Netrin-1 and its receptors play crucial roles in inducing axonal growth and neuronal migration during neuronal development.Their profound impacts then extend into adulthood to encompass the maintenance of neuronal sur...Netrin-1 and its receptors play crucial roles in inducing axonal growth and neuronal migration during neuronal development.Their profound impacts then extend into adulthood to encompass the maintenance of neuronal survival and synaptic function.Increasing amounts of evidence highlight several key points:(1)Diminished Netrin-1 levels exacerbate pathological progression in animal models of Alzheimer’s disease and Parkinson’s disease,and potentially,similar alterations occur in humans.(2)Genetic mutations of Netrin-1 receptors increase an individuals’susceptibility to neurodegenerative disorders.(3)Therapeutic approaches targeting Netrin-1 and its receptors offer the benefits of enhancing memory and motor function.(4)Netrin-1 and its receptors show genetic and epigenetic alterations in a variety of cancers.These findings provide compelling evidence that Netrin-1 and its receptors are crucial targets in neurodegenerative diseases.Through a comprehensive review of Netrin-1 signaling pathways,our objective is to uncover potential therapeutic avenues for neurodegenerative disorders.展开更多
An imbalance in adenosine-mediated signaling,particularly the increased A_(2A)R-mediated signaling,plays a role in the pathogenesis of Parkinson's disease.Existing therapeutic approaches fail to alter disease prog...An imbalance in adenosine-mediated signaling,particularly the increased A_(2A)R-mediated signaling,plays a role in the pathogenesis of Parkinson's disease.Existing therapeutic approaches fail to alter disease progression,demonstrating the need for novel approaches in PD.Repetitive transcranial magnetic stimulation is a non-invasive approach that has been shown to improve motor and non-motor symptoms of Parkinson's disease.However,the underlying mechanisms of the beneficial effects of repetitive transcranial magnetic stimulation remain unknown.The purpose of this study is to investigate the extent to which the beneficial effects of prolonged intermittent theta burst stimulation in the 6-hydroxydopamine model of experimental parkinsonism are based on modulation of adenosine-mediated signaling.Animals with unilateral 6-hydroxydopamine lesions underwent intermittent theta burst stimulation for 3 weeks and were tested for motor skills using the Rotarod test.Immunoblot,quantitative reverse transcription polymerase chain reaction,immunohistochemistry,and biochemical analysis of components of adenosine-mediated signaling were performed on the synaptosomal fraction of the lesioned caudate putamen.Prolonged intermittent theta burst stimulation improved motor symptoms in 6-hydroxydopamine-lesioned animals.A 6-hydroxydopamine lesion resulted in progressive loss of dopaminergic neurons in the caudate putamen.Treatment with intermittent theta burst stimulation began 7 days after the lesion,coinciding with the onset of motor symptoms.After treatment with prolonged intermittent theta burst stimulation,complete motor recovery was observed.This improvement was accompanied by downregulation of the e N/CD73-A_(2A)R pathway and a return to physiological levels of A_(1)R-adenosine deaminase 1 after 3 weeks of intermittent theta burst stimulation.Our results demonstrated that 6-hydroxydopamine-induced degeneration reduced the expression of A_(1)R and elevated the expression of A_(2A)R.Intermittent theta burst stimulation reversed these effects by restoring the abundances of A_(1)R and A_(2A)R to control levels.The shift in ARs expression likely restored the balance between dopamine-adenosine signaling,ultimately leading to the recovery of motor control.展开更多
