Regulation Of HIF-1α Expression Research Articles

FBP1 over-expression suppresses HIF-1α in papillary thyroid cancer

Papillary thyroid carcinoma (PTC) is generally a slow-growing disease with a favorable 10-year survival rate. However, about 10% of PTC cases show significant aggressiveness, with tendencies for local invasion or distant metastasis, the mechanisms of which remain unclear. This study aims to identify predictive indicators and explore new potential targets for clinical treatment, highlighting the need for novel biomarkers and therapeutic targets. We analyzed FBP1 expression in PTC tissues. Cell proliferation, apoptosis, and invasion were evaluated with and without FBP1 overexpression in PTC cells to assess FBP1’s effects. We then investigated whether FBP1 reduces PTC cell tumorigenesis and metastasis by regulating HIF-1α expression. FBP1 expression was reduced in PTC samples and showed a negative correlation with T stage. In vitro experiments indicated that FBP1 acts as a hypoxia response inhibitor, regulating tumor cells. Additionally, FBP1 inhibited the proliferation, apoptosis, and invasion of thyroid cancer cells by modulating HIF-1α expression. Our results provide new insights into the role of FBP1 in PTC progression and indicate that targeting the FBP1-HIF-1α axis could be a promising therapeutic approach for this disease.

N6-methyladenosine modification of hypoxia-inducible factor-1α regulates Helicobacter pylori-associated gastric cancer via the PI3K/AKT pathway.

Helicobacter pylori (H. pylori) colonizes the human gastric mucosa and is implicated in the development of gastric cancer (GC). The tumor microenvironment is characterized by hypoxia, where hypoxia-inducible factor-1α (HIF-1α) plays a key role as a transcription factor, but the mechanisms underlying H. pylori-induced HIF-1α expression and carcinogenesis remain unclear. To explore the underlying mechanism of H. pylori-induced HIF-1α expression in promoting the malignant biological behavior of gastric epithelial cells (GES-1). The study was conducted with human GES-1 cells in vitro. Relative protein levels of methyltransferase-like protein 14 (METTL14), HIF-1α, main proteins of the PI3K/AKT pathway, epithelial-mesenchymal transition (EMT) biomarkers, and invasion indicators were detected by Western blot. Relative mRNA levels of METTL14 and HIF-1α were detected by quantitative reverse transcription-polymerase chain reaction. mRNA stability was evaluated using actinomycin D, and the interaction between METTL14 and HIF-1α was confirmed by immunofluorescence staining. Cell proliferation and migration were evaluated by cell counting kit-8 assay and wound healing assay, respectively. H. pylori promoted HIF-1α expression and activated the PI3K/AKT pathway. Notably, METTL14 was downregulated in H. pylori-infected gastric mucosal epithelial cells and positively regulated HIF-1α expression. Functional experiments showed that the overexpression of HIF-1α or knockdown of METTL14 enhanced the activity of the PI3K/AKT pathway, thereby driving a series of malignant transformation, such as EMT and cell proliferation, migration, and invasion. By contrast, the knockdown of HIF-1α or overexpression of METTL14 had an opposite effect. H. pylori-induced underexpression of METTL14 promotes the translation of HIF-1α and accelerates tumor progression by activating the PI3K/AKT pathway. These results provide novel insights into the carcinogenesis of GC.

SPA inhibits hBMSC osteogenic differentiation and M1 macrophage polarization by suppressing SETD2 in acute suppurative osteomyelitis

To clarify the impact of SETD2 on macrophage function in pediatric patients with acute suppurative osteomyelitis and to elucidate the precise underlying mechanism. To gain insights into the potential functions of SETD2, a comprehensive study was conducted utilizing a co-culture model of human bone mesenchymal stem cells (hBMSCs) and bone marrow-derived macrophages (THP-1). A range of techniques were employed, including quantitative polymerase chain reaction, western blotting, ELISA, alkaline phosphatase activity assays, alizarin red S staining, luciferase reporter gene assays, and chromatin immunoprecipitation, to unravel the intricate interactions and molecular mechanisms involving SETD2 in this system. It was observed that SETD2 expression was reduced in THP-1 cells stimulated by staphylococcal protein A (SPA). Furthermore, the downregulation of SETD2 resulted in elevated M1 macrophage polarization and glycolysis, effects that were mitigated by SPA stimulation. Notably, SPA-stimulated THP-1 cells exhibited an increase in HIF-1α expression, which exhibited an inverse correlation with SETD2 levels. Moreover, it was discovered that SETD2 functioned as a catalyst for H3K36me3 and bound to the HIF-1α gene, which, in turn, regulated HIF-1α expression. Furthermore, the suppression of HIF-1α abrogated the consequences of SETD2 downregulation on glycolysis and M1 macrophage polarization. Lastly, the study demonstrated that M1 macrophage polarization serves as a mediator for BMP4’s inhibitory effect on osteogenic differentiation of hBMSCs. This research has uncovered a previously unknown role of SETD2 in macrophages during osteomyelitis, revealing its significance in the pathogenesis of this condition. These findings suggest SETD2 as a novel target for the treatment of osteomyelitis.

