RESUMEN
A short peptide termed NEMO-binding domain (NBD) peptide has an inhibitory effect on nuclear factor kappa-B (NF-κB). Despite its efficacy in inhibiting inflammatory responses, the precise neuroprotective mechanisms of NBD peptide in spinal cord injury (SCI) remain unclear. This study aims to determine whether the pyroptosis-related aspects involved in the neuroprotective effects of NBD peptide post-SCI.Using RNA sequencing, the molecular mechanisms of NBD peptide in SCI are explored. The evaluation of functional recovery is performed using the Basso mouse scale, Nissl staining, footprint analysis, Masson's trichrome staining, and HE staining. Western blotting, enzyme-linked immunosorbent assays, and immunofluorescence assays are used to examine pyroptosis, autophagy, lysosomal membrane permeabilization (LMP), acid sphingomyelinase (ASMase), and the NF-κB/p38-MAPK related signaling pathway.NBD peptide mitigated glial scar formation, reduced motor neuron death, and enhanced functional recovery in SCI mice. Additionally, NBD peptide inhibits pyroptosis, ameliorate LMP-induced autophagy flux disorder in neuron post-SCI. Mechanistically, NBD peptide alleviates LMP and subsequently enhances autophagy by inhibiting ASMase through the NF-κB/p38-MAPK/Elk-1/Egr-1 signaling cascade, thereby mitigating neuronal death. NBD peptide contributes to functional restoration by suppressing ASMase-mediated LMP and autophagy depression, and inhibiting pyroptosis in neuron following SCI, which may have potential clinical application value.
RESUMEN
Receptor-binding peptides are promising candidates for tumor target therapy. However, the inability to occupy "hot spots" on the PPI interface and rapid metabolic instability are significant limitations to their clinical application. We investigated a new strategy in which an FGFR1-binding peptide (Pep1) was site-specifically functionalized with the dinitrophenyl (DNP) hapten at the C-terminus. The resulting Pep1-DNP conjugates retained FGFR1 binding affinity and exhibited a similar potency in inhibiting FGF2-dependent cell proliferation, comparable to that of native Pep1 in vitro. In addition, three conjugates could recruit anti-DNP antibodies onto the surface of cancer cells, thereby mediating the CDC efficacy. In vivo pharmacokinetic studies and antitumor studies demonstrated that optimal conjugate 9 exhibited significantly prolonged half-lives and improved antitumor efficacy without prominent toxicity compared to those of native Pep1. This is a general and cost-effective approach for generating peptidomimetic immunotherapeutics with multiple antitumor mechanisms that may have broad applications in cancer therapy.
RESUMEN
Proviral Integrations of Moloney-2 (PIM-2) kinase is a promising target for various cancers and other diseases, and its inhibitors hold potential for treating related diseases. However, there is currently no clinically available PIM-2 inhibitor. In this study, we constructed a generative model for de novo PIM-2 inhibitor design based on artificial intelligence, performed molecular docking and molecular dynamics (MD) simulations to develop an efficient PIM-2 inhibitor generative model and discover potential PIM-2 inhibitors. First, we designed a generative model based on a Bi-directional Long Short-Term Memory (BiLSTM) framework combined with a transfer learning strategy and generated a new PIM-2 small molecule library using existing active drug databases. The generated compound library was then virtually screened by molecular docking and scaffold similarity comparison, identifying 10 initial hit compounds with better performance. Next, using the inhibitor in the crystal structure as a positive control, we performed two rounds of MD simulations, with lengths of 100 ns and 500 ns, respectively, to study the dynamic stability of the protein-ligand systems of the 10 compounds with PIM-2. Analyzed the interactions with key hinge region residues, binding free energies, and changes in the ATP pocket size. The generative model demonstrates good molecular generation capability and can generate efficient novel molecules with similar physicochemical properties as active PIM-2 drugs. Among the 10 initially selected hit compounds, 5 compounds C3 (- 29.69 kcal/mol), C4 (- 33.31 kcal/mol), C5 (- 28.59 kcal/mol), C8 (- 34.68 kcal/mol), and C9 (- 25.88 kcal/mol) have higher binding energies with PIM-2 than the positive drug 3YR (- 26.18 kcal/mol). The MD simulation results are consistent with the docking analysis, these compounds have lower and more stable RMSD values for the complex systems with the reported positive drug 3YR and PIM-2 complex system. They can form long-term stable interactions with active site and the hinge region of PIM-2, which suggests these compounds are likely to have potent inhibitory effects on PIM-2. This study provides an efficient generative model for PIM-2 inhibitor research and discovers 5 potential novel PIM-2 inhibitors.