AIM: To investigate the molecular mechanisms underlying the reversal effect of emodin on platinum resistance in hepatocellular carcinoma. METHODS: After the addition of 10 μmol/L emodin to HepG2/oxaliplatin (OXA) cel...AIM: To investigate the molecular mechanisms underlying the reversal effect of emodin on platinum resistance in hepatocellular carcinoma. METHODS: After the addition of 10 μmol/L emodin to HepG2/oxaliplatin (OXA) cells, the inhibition rate (IR), 50% inhibitory concentration (IC 50 ) and reversal index (IC 50 in experimental group/IC 50 in control group) were calculated. For HepG2, HepG2/OXA, HepG2/OXA/T, each cell line was divided into a control group, OXA group, OXA + fibroblast growth factor 7 (FGF7) group and OXA + emodin group, and the final concentrations of FGF7, emodin and OXA in each group were 5 ng/mL, 10 μg/mL and 10 μmol/L, respectively. Single-cell gel electrophoresis was conducted to detect DNA damage, and the fibroblast growth factor receptor 2 (FGFR2), phosphorylated extracellular signal-regulated kinase 1/2 (p-ERK1/2) and excision repair cross-complementing gene 1 (ERCC1) protein expression levels in each group were examined by Western blotting. RESULTS: Compared with the IC50 of 120.78 μmol/L in HepG2/OXA cells, the IC 50 decreased to 39.65 μmol/L after treatment with 10 μmol/L emodin; thus, the reversal index was 3.05. Compared with the control group, the tail length and Olive tail length in the OXA group, OXA + FGF7 group and OXA + emodin group were significantly increased, and the differences were statistically significant (P < 0.01). The tail length and Olive tail length were lower in the OXA + FGF7 group than in the OXA group, and this difference was also statistically significant. Compared with the OXA + FGF7 group, the tail extent, the Olive tail moment and the percentage of tail DNA were significantly increased in the OXA + emodin group, and these differences were statistically significant (P < 0.01). In comparison with its parental cell line HepG2, the HepG2/OXA cells demonstrated significantly increased FGFR2, p-ERK1/2 and ERCC1 expression levels, whereas the expression of all three molecules was significantly inhibited in HepG2/ OXA/T cells, in which FGFR2 was silenced by FGFR2 shRNA. In the examined HepG2 cells, the FGFR2, p-ERK1/2 and ERCC1 expression levels demonstrated increasing trends in the OXA group and OXA + FGF7 group. Compared with the OXA group and OXA + FGF7 group, the FGFR2, p-ERK1/2, and ERCC1 expression levels were significantly lower in the OXA + emodin group, and these differences were statistically significant. In the HepG2/OXA/T cell line that was transfected with FGFR2 shRNA, the FGFR2, p-ERK1/2 and ERCC1 expression levels were significantly inhibited, but there were no significant differences in these expression levels among the OXA, OXA + FGF7 and OXA + emodin groups. CONCLUSION: Emodin markedly reversed OXA resistance by enhancing OXA DNA damage in HepG2/OXA cells, and the molecular mechanism was related to the inhibitory effect on ERCC1 expression being mediated by the FGFR2/ERK1/2 signaling pathway.展开更多
Aim Alpha7 nicotinic acetylcholine receptor （α7nAChR）, a subtype of nAChR regulating neurotrans- mission in central nervous system, is an essential regulator of cholinergic antiinflammatory pathway in periphery. Th...Aim Alpha7 nicotinic acetylcholine receptor （α7nAChR）, a subtype of nAChR regulating neurotrans- mission in central nervous system, is an essential regulator of cholinergic antiinflammatory pathway in periphery. The present study was to determine the effects of activation of α7nAChR on oxidant stress-induced injury in endo- thelial cells. Methods Cultured human umbilical vein endothelial cells were treated with H202 （400 μmol · L^-1） or H202plus PNU-282987 （ 10 μmol · L^-1 ）. Cell viability and membrane integrity were measured. AnnexinV ＋ PI assay, immunoblotting of bcl-2, bax and cleaved caspase-3, and immunofluorescence of apoptosis inducing factor （AIF） were performed to evaluate apoptosis. Protein expression of vascular peroxidase-1 （ VPO-1 ） and phosphor- JNK were measured by immunoblotting. Results Activation of α7nAChR by a selective agonist PNU-282987 pre-vented H202-indced decrease of cell viability and increase of lactate dehydrogenase release. Activation of α7nAChR markedly reduced cell apoptosis and intracellular oxidative stress level. Moreover, activation of α7nAChR reduced H2 02 -induced VPO-1 protein upregulation and JNK1/2 phosphorylation. The inhibitory effect of α7nAChR activa- tion on VPO-1 was blocked by JNK inhibitor SP600125. In addition, pretreatment of α7nAChR antagonist methyl- lycaconitine blocked the cytoprotective effect of PNU-282987. Conclusion These results provide the first evidence that activation of α7nAChR protects against oxidant stress-induced damage by suppressing VPO-1 in a JNK signa- ling pathway-dependent manner in endothelial cells.展开更多