MicroRNA-138 inhibits hypoxia-inducible factor 1α expression in breast cancer cells

Background Hypoxia, a critical feature during cancer development, leads to the stabilization and activation of the hypoxia-inducible factor 1-alpha (HIF-1α) to drive the expression of many target genes which in turn can promote many aspects of breast cancer biology, mainly metastasis and resistance to therapy. MicroRNAs are known to modulate the expression of many genes involved in breast cancer tumorigenesis. In this study, we examined the regulatory effect of miRNAs on HIF1α expression. Methods MCF-7 and MDA-MB-231 were cultivated under normoxia or hypoxia conditions. TaqMan-Low Density Array (TLDA) was used to characterize the miRNA signatures. Wild-Type (WT) or mutated fragments of HIF-1α 3’UTR containing the miR-138 potential target site were cloned downstream of the Renilla luciferase gene in the psiCHECK-1 plasmid. Luciferase assays were then carried out. A lentiviral vector containing copGFP as a reporter gene was prepared and transduced into MCF-7 and MDA-MB-231 cells to assess the effect of identified deregulated miRNAs on HIF-1α expression. Results Under hypoxic conditions, MCF-7 cells showed deregulated expression for 12 miRNAs. In the case of MDA-MB-231 cells, 16 miRNAs were deregulated in response to hypoxia. Interestingly, miR-138 that was downregulated in both MCF-7 and MDA-MB-231 cells cultivated under hypoxic conditions appeared to have a binding site in 3’UTR of HIF-1α. Moreover, our results indicated that miR-138 could down regulate HIF-1α expression, upon binding directly to its 3’UTR. Conclusions Interestingly, our data highlights miR-138 as a potential therapeutic target to reduce HIF-1α expression and subsequently restrain breast cancer invasion and metastasis.

Hypoxia-Inducible Factor-1α Regulates High Phosphate-Induced Vascular Calcification via Type III Sodium-Dependent Phosphate Cotransporter 1.

Vascular calcification (VC) has a high incidence in patients with chronic kidney disease, which is a worldwide public health problem and presents a heavy burden to society. Hypoxia-inducible factor (HIF)-1α, the active subunit of HIF-1, has been reported to play a vital role in high phosphate-induced VC. However, the underlying mechanism is still undetermined, and effective treatment is unavailable. In the present study, human aortic smooth muscle cells (HASMCs) were cultured under normal or high phosphate media conditions. HIF-1α small interfering RNA and overexpression plasmids were employed to regulate HIF-1α expression. Phosphonoformic acid was employed to restrain the function of type III sodium-dependent phosphate cotransporter 1 (Pit-1). The expression levels of HIF-1α, Pit-1, runt-related transcription factor 2 (Runx2), and smooth muscle 22 alpha (SM22α) were evaluated, and the calcium contents were also examined. Cell growth was assessed using an MTT assay. High phosphate stimulation caused an upregulation in HIF-1α and Pit-1 expression levels and induced calcium depositions in HASMCs. Upregulation of Runx2 expression accompanied by downregulation of SM22α expression was observed in the high phosphate group. Following the suppression of HIF-1α expression, there was a concomitant attenuation in Pit-1 expression, calcium deposition, the alteration of phenotypic transition marker genes, and vice versa. The most serious calcium deposition was noted in HASMCs cultured under high phosphate conditions which were pretreated with a HIF-1α overexpression plasmid. However, when the biological functions of Pit-1 were restrained, the putative serious calcium deposition was not formed even in HASMCs transfected with a HIF-1α overexpression plasmid. The findings confirmed that HIF-1α regulated Pit-1 expression and exerted its pro-calcifying effect through Pit-1, which identified HIF-1α and Pit-1 as therapeutic targets for high phosphate-induced VC.

