin (CP) and SLC7A11, which jointly inhibited ferroptosis in HCC cells. In addition, elevated CP promoted HIF1α expression by reducing Fe, forming a positive feedback loop. COMMD10 inhibits the HIF1α/CP loop to enhance ferroptosis and radiosensitivity by disrupting Cu-Fe homeostasis in HCC. This work provides new targets and treatment strategies for overcoming radioresistance in HCC. Radiotherapy benefits patients with unresectable or advanced hepatocellular carcinoma (HCC), but its effectiveness is hampered by radioresistance. Herein, we uncovered a novel role for copper in promoting the radioresistance of HCCs. This work has revealed new targets and potential treatment strategies that could be used to sensitize HCC to radiotherapy.

Hypoxia-Mimetic CoCl2 Agent Enhances Pro-Angiogenic Activities in Ovine Amniotic Epithelial Cells-Derived Conditioned Medium.

Amniotic epithelial stem cells (AECs) are largely studied for their pro-regenerative properties. However, it remains undetermined if low oxygen (O2) levels that AECs experience in vivo can be of value in maintaining their biological properties after isolation. To this aim, the present study has been designed to evaluate the effects of a hypoxia-mimetic agent, cobalt chloride (CoCl2), on AECs’ stemness and angiogenic activities. First, a CoCl2 dose-effect was performed to select the concentration able to induce hypoxia, through HIF-1α stabilization, without promoting any cytotoxicity effect assessed through the analysis of cell vitality, proliferation, and apoptotic-related events. Then, the identified CoCl2 dose was evaluated on the expression and angiogenic properties of AECs’ stemness markers (OCT-4, NANOG, SOX-2) by analysing VEGF expression, angiogenic chemokines’ profiles, and AEC-derived conditioned media activity through an in vitro angiogenic xeno-assay. Results demonstrated that AECs are sensitive to the cytotoxicity effects of CoCl2. The unique concentration leading to HIF-1α stabilization and nuclear translocation was 10 µM, preserving cell viability and proliferation up to 48 h. CoCl2 exposure did not modulate stemness markers in AECs while progressively decreasing VEGF expression. On the contrary, CoCl2 treatment promoted a significant short-term release of angiogenic chemokines in culture media (CM). The enrichment in bio-active factors was confirmed by the ability of CoCl2-derived CM to induce HUVEC growth and the cells’ organization in tubule-like structures. These findings demonstrate that an appropriate dose of CoCl2 can be adopted as a hypoxia-mimetic agent in AECs. The short-term, chemical-induced hypoxic condition can be targeted to enhance AECs’ pro-angiogenic properties by providing a novel approach for stem cell-free therapy protocols.

Migration enhancement and HIF-1α nuclear translocation of vascular endothelial cells by flow and hypoxic exposure

Migration enhancement and HIF-1α nuclear translocation of vascular endothelial cells by flow and hypoxic exposure

Anti-Angiogenic Potential of Novel 31kDa Protein of Zanthoxylum Rhesta is Mediated by Inhibition of HIF-1α Nuclear Translocation in Vivo

Anti-Angiogenic Potential of Novel 31kDa Protein of Zanthoxylum Rhesta is Mediated by Inhibition of HIF-1α Nuclear Translocation in Vivo

Mechanical stimulation of chondrocytes regulates HIF-1α under hypoxic conditions

We investigated the effects of hypoxia-inducible factor (HIF)-1α on articular cartilage under mechanical stimulation and the associated mechanisms. Chondrocytes, isolated from articular cartilage from the knee, hip, and shoulder joints of Wistar rats, were subjected to 20 % tensile stress under hypoxic (5% O2) conditions for 24 h. HIF-1α and aggrecan expression was significantly enhanced with mechanical stimulation under hypoxia but not significantly altered with mechanical stimulation under normoxia. The nuclear translocation of HIF-1α was enhanced by mechanical stress under hypoxia. Under both normoxia and hypoxia, a disintegrin and metalloproteinase with thrombospondin motifs (ADAM-TS) 5 expression was significantly reduced with mechanical stimulation compared to that in the group without mechanical stimulation. However, HIF-1α knockdown mitigated changes in aggrecan and ADAM-TS5 expression mediated by mechanical stimulation under hypoxia. The effects of treadmill running on HIF-1α production in the articular cartilage of rat knee joints were also analyzed. HIF-1α production increased in the moderate running group and decreased to the same levels as those in the control group in the excessive running group. This suggests that HIF-1α regulates aggrecan and ADAM-TS5 expression in response to mechanical stimulation under hypoxia and general mechanical stimulation in articular cartilage under hypoxia, while controlling cartilage homeostasis.

Development of a constant pressure perfused ex vivo model of the equine larynx.

Distal axonopathy is seen in a broad range of species including equine patients. In horses, this degenerative disorder of the recurrent laryngeal nerve is described as recurrent laryngeal neuropathy (RLN). The dysfunctional innervation of the cricoarytenoideus dorsalis muscle (CAD) leads to a loss of performance in affected horses. In general, ex vivo models of the larynx are rare and for equine patients, just one short report is available. To allow for testing new therapy approaches in an isolated organ model, we examined equine larynges in a constant pressure perfused setup. In order to check the vitality and functionality of the isolated larynx, the vessels´ reaction to norepinephrine (NE) and sodium nitroprusside (NP) as vasoactive agents was tested. Additionally, the contractility of the CAD was checked via electrical stimulation. To determine the extent of hypoxic alterations, lactate dehydrogenase (LDH) and lactate were measured and an immunofluorescent analysis of hypoxia-inducible factor (HIF-1α), a key transcription factor in hypoxia, was performed. For this, a hypoxia-induced cell culture for HIF-1α was developed. The application of NE led to an expected vasoconstriction while NP caused the expected vasodilation. During a perfusion period of 352 ±20.78 min, LDH values were in the reference range and lactate values slightly exceeded the reference range at the end of the perfusion. HIF-1α nuclear translocation could reliably be detected in the hypoxia-induced cell cultures, but not in sections of the perfused CAD. With the approach presented here, a solid basis for perfusing equine larynges was established and may serve as a tool for further investigations of equine larynx disorders as well as a transferrable model for other species.