AIM To investigate the mechanism by which hepatitis C virus(HCV) core protein-induced mi R-93-5 p up-regulation regulates the interferon(IFN) signaling pathway.METHODS HCV-1 b core protein was exogenously expressed in...AIM To investigate the mechanism by which hepatitis C virus(HCV) core protein-induced mi R-93-5 p up-regulation regulates the interferon(IFN) signaling pathway.METHODS HCV-1 b core protein was exogenously expressed in Huh7 cells using pc DNA3.1(+) vector. The expression of mi R-93-5 p and interferon receptor 1(IFNAR1) was measured using quantitative reverse transcriptionpolymerase chain reaction and Western blot. The protein expression and phosphorylation level of STAT1 were evaluated by Western blot. The overexpression and silencing of mi R-93-5 p and IFNAR1 were performed using mi R-93-5 p agomir and antagomir, and pc DNA3.1-IFNAR1 and IFNAR1 si RNA, respectively. Luciferase assay was used to identify whether IFNAR1 is a target of mi R-93-5 p. Cellular experiments were also conducted.RESULTS Serum mi R-93-5 p level was increased in patients with HCV-1 b infection and decreased to normal level after HCV-1 b clearance, but persistently increased in those with pegylated interferon-α resistance, compared with healthy subjects. Serum mi R-93-5 p expression had an AUC value of 0.8359 in distinguishing patients with pegylated interferon-α resistance from those with pegylated interferon-α sensitivity. HCV-1 b core protein increased mi R-93-5 p expression and induced inactivation of the IFN signaling pathway in Huh7 cells. Furthermore, IFNAR1 was identified as a direct target of mi R-93-5 p, and IFNAR1 restore could rescue mi R-93-5 p-reduced STAT1 phosphorylation, suggesting that the mi R-93-5 p-IFNAR1 axis regulates the IFN signaling pathway.CONCLUSION HCV-1 b core protein-induced mi R-93-5 p up-regulation inhibits the IFN signaling pathway by directly targeting IFNAR1, and the mi R-93-5 p-IFNAR1 axis regulates STAT1 phosphorylation. This axis may be a potential therapeutic target for HCV-1 b infection.展开更多
The present study investigated a possible mechanism for endogenous endothelin-1 (ET-1) regulation of atrial natriuretic peptide (ANP) secretion in isolated perfused acute hypoxic rabbit atria. Acute hypoxia significan...The present study investigated a possible mechanism for endogenous endothelin-1 (ET-1) regulation of atrial natriuretic peptide (ANP) secretion in isolated perfused acute hypoxic rabbit atria. Acute hypoxia significantly enhanced the release of ET-1 and the expression of the ET receptor (ETR) type A and B (ETR<sub>A</sub> and ETR<sub>B</sub>) in atrial tissues, with a concomitant increase in ANP secretion. The ETR<sub>A</sub> or ETR<sub>B</sub> antagonist, BQ123 (0.3 μmol/L) or BQ788 (0.3 μmol/L), respectively attenuated hypoxia-induced ANP secretion. Both antagonists significantly attenuated the levels of hypoxiainduced atrial phosphorylated (p)-extracellular signal-regulated kinase (ERK) and p-protein kinase B (Akt). The ERK and Akt inhibitors, PD098059 (30 μmol/L) and LY294002 (30 μmol/L), respectively mimicked the effect of the ETR antagonists. These results demonstrated that acute hypoxia- mediated atrial ET-1 regulated ANP secretion through ETR and the subsequent mitogenactivated protein kinase (MAPK)/ERK and ETR-phosphatidylinositol-3-kinase (PI3K)/Akt signaling pathways. These pathways may mediate atrial endocrine functions under hypoxic conditions.展开更多
A major problem which is poorly understood in the management of bladder cancer is low sensitivity to chemotherapy and high recurrence after transurethral resection. Insulin-like growth factor 1 receptor (IGF-1R) signa...A major problem which is poorly understood in the management of bladder cancer is low sensitivity to chemotherapy and high recurrence after transurethral resection. Insulin-like growth factor 1 receptor (IGF-1R) signaling plays a very important role in progression, invasion and metastasis of bladder cancer cells. In this study, we investigated whether IGF-1R was involved in the growth stimulating activity and drug resistance of bladder cancer cells. The results showed: The mRNAs of IGF-1, IGF-2 and IGF-1R were strongly expressed in serum-free cultured T24 cell line, whereas normal urothelial cells did not express these factors/receptors or only in trace levels; T24 cell responded far better to growth stimulation by IGF-1 than did normal urothelial cells; blockage of IGF1R by antisense oligodeoxynucleotide (ODN) significantly inhibited the growth of T24 cell and enhanced sensitivity and apoptosis of T24 cells to mitomycin (MMC). These results suggested that blockage of IGF-IR signaling might potentially contribute to the treatment of bladder cancer cells which are insensitive to chemotherapy.展开更多