Abstract 4590: Antitumor effects of histone deacetylase inhibitors on breast cancer cell viability and mammosphere formation

Abstract Histone deacetylases (HDACs) are overexpressed in various types of primary human cancer and have become attractive targets for cancer therapy. This study aimed to evaluate the cytotoxic properties of three HDAC inhibitors (suberoylanilide hydroxamic acid (SAHA, or Vorinostat), Valproic Acid (VPA) and CAY10603) and compare them to Paclitaxel (PTX), a chemotherapy agent used to treat Triple Negative Breast Cancers (TNBCs). In the present study, drug response in the MDA-MB-231, MDA-MB-468, and MCF-10A cell lines was evaluated by cell viability assays. Apoptosis and cell cycle distribution were analyzed by flow cytometry. Mammospheres generated from MDA-MB-231 and MDA-MB-468 breast cancer cells were also subjected to drug treatment; their response was assessed under both normoxic and hypoxic conditions. We found that SAHA and CAY10603 inhibited breast cancer cell viability without significantly affecting MCF-10A cells. SAHA, CAY10603, and VPA all arrested a significant percentage of MDA-MB-231 cells at G2/M phase after 24h compared to controls. TNBC cells experienced an increase in both early and late apoptosis following treatment with the most effective concentrations of SAHA, CAY10603, and VPA (1.8µM, 2µM, and 0.4mM, respectively). SAHA proved to be the most effective drug for induction of apoptosis compared to the control, followed by CAY10603 (8.26% and 6%, 3.67%). In both TNBCs and mammospheres under hypoxic conditions, SAHA and CAY10603 down regulated HIF-1α expression significantly, but PTX did not. In contrast, VPA and PTX down regulated CTNNB1 gene expression. This study indicates that epi-drugs exert an anti-tumor effect on TNBCs, suggesting a potential role for HDAC inhibitors as novel therapeutics for breast cancer patients. Citation Format: Golnaz Asaadi Tehrani, Becca Kubick, Meenal Datta. Antitumor effects of histone deacetylase inhibitors on breast cancer cell viability and mammosphere formation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4590.

Abstract 373: Transcriptional and post-transcriptional positive regulatory loops between HIF-1a and the RNA-Binding protein hnRNPA18

Abstract Hypoxia, the scarcity of oxygen, is a hallmark of solid tumors to which they adapt by activating the hypoxia-inducible transcription factor 1 (HIF-1). HIF activates the transcription of genes bearing hypoxia responsive elements (RE) in their promoter regions including genes related to angiogenesis, glucose metabolism, cell proliferation, survival, invasion, and metastasis. The HIF-1 pathway is thus an attractive target to prevent cancer aggressiveness and improve the effectiveness of cancer therapy. HIF-1α activity is known to be mainly regulated through post-translational modification by prolyl hydroxylase domain (PHD) enzymes, but accumulating evidence indicate that it is also regulated by other mechanisms such as transcriptional initiation, translational initiation, protein-protein interaction, post-translational modifications and post-transcriptionally, primarily through the action of trans-acting factors (noncoding RNAs and RNA-binding proteins) that interact with the HIF-1α mRNA to regulate its decay and translational rates. We recently identified the RNA binding protein hnRNP A18 as a regulator of HIF-1α translation under hypoxic conditions. hnRNP A18 is a nuclear stress responsive protein that translocates to the cytosol in response to cellular stress including hypoxia to bind to a recognition motif in the 3’UTR of its targeted transcripts including HIF-1α to stabilize the transcripts and increase their translation. Using a bicistronic reporter plasmid containing a HIF-1α internal ribosome entry site (IRES) we now show that hnRNP A18 can also regulate HIF-1α through its IRES in the 5’UTR. In fact, cells expressing hnRNP A18 significantly increase translation of a reporter CAT protein under the control of HIF-1α IRES in the presence of the hypoxia mimetic agent CoCl2. On the other hand, deletion of hnRNP A18 prevents the translation of the CAT reporter protein even in the presence of CoCl2. Moreover, we also identified several HIF-1α REs in hnRNP A18 promoter. Two of the predicted HIF-1α REs, located at -57 and -226 upstream of the hnRNP A18 start codon were validated as bonafide HIF-1α binding sites by Electromobility Shift Assay with recombinant HIF-1α protein and by Chromatin Immunoprecipitation assay in human pancreatic cancer cells Panc 01. These data thus indicate that a positive regulatory loop between hnRNP A18 and HIF1-1α exist to amplify HIF-1α expression under cellular stress. hnRNP A18 can upregulate HIF1α protein expression by stabilizing its transcript at the 3’UTR and by binding to its IRES. On the other hand, HIF-1α can upregulate hnRNP A18 by binding to its proximal promoter. Targeting hnRNP A18 could thus provide a new mechanism to regulate HIF-1α expression and sensitize cancer cells to therapies. Citation Format: Eduardo Solano-Gonzalez, France Carrier. Transcriptional and post-transcriptional positive regulatory loops between HIF-1a and the RNA-Binding protein hnRNPA18 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 373.