RESUMEN
Background: Ensuring the survival of the distal end of a random flap during hypoperfusion (ischaemia) is difficult in clinical practice. Effective prevention of programmed cell death is a potential strategy for inhibiting ischaemic flap necrosis. The activation of stimulator of interferon genes (STING) pathway promotes inflammation and leads to cell death. The epidermal growth factor family member neuregulin-1 (NRG1) reduces cell death by activating the protein kinase B (AKT) signalling pathway. Moreover, AKT signalling negatively regulates STING activity. We aimed to verify the efficacy of NRG1 injection in protecting against flap necrosis. Additionally, we investigated whether NRG1 effectively enhances ischemic flap survival by inhibiting pyroptosis and necroptosis through STING suppression. Methods: A random-pattern skin flap model was generated on the backs of C57BL/6 mice. The skin flap survival area was determined. The blood supply and vascular network of the flap was assessed by laser Doppler blood flow analysis. Cluster of differentiation 34 immunohistochemistry (IHC) and haematoxylin and eosin (H&E) staining of the flap sections revealed microvessels. Transcriptome sequencing analysis revealed the mechanism by which NRG1 promotes the survival of ischaemic flaps. The levels of angiogenesis, oxidative stress, necroptosis, pyroptosis and indicators associated with signalling pathways in flaps were examined by IHC, immunofluorescence and Western blotting. Packaging adeno-associated virus (AAV) was used to activate STING in flaps. Results: NRG1 promoted the survival of ischaemic flaps. An increased subcutaneous vascular network and neovascularization were found in ischaemic flaps after the application of NRG1. Transcriptomic gene ontology enrichment analysis and protein level detection indicated that necroptosis, pyroptosis and STING activity were reduced in the NRG1 group. The phosphorylation of AKT and forkhead box O3a (FOXO3a) were increased after NRG1 treatment. The increased expression of STING in flaps induced by AAV reversed the therapeutic effect of NRG1. The ability of NRG1 to phosphorylate AKT-FOXO3a, inhibit STING and promote flap survival was abolished after the application of the AKT inhibitor MK2206. Conclusions: NRG1 inhibits pyroptosis and necroptosis by activating the AKT-FOXO3a signalling pathway to suppress STING activation and promote ischaemic flap survival.
RESUMEN
Preventing and treating avascular necrosis at the distal end of the flaps are critical to surgery success, but current treatments are not ideal. A recent study shows that apoptotic bodies (ABs) generated near the site of apoptosis can be taken up and promote cell proliferation. The study reveals that ABs derived from fibroblast-like cells in the subcutaneous connective tissue (FSCT cells) of skin flaps promoted ischaemic flap survival. It is also found that ABs inhibited cell death and oxidative stress and promoted M1-to-M2 polarization in macrophages. Transcriptome sequencing and protein level testing demonstrated that ABs promoted ischaemic flap survival in endothelial cells and macrophages by inhibiting ferroptosis via the KEAP1-Nrf2 axis. Furthermore, microRNA (miR) sequencing data and in vitro and in vivo experiments demonstrated that ABs inhibited KEAP1 by delivering miR-339-5p to exert therapeutic effects. In conclusion, FSCT cell-derived ABs inhibited ferroptosis, promoted the macrophage M1-to-M2 transition via the miR-339-5p/KEAP1/Nrf2 axis and promoted ischaemic flap survival. These results provide a potential therapeutic strategy to promote ischaemic flap survival by administering ABs.