Early appearance of the serotonergic system in the fetal brain and the various effects of serotonin (5-HT) on brain morphogenesis, have given support to a neurotrophic role of serotonin. This function of serotonin is ...Early appearance of the serotonergic system in the fetal brain and the various effects of serotonin (5-HT) on brain morphogenesis, have given support to a neurotrophic role of serotonin. This function of serotonin is accomplished through a system of serotonin nerve terminals in the target regions that involves various 5-HT receptors. In visual, auditory and somatosensory cortex an early and intense serotonergic innervation is particularly important. The neuronal somata of these terminals are normally located in the mesencephalon and they have not been observed in the maturing cerebral cortex, neither in the adult brain. By using immunolabeling techniques, fluorescence and confocal microscopy, we observe the presence of both, 5-HT terminals and 5-HT cells in mesencephalon (Me, E17) and in the neopallium (Np, E13-E16) cocultures. Cells immunopositive to 5-HT and to tryptophan-5-hydroxilase are also observed in the Np on day 12 of culture. These results concerning the unexpected presence of serotonergic cells in the fetal cerebral cortex are interesting and may be of importance in corticogenesis. As it happens with other elements of the serotonergic system, the presence of these phenotypically serotonergic cells in the early cerebral cortex may be transitory and probably supporting cortex maturation processes. The molecular signaling path of the 5-HT1A receptor has also been identified.展开更多
Oxidized low-density lipoprotein receptor 1(OLR1)is upregulated in neurons and participates in hypertension-induced neuronal apoptosis.OLR1 deletion exerts protective effects on cerebral damage induced by hypertensive...Oxidized low-density lipoprotein receptor 1(OLR1)is upregulated in neurons and participates in hypertension-induced neuronal apoptosis.OLR1 deletion exerts protective effects on cerebral damage induced by hypertensive-induced stroke.Therefore,OLR1 is likely involved in the progress of intracerebral hemorrhage.In this study,we examined the potential role of OLR1 in intracerebral hemorrhage using a rat model.OLR1 small interfering RNA(10μL;50 pmol/μL)was injected into the right basal ganglia to knock down OLR1.Twenty-four hours later,0.5 U collagenase type VII was injected to induce intracerebral hemorrhage.We found that knockdown of OLR1 attenuated neurological behavior impairment in rats with intracerebral hemorrhage and reduced hematoma,neuron loss,inflammatory reaction,and oxidative stress in rat brain tissue.We also found that silencing of OLR1 suppressed ferroptosis induced by intracerebral hemorrhage and the p38 signaling pathway.Therefore,silencing OLR1 exhibits protective effects against secondary injury of intracerebral hemorrhage.These findings suggest that OLR1 may be a novel potential therapeutic target for intracerebral hemorrhage.展开更多
In the past,contraction-induced production of reactive oxygen species(ROS)has been implicated in oxidative stress to skeletal muscle.As research advances,clear evidence has revealed a more complete role of ROS under b...In the past,contraction-induced production of reactive oxygen species(ROS)has been implicated in oxidative stress to skeletal muscle.As research advances,clear evidence has revealed a more complete role of ROS under both physiologic and pathologic conditions.Central to the role of ROS is the redox signaling pathways that control exercise-induced major physiologic and cellular responses and adaptations,such as mitochondrial biogenesis,mitophagy,mitochondrial morphologic dynamics,antioxidant defense,and inflammation.The current review focuses on how muscle contraction and immobilization may activate or inhibit redox signalings and their impact on muscle mitochondrial homeostasis and physiologic implications.展开更多