NMAAP1 regulated macrophage polarizion into M1 type through glycolysis stimulated with BCG

Bacillus Calmette Guerin (BCG) perfusion is widely used as cancer adjuvant therapy, in which macrophages play an important role. Novel macrophage activated associated protein 1 (NMAAP1), upregulated after BCG's activation, was proved to promote macrophage polarization to the M1 type. We found that BCG could stimulate mice BMDM to the M1 type and kill tumor cells. After the deletion of NMAAP1, the tumor volume of mice became larger, and the number of M1 type macrophages in the tumor decreased significantly. When macrophages were induced into the M1 type, aerobic glycolysis, the Warburg effect manifested in the increased uptake of glucose and the conversion of pyruvate to lactic acid. NMAAP1 could bind with IP3R and regulate macrophage polarization to the M1 type. However, the specific mechanism of how NMAAP1 regulates macrophage polarization towards the M1 type and plays an antitumor role must be clarified. NMAAP1 could promote the release of lactic acid and pyruvate, enhance the glycolysis of macrophages, and affect the expression of HIF-1α. After inhibition of glycolysis by 2-DG and lactic acid generation by FX11, the effects of NMAAP1 promoting macrophage polarization to the antitumor M1 type were weakened. Furthermore, NMAAP1 upregulated the expression of HIF-1α, which is associated with glycolysis. Moreover, the Ca2+/NF-κB pathway regulated HIF-1α expression by NMAAP1 in the macrophages. NMAAP1 promotes the polarization of macrophages towards the M1 type by affecting the Warburg effect stimulated by BCG.

Lnc AC016727.1/BACH1/HIF-1 α signal loop promotes the progression of non-small cell lung cancer

BackgroundLong noncoding RNAs (lncRNAs) have been reported to play vital roles in the development and progression of cancer. However, their biological significance and functional mechanisms in non-small cell lung cancer (NSCLC) are mostly unclear.MethodsWe performed RNA-sequencing to predict the differential expression of lncRNAs in clinical NSCLC and paired paracancerous lung tissues. To identify lncRNA expression, quantitative polymerase chain reaction (qPCR) was used. Using both cell and mouse models, We studied lncRNA AC016727.1’s function in NSCLC growth and metastasis. Western blot assays, dual luciferase reporter assays, and chromatin immunoprecipitation were used to analyze the functional mechanism of lncRNA AC016727.1.ResultsOur larger NSCLC cohorts validated that the lncRNA AC016727.1 was upregulated in 94 paired NSCLC tissues and correlated with poor survival. Functionally, lncRNA AC016727.1 downregulation inhibited NSCLC cell proliferation, aerobic glycolysis, EMT, and migration, inducing apoptosis. Conversely, upregulated lncRNA AC016727.1 expression exhibited the opposite effect, promoting NSCLC cell survival. Importantly, lncRNA AC016727.1 knockdown inhibited lung cancer growth and slowed the progression of lung metastasis in nude mouse models. Mechanistically, lncRNA AC016727.1 upregulated BACH1 target gene expression by acting as a sponge for miR-98-5p, thereby functioning as a competing endogenous RNA. The function of lncRNA AC016727.1 is mediated by the miR-98-5p/BACH1 axis in NSCLC cells. Meanwhile, the transcription factor HIF-1α can bind to the promoter and activate lncRNA AC016727.1 transcription. lncRNA AC016727.1 regulates HIF-1α expression via BACH1 in NSCLC and forms the lncRNA AC016727.1/BACH1/HIF-1α signaling loop under hypoxic conditions.ConclusionOur study reveals a novel lncRNA AC016727.1/BACH1/HIF-1α signaling loop in the progression of NSCLC under hypoxic conditions, suggesting that lncRNA AC016727.1 could act as a useful biomarker for NSCLC and a new therapeutic target.