Asunto(s)
Ferroptosis , Fibroblastos , Proteína 1 Asociada A ECH Tipo Kelch , MicroARNs , Factor 2 Relacionado con NF-E2 , Colgajos Quirúrgicos , Animales , Ratones , Proteína 1 Asociada A ECH Tipo Kelch/metabolismo , Proteína 1 Asociada A ECH Tipo Kelch/genética , Factor 2 Relacionado con NF-E2/metabolismo , Factor 2 Relacionado con NF-E2/genética , Ferroptosis/genética , MicroARNs/genética , MicroARNs/metabolismo , Fibroblastos/metabolismo , Modelos Animales de Enfermedad , Isquemia/metabolismo , Isquemia/genética , Masculino , Apoptosis/genética , Tejido Conectivo/metabolismo , Transducción de Señal/genéticaRESUMEN
Excessive activation of FGF19/fibroblast growth factor receptor 4 (FGFR4) signaling is associated with poor survival of patients with hepatocellular carcinoma (HCC). FGFR4 inhibitors show promise for HCC treatment. F30, an indazole derivative designed through computer-aided drug design targeting FGFR4, demonstrated anti-HCC activity as described in our previous studies. However, the precise molecular mechanisms underlying F30's anticancer effects remain largely unexplored. We report here that F30 could effectively induce ferroptosis in HCC cells. The concentrations of cellular ferrous iron, the peroxidation of cell membranes and the homeostasis of reduced glutathione (GSH)/oxidized glutathione disulfide (GSSG) were dysregulated by F30, thereby affecting cellular redox status. Induction of ferroptosis in HCC by F30 was inhibited by specific ferroptosis inhibitor ferrostatin-1. F30 upregulates various ferroptosis-related genes, including the heme oxygenase enzymes 1 (HMOX1), a key mediator of redox regulation. Surprisingly, F30-induced ferroptosis in HCC is dependent on HMOX1. The dysregulation of cellular ferrous iron concentrations and cell membrane peroxidation was rescued when knocking down HMOX1 with specific small interfering RNA. These findings shed light on the molecular mechanisms underlying FGFR4-targeting F30's anti-HCC effects and suggest that FGFR4 inactivation could be beneficial for HCC treatment involving ferroptosis.
Asunto(s)
Carcinoma Hepatocelular , Ferroptosis , Neoplasias Hepáticas , Humanos , Carcinoma Hepatocelular/genética , Neoplasias Hepáticas/genética , Receptor Tipo 4 de Factor de Crecimiento de Fibroblastos/genética , Receptor Tipo 4 de Factor de Crecimiento de Fibroblastos/metabolismo , Línea Celular Tumoral , Proliferación Celular , Hierro , Hemo-Oxigenasa 1RESUMEN
BACKGROUND AND PURPOSE: Autophagy is a protective factor for controlling neuronal damage, while necroptosis promotes neuroinflammation after spinal cord injury (SCI). DADLE (D-Ala2 , D-Leu5 ]-enkephalin) is a selective agonist for delta (δ) opioid receptor and has been identified as a promising drug for neuroprotection. The aim of this study was to investigate the mechanism/s by which DADLE causes locomotor recovery following SCI. EXPERIMENTAL APPROACH: Spinal cord contusion model was used and DADLE was given by i.p. (16 mg·kg-1 ) in mice for following experiments. Motor function was assessed by footprint and Basso mouse scale (BMS) score analysis. Western blotting used to evaluate related protein expression. Immunofluorescence showed the protein expression in each cell and its distribution. Network pharmacology analysis was used to find the related signalling pathways. KEY RESULTS: DADLE promoted functional recovery after SCI. In SCI model of mice, DADLE significantly increased autophagic flux and inhibited necroptosis. Concurrently, DADLE restored autophagic flux by decreasing lysosomal membrane permeabilization (LMP). Additionally, chloroquine administration reversed the protective effect of DADLE to inhibit necroptosis. Further analysis showed that DADLE decreased phosphorylated cPLA2 , overexpression of cPLA2 partially reversed DADLE inhibitory effect on LMP and necroptosis, as well as the promotion autophagy. Finally, AMPK/SIRT1/p38 pathway regulating cPLA2 is involved in the action DADLE on SCI and naltrindole inhibited DADLE action on δ receptor and on AMPK signalling pathway. CONCLUSION AND IMPLICATION: DADLE causes its neuroprotective effects on SCI by promoting autophagic flux and inhibiting necroptosis by decreasing LMP via activating δ receptor/AMPK/SIRT1/p38/cPLA2 pathway.