Many researchers employed mammalian expression system to artificially express cannabinoid receptors, but immunoblot data that directly prove efficient protein expression can hardly be seen in related research reports....Many researchers employed mammalian expression system to artificially express cannabinoid receptors, but immunoblot data that directly prove efficient protein expression can hardly be seen in related research reports. In present study, we demonstrated cannabinoid receptor protein was not able to be properly expressed with routine mammalian expression system. This inefficient expression was rescued by endowing an exogenous signal peptide ahead of cannabinoid receptor peptide. In addition, the artificially synthesized cannabinoid receptor was found to aggregate under routine sample denaturing temperatures (i.e.,≥95°C), forming a large molecular weight band when analyzed by immuno-blotting. Only denaturing temperatures ≤75°C yielded a clear band at the predicted molecular weight. Collectively, we showed that efficient mammalian expression of cannabinoid receptors need a signal peptide sequence, and described the requirement for a low sample denaturing temperature in immuno-blot analysis. These findings provide very useful information for efficient mammalian expression and immuno-blotting of membrane receptors.展开更多
The activation of the mitogen-activated protein(MAP) kinases extracellular signal-regulated kinase(ERK)1/2 was traditionally used as a readout of signaling of G protein-coupled receptors(GPCRs) via arrestins, as oppos...The activation of the mitogen-activated protein(MAP) kinases extracellular signal-regulated kinase(ERK)1/2 was traditionally used as a readout of signaling of G protein-coupled receptors(GPCRs) via arrestins, as opposed to conventional GPCR signaling via G proteins. Several recent studies using HEK293 cells where all G proteins were genetically ablated or inactivated, or both non-visual arrestins were knocked out, demonstrated that ERK1/2 phosphorylation requires G protein activity, but does not necessarily require the presence of non-visual arrestins. This appears to contradict the prevailing paradigm. Here we discuss these results along with the recent data on gene edited cells and arrestinmediated signaling. We suggest that there is no real controversy. G proteins might be involved in the activation of the upstream-most MAP3Ks, although in vivo most MAP3K activation is independent of heterotrimeric G proteins, being initiated by receptor tyrosine kinases and/or integrins. As far as MAP kinases are concerned, the best-established role of arrestins is scaffolding of the three-tiered cascades(MAP3K-MAP2 K-MAPK). Thus, it seems likely that arrestins, GPCRbound and free, facilitate the propagation of signals in these cascades, whereas signal initiation via MAP3K activation may be independent of arrestins. Different MAP3Ks are activated by various inputs, some of which are mediated by G proteins, particularly in cell culture, where we artificially prevent signaling by receptor tyrosine kinases and integrins, thereby favoring GPCR-induced signaling. Thus, there is no reason to change the paradigm: Arrestins and G proteins play distinct non-overlapping roles in cell signaling.展开更多
基金supported by the National Natural Science Foundation of China(Youth Science Fund Project),No.81901292(to GC)the National Key Research and Development Program of China,No.2021YFC2502100(to GC)the National Natural Science Foundation of China,No.82071183(to ZZ).
文摘Netrin-1 and its receptors play crucial roles in inducing axonal growth and neuronal migration during neuronal development.Their profound impacts then extend into adulthood to encompass the maintenance of neuronal survival and synaptic function.Increasing amounts of evidence highlight several key points:(1)Diminished Netrin-1 levels exacerbate pathological progression in animal models of Alzheimer’s disease and Parkinson’s disease,and potentially,similar alterations occur in humans.(2)Genetic mutations of Netrin-1 receptors increase an individuals’susceptibility to neurodegenerative disorders.(3)Therapeutic approaches targeting Netrin-1 and its receptors offer the benefits of enhancing memory and motor function.(4)Netrin-1 and its receptors show genetic and epigenetic alterations in a variety of cancers.These findings provide compelling evidence that Netrin-1 and its receptors are crucial targets in neurodegenerative diseases.Through a comprehensive review of Netrin-1 signaling pathways,our objective is to uncover potential therapeutic avenues for neurodegenerative disorders.