Iguratimod promotes functional recovery after SCI by repairing endothelial cell tight junctions

Destruction of the blood-spinal cord barrier (BSCB) after spinal cord injury (SCI) is an important factor promoting the progression of the injury. This study addressed how to repair the BSCB in order to promote the repair of injured spinal cords. Iguratimod (IGU), an anti-rheumatic drug, has been approved for clinical use. A spinal cord injury mouse model and TNF-α-stimulated bEnd.3 cells were used to investigate the effect and mechanism of IGU on injured BSCB. An intracerebroventricular osmotic pump was used to administer drugs to the SCI mouse model. The results showed that the SCI mice in the treatment group had better recovery of neurological function than the control group. Examination of the tissue revealed better repair of the BSCB in injured spinal cords after medication. According to the results from the cell model, IGU promoted the expression of tight junction proteins and reduced cell permeability. Further research found that IGU repaired the barrier function by regulating glycolysis levels in the injured endothelial cells. In studying the mechanism, IGU was found to regulate HIF-1α expression through the NF-κB pathway, thereby regulating the expression of the glycolytic enzymes related to endothelial injury. In summary, IGU promoted functional recovery in vivo by repairing the BSCB. In vitro, IGU regulated the level of glycolysis in the damaged endothelium through the NF-κB pathway, thereby repairing the tight junctions between the endothelium. Therefore, IGU may become a potential drug for treating spinal cord injury.

Ponatinib delays the growth of solid tumours by remodelling immunosuppressive tumour microenvironment through the inhibition of induced PD-L1 expression.

Therapeutic modalities including chemo, radiation, immunotherapy, etc. induce PD-L1 expression that facilitates the adaptive immune resistance to evade the antitumour immune response. IFN-γ and hypoxia are some of the crucial inducers of PD-L1 expression in tumour and systemic microenvironment which regulate the expression of PD-L1 via various factors including HIF-1α and MAPK signalling. Hence, inhibition of these factors is crucial to regulate the induced PD-L1 expression and to achieve a durable therapeutic outcome by averting the immunosuppression. B16-F10 melanoma, 4T1 breast carcinoma, and GL261 glioblastoma murine models were established to investigate the in vivo antitumour efficacy of Ponatinib. Western blot, immunohistochemistry, and ELISA were performed to determine the effect of Ponatinib on the immunomodulation of tumour microenvironment (TME). CTL assay and flow cytometry were such as p-MAPK, p-JNK, p-Erk, and cleaved caspase-3 carried out to evaluate the systemic immunity induced by Ponatinib. RNA sequencing, immunofluorescence and Western blot analysis were used to determine the mechanism of PD-L1 regulation by Ponatinib. Antitumour immunity induced by Ponatinib were compared with Dasatinib. Here, Ponatinib treatment delayed the growth of tumours by inhibiting PD-L1 and modulating TME. It also downregulated the level of PD-L1 downstream signalling molecules. Ponatinib enhanced the CD8 T cell infiltration, regulated Th1/Th2 ratio and depleted tumour associated macrophages (TAMs) in TME. It induced a favourable systemic antitumour immunity by enhancing CD8 T cell population, tumour specific CTL activity, balancing the Th1/Th2 ratio and lowering PD-L1 expression. Ponatinib inhibited FoxP3 expression in tumour and spleen. RNA sequencing data revealed that Ponatinib treatment downregulated the genes related to transcription including HIF-1α. Further mechanistic studies showed that it inhibited the IFN-γ and hypoxia induced PD-L1 expression via regulating HIF-1α. Dasatinib was used as control to prove that Ponatinib induced antitumour immunity is via PD-L1 inhibition mediated T cell activation. RNA sequencing data along with rigorous in vitro and in vivo studies revealed a novel molecular mechanism by which Ponatinib can inhibit the induced PD-L1 levels via regulating HIF-1α expression which leads to modulation of tumour microenvironment. Thus, our study provides a novel therapeutic insight of Ponatinib for the treatment of solid tumours where it can be used alone or in combination with other drugs which are known to induce PD-L1 expression and generate adaptive resistance.

NSD1 promotes esophageal cancer tumorigenesis via HIF1α signaling.

Unlike angiogenesis in normal tissues, tumor angiogenesis is typically dysregulated, during which the HIF1/VEGFA signaling pathway plays a pivotal role. Solid tumors generate immature vessels, which promote tumor progression and treatment resistance. NSD1 can di-methylate histone 3 lysine 36 and regulate transcription factors binding to the promoters of various genes. However, the role of NSD1 in tumorigenesis remains elusive. Here, we evaluated the relationship between NSD1 signaling and HIF1 signaling. It was found that NSD1 transcriptionally regulates HIF1α expression by recruiting STAT3 molecule into the HIF1α promoter. In vivo xenograft experiments further confirmed that HIF1α and STAT3 maintenance is essential for NSD1-mediated tumor progression and angiogenesis. Therefore, the NSD1/STAT3/HIF1α signaling pathway may be a novel and effective treatment target for ESCA.