Asunto(s)
Leucina Encefalina-2-Alanina , Traumatismos de la Médula Espinal , Animales , Ratones , Proteínas Quinasas Activadas por AMP/metabolismo , Leucina Encefalina-2-Alanina/metabolismo , Leucina Encefalina-2-Alanina/farmacología , Lisosomas/metabolismo , Fosfolipasas/metabolismo , Receptores Opioides delta/metabolismo , Recuperación de la Función , Sirtuina 1/metabolismo , Médula Espinal/metabolismo , Traumatismos de la Médula Espinal/tratamiento farmacológico , Traumatismos de la Médula Espinal/metabolismoRESUMEN
Spinal cord injury (SCI) is a severe medical condition with lasting effects. The efficacy of numerous clinical treatments is hampered by the intricate pathophysiological mechanism of SCI. Fibroblast growth factor 18 (FGF-18) has been found to exert neuroprotective effects after brain ischaemia, but its effect after SCI has not been well explored. The aim of the present study was to explore the therapeutic effect of FGF-18 on SCI and the related mechanism. In the present study, a mouse model of SCI was used, and the results showed that FGF-18 may significantly affect functional recovery. The present findings demonstrated that FGF-18 directly promoted functional recovery by increasing autophagy and decreasing pyroptosis. In addition, FGF-18 increased autophagy, and the well-known autophagy inhibitor 3-methyladenine (3MA) reversed the therapeutic benefits of FGF-18 after SCI, suggesting that autophagy mediates the therapeutic effects of FGF-18 on SCI. A mechanistic study revealed that after stimulation of the protein kinase B (AKT)-transient receptor potential mucolipin 1 (TRPML1)-calcineurin signalling pathway, the FGF-18-induced increase in autophagy was mediated by the dephosphorylation and nuclear translocation of transcription factor E3 (TFE3). Together, these findings indicated that FGF-18 is a robust autophagy modulator capable of accelerating functional recovery after SCI, suggesting that it may be a promising treatment for SCI in the clinic.
Asunto(s)
Factores de Crecimiento de Fibroblastos , Proteínas Proto-Oncogénicas c-akt , Traumatismos de la Médula Espinal , Ratas , Ratones , Animales , Proteínas Proto-Oncogénicas c-akt/metabolismo , Piroptosis , Ratas Sprague-Dawley , Serina-Treonina Quinasas TOR/metabolismo , Médula Espinal/metabolismo , Traumatismos de la Médula Espinal/metabolismo , AutofagiaRESUMEN
BACKGROUND AND PURPOSE: Ischaemia-reperfusion (I/R) injury is a major contributor to skin flap necrosis, which presents a challenge in achieving satisfactory therapeutic outcomes. Previous studies showed that cathelicidin-BF (BF-30) protects tissues from I/R injury. In this investigation, BF-30 was synthesized and its role and mechanism in promoting survival of I/R-injured skin flaps explored. EXPERIMENTAL APPROACH: Survival rate analysis and laser Doppler blood flow analysis were used to evaluate I/R-injured flap viability. Western blotting, immunofluorescence, TdT-mediated dUTP nick end labelling (TUNEL) and dihydroethidium were utilized to examine the levels of apoptosis, pyroptosis, oxidative stress, transcription factor EB (TFEB)-mediated autophagy and molecules related to the adenosine 5'-monophosphate-activated protein kinase (AMPK)-transient receptor potential mucolipin 1 (TRPML1)-calcineurin signalling pathway. KEY RESULTS: The outcomes revealed that BF-30 enhanced I/R-injured island skin flap viability. Autophagy, oxidative stress, pyroptosis and apoptosis were related to the BF-30 capability to enhance I/R-injured flap survival. Improved autophagy flux and tolerance to oxidative stress promoted the inhibition of apoptosis and pyroptosis in vascular endothelial cells. Activation of TFEB increased autophagy and inhibited endothelial cell oxidative stress in I/R-injured flaps. A reduction in TFEB level led to a loss of the protective effect of BF-30, by reducing autophagy flux and increasing the accumulation of reactive oxygen species (ROS) in endothelial cells. Additionally, BF-30 modulated TFEB activity via the AMPK-TRPML1-calcineurin signalling pathway. CONCLUSION AND IMPLICATIONS: BF-30 promotes I/R-injured skin flap survival by TFEB-mediated up-regulation of autophagy and inhibition of oxidative stress, which may have possible clinical applications.