基金supported by a grant from Ministry of Science,Technological Development and Innovation,Serbia,No.451-03-68/2022-14/200178(to NN)University of Defence,No.MFVMA/02/22-24(to MN)。
文摘An imbalance in adenosine-mediated signaling,particularly the increased A_(2A)R-mediated signaling,plays a role in the pathogenesis of Parkinson's disease.Existing therapeutic approaches fail to alter disease progression,demonstrating the need for novel approaches in PD.Repetitive transcranial magnetic stimulation is a non-invasive approach that has been shown to improve motor and non-motor symptoms of Parkinson's disease.However,the underlying mechanisms of the beneficial effects of repetitive transcranial magnetic stimulation remain unknown.The purpose of this study is to investigate the extent to which the beneficial effects of prolonged intermittent theta burst stimulation in the 6-hydroxydopamine model of experimental parkinsonism are based on modulation of adenosine-mediated signaling.Animals with unilateral 6-hydroxydopamine lesions underwent intermittent theta burst stimulation for 3 weeks and were tested for motor skills using the Rotarod test.Immunoblot,quantitative reverse transcription polymerase chain reaction,immunohistochemistry,and biochemical analysis of components of adenosine-mediated signaling were performed on the synaptosomal fraction of the lesioned caudate putamen.Prolonged intermittent theta burst stimulation improved motor symptoms in 6-hydroxydopamine-lesioned animals.A 6-hydroxydopamine lesion resulted in progressive loss of dopaminergic neurons in the caudate putamen.Treatment with intermittent theta burst stimulation began 7 days after the lesion,coinciding with the onset of motor symptoms.After treatment with prolonged intermittent theta burst stimulation,complete motor recovery was observed.This improvement was accompanied by downregulation of the e N/CD73-A_(2A)R pathway and a return to physiological levels of A_(1)R-adenosine deaminase 1 after 3 weeks of intermittent theta burst stimulation.Our results demonstrated that 6-hydroxydopamine-induced degeneration reduced the expression of A_(1)R and elevated the expression of A_(2A)R.Intermittent theta burst stimulation reversed these effects by restoring the abundances of A_(1)R and A_(2A)R to control levels.The shift in ARs expression likely restored the balance between dopamine-adenosine signaling,ultimately leading to the recovery of motor control.
基金Supported by National Natural Sciences Foundation of China,No. 81001067the Ministry of Science and Technology International Cooperation Project, No. 2010DFA31870the AstraZeneca Special Research Foundation for Targeted Therapy of the Wu Jieping Medical Foundation, No. 320.6700.09068
文摘AIM: To investigate the molecular mechanisms underlying the reversal effect of emodin on platinum resistance in hepatocellular carcinoma. METHODS: After the addition of 10 μmol/L emodin to HepG2/oxaliplatin (OXA) cells, the inhibition rate (IR), 50% inhibitory concentration (IC 50 ) and reversal index (IC 50 in experimental group/IC 50 in control group) were calculated. For HepG2, HepG2/OXA, HepG2/OXA/T, each cell line was divided into a control group, OXA group, OXA + fibroblast growth factor 7 (FGF7) group and OXA + emodin group, and the final concentrations of FGF7, emodin and OXA in each group were 5 ng/mL, 10 μg/mL and 10 μmol/L, respectively. Single-cell gel electrophoresis was conducted to detect DNA damage, and the fibroblast growth factor receptor 2 (FGFR2), phosphorylated extracellular signal-regulated kinase 1/2 (p-ERK1/2) and excision repair cross-complementing gene 1 (ERCC1) protein expression levels in each group were examined by Western blotting. RESULTS: Compared with the IC50 of 120.78 μmol/L in HepG2/OXA cells, the IC 50 decreased to 39.65 μmol/L after treatment with 10 μmol/L emodin; thus, the reversal index was 3.05. Compared with the control group, the tail length and Olive tail length in the OXA group, OXA + FGF7 group and OXA + emodin group were significantly increased, and the differences were statistically significant (P < 0.01). The tail