Ligand-gated ion channel P2X7 regulates hypoxia-induced factor-1α mediated pain induced by dental pulpitis in the medullary dorsal horn.

Dental pulpitis often induces severe pain, and the molecular immune response is remarkable in both peripheral and central nervous system. Accumulating evidence indicates that activated microglia in the medullary dorsal horn (MDH) contribute to dental pulpitis induced pain. The P2X7 receptor plays an important role in driving pain and inflammatory processes, and its downstream target hypoxia-induced factor-1α (HIF-1α) has a crucial role in maintaining inflammation. However, the relationship between P2X7 and HIF-1α in dental inflammatory pain remains unclear. This study demonstrated that the degree of inflammation in the dental pulp tissue became more severe in a time-dependent manner by establishing a rat dental pulpitis model via pulp exposure. Meanwhile, the expression of P2X7, HIF-1α, IL-1β, and IL-18 in the MDH increased most on the seventh day when the pain threshold was the lowest in the dental pulpitis model. Furthermore, lipopolysaccharides (LPS) increased P2X7-mediated HIF-1α expression in microglia. Notably, the suppression of P2X7 caused less IL-1β and IL-18 release and lower HIF-1α expression, and P2X7 antagonist Brilliant Blue G (BBG) could alleviate pain behaviors of the dental pulpitis rats. In conclusion, our results provide further evidence that P2X7 is a key molecule, which regulates HIF-1α expression and inflammation in dental pulpitis-induced pain.

PARP14 promotes the growth and glycolysis of acute myeloid leukemia cells by regulating HIF-1α expression

ObjectiveAcute myeloid leukemia (AML) is an aggressive hematological malignancy with a poor prognosis. This study aimed to investigate the action of PARP14 in the growth and glycolysis of AML. MethodsThe clinical samples of AML patients were collected, and the expression of PARP14 was detected. AML cells were transfected with PARP14, HIF-1α or treated with NF-KB inhibitor (BAY11–7082) or PARP14 inhibitor (RBN012759). Cell proliferation was detected by CCK-8 and colony formation assays, apoptosis by flow cytometry, glucose consumption and lactate production by glucose and lactate kits, ECAR and OCR by XF96 bioenergy analyzer, and related protein levels by Western blot. A mouse xenograft tumor model was established to evaluate the effect of PARP14 on tumor formation. ResultsSignificant upregulation of PARP14 expression was observed in AML. PARP14 promoted AML cell proliferation and glycolysis and inhibited apoptosis, while PARP14 deficiency had the opposite effect. PARP14 promoted HIF-1α expression by activating NF-κB. HIF-1α silencing reversed the cancer-promoting effect of PARP14. In vivo results suggested that PARP14 promoted tumor formation. ConclusionPARP14 induces AML cell growth and glycolysis by activating NF-κB and promoting HIF-1α expression, which may suggest new insights into the pathogenesis of AML.

Anti-Growth, Anti-Angiogenic, and Pro-Apoptotic Effects by CX-4945, an Inhibitor of Casein Kinase 2, on HuCCT-1 Human Cholangiocarcinoma Cells via Control of Caspase-9/3, DR-4, STAT-3/STAT-5, Mcl-1, eIF-2α, and HIF-1α.