Asunto(s)
Piroptosis , Daño por Reperfusión , Humanos , Especies Reactivas de Oxígeno/metabolismo , Catelicidinas/farmacología , Proteínas Quinasas Activadas por AMP/metabolismo , Células Endoteliales/metabolismo , Calcineurina/farmacología , Autofagia , Daño por Reperfusión/metabolismo , Factores de TranscripciónRESUMEN
We have developed an efficient and green strategy for the synthesis of C2-amino indolin-3-ones and C2-acyloxy indolin-3-ones via KI-catalyzed C(sp3)-H amination and acyloxylation of indolin-3-ones using air as the oxidant. The reaction provides straightforward access to 2-substituted indolin-3-ones by the direct functionalization of indolin-3-ones at the C2 position under mild conditions. Moreover, the conditions enable direct functionalization of a range of complex pharmaceuticals, providing attractive products for medicinal chemistry programs.
RESUMEN
BACKGROUND: CCN6 is a matricellular protein that critically regulates the tumourigenesis and progression of breast cancer. Although the tumour-suppressive function of CCN6 has been extensively studied, molecular mechanisms regulating protein levels of CCN6 remain largely unclear. This study aims to investigate the regulation of CCN6 by ubiquitination and deubiquitinating enzymes (DUBs) in breast cancer. METHODS: A screening assay was performed to identify OTUB1 as the DUB for CCN6. Various biochemical methods were applied to elucidate the molecular mechanism of OTUB1 in the regulation of CCN6. The role of OTUB1-CCN6 interaction in breast cancer was studied with cell experiments and the allograft model. The correlation of OTUB1 and CCN6 in human breast cancer was determined by immunohistochemistry and Western blot. RESULTS: We found that CCN6 protein levels were controlled by the ubiquitin-proteasome system. The K48 ubiquitination and degradation of CCN6 was inhibited by OTUB1, which directly interacted with CCN6 through its linker domain. Furthermore, OTUB1 inhibited the ubiquitination of CCN6 in a non-canonical manner. Deletion of OTUB1, concomitant with reduced CCN6 abundance, increased the migration, proliferation and viability of breast cancer cells. Supplementation of CCN6 abolished the effect of OTUB1 deletion on breast cancer. Importantly, OTUB1 expression was downregulated in human breast cancer and positively correlated with CCN6 levels. CONCLUSION: This study identified OTUB1 as a novel regulator of CCN6 in breast cancer.
Asunto(s)
Proteínas CCN de Señalización Intercelular , Carcinogénesis , Transformación Celular Neoplásica , Enzimas Desubicuitinizantes , Humanos , Western Blotting , Citoplasma , Complejo de la Endopetidasa Proteasomal , Enzimas Desubicuitinizantes/metabolismo , Proteínas CCN de Señalización Intercelular/metabolismoRESUMEN
BACKGROUND: Necroptosis and pyroptosis, two types of proinflammatory programmed cell death, were recently found to play important roles in spinal cord injury (SCI). Moreover, cyclic helix B peptide (CHBP) was designed to maintain erythropoietin (EPO) activity and protect tissue against the adverse effects of EPO. However, the protective mechanism of CHBP following SCI is still unknown. This research explored the necroptosis- and pyroptosis-related mechanism underlying the neuroprotective effect of CHBP after SCI. METHODS: Gene Expression Omnibus (GEO) datasets and RNA sequencing were used to identify the molecular mechanisms of CHBP for SCI. A mouse model of contusion SCI was constructed, and HE staining, Nissl staining, Masson staining, footprint analysis and the Basso Mouse Scale (BMS) were applied for histological and behavioural analyses. qPCR, Western blot analysis, immunoprecipitation and immunofluorescence were utilized to analyse the levels of necroptosis, pyroptosis, autophagy and molecules associated with the AMPK signalling pathway. RESULTS: The results revealed that CHBP significantly improved functional restoration, elevated autophagy, suppressed pyroptosis, and mitigated necroptosis after SCI. 3-Methyladenine (3-MA), an autophagy inhibitor, attenuated these beneficial effects of CHBP. Furthermore, CHBP-triggered elevation of autophagy was mediated by the dephosphorylation and nuclear translocation of TFEB, and this effect was due to stimulation of the AMPK-FOXO3a-SPK2-CARM1 and AMPK-mTOR signalling pathways. CONCLUSION: CHBP acts as a powerful regulator of autophagy that improves functional recovery by alleviating proinflammatory cell death after SCI and thus might be a prospective therapeutic agent for clinical application.
Asunto(s)
Péptidos Cíclicos , Traumatismos de la Médula Espinal , Ratones , Animales , Péptidos Cíclicos/farmacología , Péptidos Cíclicos/uso terapéutico , Proteínas Quinasas Activadas por AMP/metabolismo , Traumatismos de la Médula Espinal/tratamiento farmacológico , Traumatismos de la Médula Espinal/genética , Traumatismos de la Médula Espinal/metabolismo , Apoptosis , Transducción de Señal , AutofagiaRESUMEN
A copper-catalyzed C3 amination of 2H-indazoles with 2H-indazoles and indazol-3(2H)-ones under mild conditions was developed. A series of indazole-containing indazol-3(2H)-one derivatives were produced in moderate to excellent yields. The mechanistic studies suggest that the reactions probably proceed through a radical pathway.
RESUMEN
Background: Caloric restriction mimetics (CRMs) mimic the favourable effects of caloric restriction (CR) and have been shown to have therapeutic effects in neuroinflammatory disease. However, whether CRMs improve the functional recovery from spinal cord injury (SCI) and the underlying mechanism of action remain unclear. In this study, we used the CRMs 3,4-dimethoxychalcone (3,4-DC) to evaluate the therapeutic value of CRMs for SCI. Methods: HE, Masson and Nissl staining; footprint analysis; and the Basso mouse scale were used to determine the functional recovery from SCI after 3,4-DC treatment. RNA sequencing was used to identify the mechanisms of 3,4-DC in SCI. Western blotting, qPCR and immunofluorescence were used to detect the levels of pyroptosis, necroptosis, autophagy and the AMPK-TRPML1-calcineurin signalling pathway. We employed a dual-luciferase reporter assay in vitro and applied AAV vectors to inhibit TFEB in vivo to explore the mechanism of 3,4-DC. Results: 3,4-DC reduced glial scar area and motor neuron death and improved functional recovery after SCI. RNA-sequencing results indicated that oxidative stress, pyroptosis, necroptosis, and autophagy may be involved in the ability of 3,4-DC to improve functional recovery. Furthermore, 3,4-DC inhibited pyroptosis and necroptosis by enhancing autophagy. We also found that 3,4-DC enhances autophagy by promoting TFEB activity. A decrease in the TFEB level abolished the protective effect of 3,4-DC. In addition, 3,4-DC partially regulated TFEB activity through the AMPK-TRPML1-calcineurin signalling pathway. Conclusions: 3,4-DC promotes functional recovery by upregulating TFEB-mediated autophagy and inhibiting pyroptosis and necroptosis after SCI, which may have potential clinical application value.
Asunto(s)
Restricción Calórica , Necroptosis , Piroptosis , Traumatismos de la Médula Espinal , Animales , Ratones , Proteínas Quinasas Activadas por AMP/metabolismo , Autofagia , Calcineurina/metabolismo , Necroptosis/efectos de los fármacos , Piroptosis/efectos de los fármacos , Traumatismos de la Médula Espinal/tratamiento farmacológico , Traumatismos de la Médula Espinal/patologíaRESUMEN
Background: Increasing evidence suggests that acute traumatic spinal cord injury (SCI)-induced defects in autophagy and autophagy-lysosomal pathway (ALP) may contribute to endothelial barrier disruption following injury. Recently, Kruppel-like factor 2 (KLF2) was reported as a key molecular switch on regulating autophagy. Whether KLF2 coordinates endothelial endothelial ALP in SCI is not known. Methods: Genetic manipulations of KLF2 were performed in bEnd.3 cells and SCI model. Western blot, qRT-PCR, immunofluorescence staining and Lyso-Tracker Red staining, Evans blue dye extravasation, behavioral assessment via Basso mouse scale (BMS), electrophysiology and footprint analysis were performed. Results: In SCI, autophagy flux disruption in endothelial cells contributes to TJ proteins degradation, leading to blood-spinal cord barrier (BSCB) impairment. Furthermore, the KLF2 level was decreased in SCI, overexpression of which alleviated TJ proteins loss and BSCB damage, which improve motor function recovery in SCI mice, while knockdown of KLF2 displayed the opposite effects. At the molecular level, KLF2 overexpression alleviated the TJ proteins degradation and the endothelial permeability by tuning the ALP dysfunction caused by SCI and oxygen glucose deprivation (OGD). Conclusions: Endothelial KLF2 as one of the key contributors to SCI-mediated ALP dysfunction and BSCB disruption. KLF2 could be a promising pharmacological target for the management and treatment of SCI.