Overexpression of casein kinase 2 (CK2) has an oncogenic and pro-survival role in many cancers. CX-4945 (Silmitasertib) is a CK2 inhibitor with anti-cancerous and anti-angiogenic effects. Up to date, the anti-cancer effect and mechanism of CX-4945 on human cholangiocarcinoma (CCA) remain unclear. This study investigated whether CX-4945 inhibits growth and induces apoptosis of HuCCT-1 cells, a human CCA cell line. Of note, treatment with CX-4945 at 20 μM markedly reduced survival and induced apoptosis of HuCCT-1 cells, as evidenced by nuclear DNA fragmentation, PARP cleavage, activation of caspase-9/3, and up-regulation of DR-4. Although CX-4945 did not affect the phosphorylation and expression of CK2, it vastly inhibited the phosphorylation of CK2 substrates, supporting the drug’s efficacy in inhibiting CK2 and its downstream pathway. Importantly, knockdown of CK2 that partially suppressed the phosphorylation of CK2 substrates resulted in a significant reduction of HuCCT-1 cell survival. In addition, CX-4945 reduced the phosphorylation and expression of STAT-3 and STAT-5 in HuCCT-1 cells, and pharmacological inhibition or respective knockdown of these proteins resulted in significant growth suppression of HuCCT-1 cells. CX-4945 also had abilities to decrease Mcl-1 expression while increasing eIF-2α phosphorylation in HuCCT-1 cells. Furthermore, there was a time-differential negative regulation of HIF-1α expression by CX-4945 in HuCCT-1 cells, and knockdown of HIF-1α caused a significant reduction of the cell survival. In summary, these results demonstrated that CX-4945 has anti-growth, anti-angiogenic, and pro-apoptotic effects on HuCCT-1 cells, which are mediated through control of CK2, caspase-9/3, DR-4, STAT-3/5, Mcl-1, eIF-2α, and HIF-1α.

LAT1-specific inhibitor ameliorates severe autoimmune arthritis in SKG mouse

L-type amino acid transporter 1 (LAT1, slc7a5) supplies large neutral amino acids to highly proliferative cells. LAT1 is an attractive therapeutic target for treating overactive T cell-mediated immune disorders due to its high expression in activated T cells, but not in resting T cells. Here, we demonstrate that LAT1 plays a crucial role in T helper (Th) 17-mediated autoimmune arthritis in SKG mice, an animal model of human rheumatoid arthritis (RA). Administration of JPH203, a LAT1-specific inhibitor, suppressed mannan-induced joint swelling, synoviocyte proliferation and inflammatory cell infiltration in SKG mice. A diminished metabolic reprogramming, including a decrease in oxidative phosphorylation that regulates Hif-1α expression and subsequent control of glycolysis enzymes, was involved in the downregulation of Th17 differentiation by LAT1 inhibition. Moreover, publicly released database analysis revealed facilitated expression of LAT1 in T cells with cytotoxic features in patients with RA. Our results demonstrate the essential contribution of LAT1 to the development of RA, proposing a potential therapeutic approach targeting amino acid transporters for treating hypersensitive immune diseases.

Methylsulfonylmethane relieves cobalt chloride-induced hypoxic toxicity in C2C12 myoblasts

AimsIn biology and medicine, hypoxia refers to reduced oxygen tension or oxygen starvation resulting from various environmental or pathological conditions. Prolonged hypoxia may lead to an imbalance in protein production and a loss of muscle mass in animals. The physiological response to hypoxia includes oxidative stress-induced activation of complex cell-signaling networks such as hypoxia-inducible factor (HIF), phosphoinositide 3-kinase (PI3K), and Janus kinase/signal transducer and activator of transcription (JAK-STAT). Methylsulfonylmethane (MSM) is a natural sulfur compound that regulates HIF-1α expression and provides cytoprotection from oxidative stress. In this study, we explored the anti-hypoxic activity and cytoprotective effect of MSM in cobalt chloride (CoCl2)-induced hypoxic C2C12 mouse myoblast culture. Materials and methodsWe used western blotting, real time PCR, flow cytometry for molecular signaling studies and we also used MTT assay and ChIP assay along with comet assay for cellular processes. Key findingsMSM prevented the CoCl2 induced cytotoxicity. Molecular markers of hypoxia, induced by CoCl2, were normalized or reduced by MSM, which also inhibited the effect of CoCl2-induced JAK2/STAT5b/Cyclin D1 and PI3K/AKT signaling. CoCl2-induced oxidative stress results in activation of the NRF2/HO-1-mediated cell survival pathway and inhibition of DNA repair, both of which were prevented by MSM. SignificanceWe suggest MSM can be considered as a candidate drug for reducing the effects of hypoxia in both animals and humans.

Downregulation of the long non-coding RNA MALAT1 in tenofovir-treated pregnant women with hepatitis B virus infection promotes immune recovery of natural killer cells via the has-miR-155-5p/HIF-1α axis