Asunto(s)
Autofagia , Barrera Hematoencefálica , Factores de Transcripción de Tipo Kruppel , Traumatismos de la Médula Espinal , Animales , Ratones , Barrera Hematoencefálica/metabolismo , Células Endoteliales/metabolismo , Factores de Transcripción de Tipo Kruppel/genética , Factores de Transcripción de Tipo Kruppel/metabolismo , Recuperación de la Función , Traumatismos de la Médula Espinal/metabolismo , Factores de Transcripción/metabolismoRESUMEN
Mitochondrial dynamics, including mitochondrial fission and fusion, are critical for maintaining mitochondrial functions. Evidence shows that TANK-binding kinase 1 (TBK1) regulates mitochondrial fusion and fission and then mitophagy. Since a previous study demonstrates a strong correlation between mitophagy and osteoarthritis (OA), we herein investigated the potential role of TBK1 in OA process and mitochondrial functions. We demonstrated a strong correlation between TBK1 and OA, evidenced by significantly downregulated expression of TBK1 in cartilage tissue samples of OA patients and in the chondrocytes of aged mice, as well as TNF-α-stimulated phosphorylation of TBK1 in primary mouse chondrocytes. TBK1 overexpression significantly attenuated TNF-α-induced apoptosis and abnormal mitochondrial function in primary mouse chondrocytes. Furthermore, TBK1 overexpression induced remodeling of mitochondrial morphology by directly phosphorylating dynamin-related protein 1 (DRP1) at Ser637, abolishing the fission of DRP1 and preventing its fragmentation function. Moreover, TBK1 recruitment and DRP1 phosphorylation at Ser637 was necessary for engulfing damaged mitochondria by autophagosomal membranes during mitophagy. Moreover, we demonstrated that APMK/ULK1 signaling contributed to TBK1 activation. In OA mouse models established by surgical destabilization of the medial meniscus, intraarticular injection of lentivirus-TBK1 significantly ameliorated cartilage degradation via regulation of autophagy and alleviation of cell apoptosis. In conclusion, our results suggest that the TBK1/DRP1 pathway is involved in OA and pharmacological targeting of the TBK1-DRP1 cascade provides prospective therapeutic benefits for the treatment of OA.
Asunto(s)
Dinámicas Mitocondriales , Factor de Necrosis Tumoral alfa , Ratones , Animales , Fosforilación , Factor de Necrosis Tumoral alfa/metabolismo , Autofagia/fisiología , Dinaminas/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismoRESUMEN
The 4-Aminopyrazolopyrimidine scaffold has been an interesting pharmacophore since the disclosure of the intimate connection between small-molecule inhibitors and the treatment of diseases. Modification of the 4-aminopyrazolopyrimidine scaffold according to different targets, especially tyrosine kinase and serine/threonine kinase, has resulted in a variety of small-molecule inhibitors. Kinase inhibitors with 4-aminopyrazolopyrimidine derivatives as scaffolds have been widely applied in the treatment of diseases. In this article, we summarized the reports on 4-aminopyrazolopyrimidine as well as its deformation and the application of its derivatives in designing small-molecule inhibitors and the treatment of diseases.
RESUMEN
BACKGROUND: Fibroblast growth factor receptor (FGFR) signaling influenced tumour occurrence and development. Overexpression of FGFR had been observed in many types of cancers, including colon cancer. FGFR inhibitor is considered to be effective in treating colon cancer patients. METHODS: First, the kinase inhibition rate was determined. MTT, western blotting, colony formation, EdU and comet assays were performed to evaluate the anti-tumour effects of F1-7 in vitro. RNA-seq and bioinformatics analysis were used for further verification. Additionally, a xenograft model was generated to investigate the anti-tumour effect of F1-7. RESULTS: F1-7 can inhibit the proliferation of colon cancer cells in vitro. It could significantly inhibit FGFR phosphorylation and its downstream signaling pathway. Whole-genome RNA-seq analysis found that the changed genes were not only functionally focused on MAPK signaling pathway but also related to cell apoptosis and ferroptosis. Experimental evidence demonstrated that F1-7 can directly increase the level of cellular DNA damage. The occurrence of DNA damage led to cell cycle arrest and inhibition of cell metastasis and cell apoptosis. Mouse model experiments also confirmed that F1-7 could inhibit tumour growth by inhibiting the FGFR pathway. CONCLUSIONS: F1-7 exhibits anti-tumour activity by inhibiting the FGFR pathway. It could be a novel therapeutic agent for targeting colon cancer cells.