BackgroundCurrently, tenofovir disoproxil fumarate (TDF) treatment in pregnant women with hepatitis B virus (HBV) infection in the second trimester of pregnancy to reduce the HBV DNA load and block the mother-to-child transmission of HBV has become a consensus. An increasing number of studies have shown that lncRNAs play an important role in immune regulation; however, there are only a few studies on how lncRNAs affect the immune function of TDF-treated pregnant women with HBV infection. MethodsThe peripheral blood of pregnant women with HBV infection was collected before and after TDF treatment for whole-transcriptome sequencing; the differential expression of lncRNA MALAT1 in PBMCs and natural killer (NK) cells was verified by qRT-PCR. ELISA and flow cytometry were used to detect the effect of MALAT1 on NK-92 cells on IFN-γ secretion. Dual-luciferase reporter, qRT-PCR, and western blot analyses were used to verify the relationship between the expression levels of MALAT1, has-miR-155-5p and HIF-1α. ResultsAfter TDF treatment, the MALAT1 expression in the PBMCs of pregnant women with HBV infection, especially in NK cells, was significantly reduced. MALAT1 overexpression decreased IFN-γ secretion in NK-92 cells, while IFN-γ secretion increased after MALAT1 knockdown. Mechanistically, we identified MALAT1 as a competitive endogenous RNA that regulates HIF-1α expression by sponging has-miR-155-5p. Low HIF-1α expression resulted in increased IFNG expression in, and IFN-γ secretion by, NK cells. ConclusionsThus, MALAT1 may play an important role in NK cell-mediated immunity in TDF-treated pregnant women with HBV infection by regulating the has-miR-155-5p/HIF-1α axis.

The Regulation of Hypoxia Inducible Factor (HIF)1α Expression by Quercetin: an In Silico Study

Background:Cancer disease is a growing health problem in developing and developed countries. Hypoxia-inducible factor-1a (HIF1α) is a transcription factor responsible for expressing several proteins involved in angiogenesis. Quercetin can suppress HIF1α expression due to the inhibition of protein synthesis. However, to date, the study exploring the potential of quercetin in repressing HIF1α through its degradation mechanism has never been done. An in silico study is needed as a preliminary study to understand the mechanism underlining this possibility.Objective:This study aimed to investigate the potential of quercetin in regulating HIF1α expression through the ubiquitin degradation pathway by in silico study.Methods:This study was performed by in silico analysis, including biological activity prediction, 3D protein structure collection, protein-ligand and protein-protein docking, and the visualization of the docking results.Results:The probability activity (Pa) score of quercetin as an HIF1α expression inhibitor was 0.969. In the absence of quercetin, the center-weighted score of HIF1α - pVHL, HIF1α - FIH, and HIF1α - PHD2 was -699.4 kJ/mol, -846.0 kJ/mol, and -650.5 kJ/mol, respectively. In the presence of quercetin, the weighted score of HIF1α - pVHL, HIF1α - FIH, and HIF1α - PHD2 was reduced to -728.1 kJ/mol, -854.2 kJ/mol, and -650.5 kJ/mol, respectively.Conclusion:Quercetin could directly promote HIF1α and pVHL interaction, thus increasing the degradation of HIF1α by ubiquitin-dependent pathway.

HIF-1\u03b1-activated long non-coding RNA KDM4A-AS1 promotes hepatocellular carcinoma progression via the miR-411-5p/KPNA2/AKT pathway

Hepatocellular carcinoma (HCC) is the most common type of liver cancer with poor clinical outcomes. Long non-coding RNAs (lncRNAs) are extensively involved in the tumorigenesis and progression of HCC. However, more investigations should be carried out on novel lncRNAs and their effects on HCC. Here we identified a novel lncRNA KDM4A-AS1, which was aberrantly overexpressed in HCC tissues, associated with unfavorable clinical features and poor prognosis of patients. KDM4A-AS1 promoted HCC cell proliferation, migration, and invasion in vitro and contributed to HCC growth and lung metastasis in vivo. Mechanistically, KDM4A-AS1 was inversely modulated by miR-411-5p at the post-transcriptional level and facilitated Karyopherin α2 (KPNA2) expression by competitively binding miR-411-5p, thereby activating the AKT pathway. KPNA2 silencing, miR-411-5p overexpression, and AKT inhibitor (MK2206) consistently reversed KDM4A-AS1-enhanced proliferation, mobility, and EMT of HCC cells. KDM4A-AS1 was identified as a novel hypoxia-responsive gene and transactivated by hypoxia-inducible factor 1α (HIF-1α) in HCC cells. In turn, KDM4A-AS1 regulated HIF-1α expression through the KPNA2/AKT signaling pathway. Hence, this study revealed a novel hypoxia-responsive lncRNA, KDM4A-AS1, which contributed to HCC growth and metastasis via the KDM4A-AS1/KPNA2/HIF-1α signaling loop. Our findings provide a promising prognostic and therapeutic target for HCC.