Asunto(s)
Neoplasias del Colon , Inhibidores de Proteínas Quinasas , Animales , Muerte Celular , Línea Celular Tumoral , Proliferación Celular , Neoplasias del Colon/tratamiento farmacológico , Neoplasias del Colon/genética , Daño del ADN , Humanos , Ratones , Inhibidores de Proteínas Quinasas/farmacología , Receptores de Factores de Crecimiento de Fibroblastos/genéticaRESUMEN
BACKGROUND AND PURPOSE: Necrosis of random-pattern skin flaps limits their clinical application. Helix B surface peptide (HBSP) protects tissues from ischaemia-reperfusion injury but its short plasma half-life limits its applications. Here, we have synthesized cyclic helix B peptide (CHBP) and investigated its role in flap survival and the underlying mechanisms. EXPERIMENTAL APPROACH: Flap viability was evaluated by survival area analysis, laser Doppler blood flow and histological analysis. RNA sequencing was used to identify mechanisms underlying the effects of CHBP. Levels of autophagy, oxidative stress, pyroptosis, necroptosis and molecules related to the AMP-activated protein kinase (AMPK)-TRPML1-calcineurin signalling pathway were assayed with Western blotting, RT-qPCR, immunohistochemistry and immunofluorescence. KEY RESULTS: The results indicated that CHBP promoted the survival of random-pattern skin flaps. The results of RNA sequencing analysis indicated that autophagy, oxidative stress, pyroptosis and necroptosis were involved in the ability of CHBP to promote skin flap survival. Restoration of autophagy flux and enhanced resistance to oxidative stress contributed to inhibition of pyroptosis and necroptosis. Increased autophagy and inhibition of oxidative stress in the ischaemic flaps were regulated by transcription factor E3 (TFE3). A decrease in the levels of TFE3 caused a reduction in autophagy flux and accumulation of ROS and eliminated the protective effect of CHBP. Moreover, CHBP regulated the activity of TFE3 via the AMPK-TRPML1-calcineurin signalling pathway. CONCLUSION AND IMPLICATIONS: CHBP promotes skin flap survival by up-regulating autophagy and inhibiting oxidative stress in the ischaemic flap and may have potential clinical applications.
Asunto(s)
Proteínas Quinasas Activadas por AMP , Calcineurina , Proteínas Quinasas Activadas por AMP/metabolismo , Autofagia , Factores de Transcripción Básicos con Cremalleras de Leucinas y Motivos Hélice-Asa-Hélice/genética , Factores de Transcripción Básicos con Cremalleras de Leucinas y Motivos Hélice-Asa-Hélice/metabolismo , Factores de Transcripción Básicos con Cremalleras de Leucinas y Motivos Hélice-Asa-Hélice/farmacología , Calcineurina/metabolismo , Calcineurina/farmacología , Péptidos Cíclicos/farmacología , Especies Reactivas de Oxígeno/metabolismoRESUMEN
Myeloid differentiation factor 88 (MyD88) is a hub protein in the Toll-like receptor signaling pathway, which acts as a master switch for numerous inflammatory diseases, including acute lung injury (ALI). Although this protein is considered as a crucial therapeutic target, there are currently no clinically approved MyD88-targeting drugs. Based on previous literature, here we report the discovery via computer-aided drug design (CADD) of a small molecule, M20, which functions as a novel MyD88 inhibitor to efficiently relieve lipopolysaccharide-induced inflammation both in vitro and in vivo. Computational chemistry, surface plasmon resonance detection (SPR) and biological experiments demonstrated that M20 forms an important interaction with the MyD88-Toll/interleukin-1 receptor domain and thereby inhibits the protein dimerization. Taken together, this study found a MyD88 inhibitor, M20, with a novel skeleton, which provides a crucial understanding in the development and modification of MyD88 inhibitors. Meanwhile, the favorable bioactivity of the hit compound is also conducive to the treatment of acute lung injury or other more inflammatory diseases.