length and Olive tail length were lower in the OXA + FGF7 group than in the OXA group, and this difference was also statistically significant. Compared with the OXA + FGF7 group, the tail extent, the Olive tail moment and the percentage of tail DNA were significantly increased in the OXA + emodin group, and these differences were statistically significant (P < 0.01). In comparison with its parental cell line HepG2, the HepG2/OXA cells demonstrated significantly increased FGFR2, p-ERK1/2 and ERCC1 expression levels, whereas the expression of all three molecules was significantly inhibited in HepG2/ OXA/T cells, in which FGFR2 was silenced by FGFR2 shRNA. In the examined HepG2 cells, the FGFR2, p-ERK1/2 and ERCC1 expression levels demonstrated increasing trends in the OXA group and OXA + FGF7 group. Compared with the OXA group and OXA + FGF7 group, the FGFR2, p-ERK1/2, and ERCC1 expression levels were significantly lower in the OXA + emodin group, and these differences were statistically significant. In the HepG2/OXA/T cell line that was transfected with FGFR2 shRNA, the FGFR2, p-ERK1/2 and ERCC1 expression levels were significantly inhibited, but there were no significant differences in these expression levels among the OXA, OXA + FGF7 and OXA + emodin groups. CONCLUSION: Emodin markedly reversed OXA resistance by enhancing OXA DNA damage in HepG2/OXA cells, and the molecular mechanism was related to the inhibitory effect on ERCC1 expression being mediated by the FGFR2/ERK1/2 signaling pathway.
文摘Aim Alpha7 nicotinic acetylcholine receptor （α7nAChR）, a subtype of nAChR regulating neurotrans- mission in central nervous system, is an essential regulator of cholinergic antiinflammatory pathway in periphery. The present study was to determine the effects of activation of α7nAChR on oxidant stress-induced injury in endo- thelial cells. Methods Cultured human umbilical vein endothelial cells were treated with H202 （400 μmol · L^-1） or H202plus PNU-282987 （ 10 μmol · L^-1 ）. Cell viability and membrane integrity were measured. AnnexinV ＋ PI assay, immunoblotting of bcl-2, bax and cleaved caspase-3, and immunofluorescence of apoptosis inducing factor （AIF） were performed to evaluate apoptosis. Protein expression of vascular peroxidase-1 （ VPO-1 ） and phosphor- JNK were measured by immunoblotting. Results Activation of α7nAChR by a selective agonist PNU-282987 pre-vented H202-indced decrease of cell viability and increase of lactate dehydrogenase release. Activation of α7nAChR markedly reduced cell apoptosis and intracellular oxidative stress level. Moreover, activation of α7nAChR reduced H2 02 -induced VPO-1 protein upregulation and JNK1/2 phosphorylation. The inhibitory effect of α7nAChR activa- tion on VPO-1 was blocked by JNK inhibitor SP600125. In addition, pretreatment of α7nAChR antagonist methyl- lycaconitine blocked the cytoprotective effect of PNU-282987. Conclusion These results provide the first evidence that activation of α7nAChR protects against oxidant stress-induced damage by suppressing VPO-1 in a JNK signa- ling pathway-dependent manner in endothelial cells.
基金Supported by National Natural Science Foundation of China,No.81371849the TMMU Key Project for Clinical Research,No.2012XLC05
文摘AIM To investigate the mechanism by which hepatitis C virus(HCV) core protein-induced mi R-93-5 p up-regulation regulates the interferon(IFN) signaling pathway.METHODS HCV-1 b core protein was exogenously expressed in Huh7 cells using pc DNA3.1(+) vector. The expression of mi R-93-5 p and interferon receptor 1(IFNAR1) was measured using quantitative reverse transcriptionpolymerase chain reaction and Western blot. The protein expression and phosphorylation level of STAT1 were evaluated by Western blot. The overexpression and silencing of mi R-93-5 p and IFNAR1 were performed using mi R-93-5 p agomir and antagomir, and pc DNA3.1-IFNAR1 and IFNAR1 si RNA, respectively. Luciferase assay was used to identify whether IFNAR1 is a target of mi R-93-5 p. Cellular experiments were also conducted.RESULTS Serum mi R-93-5 p level was increased in patients with HCV-1 b infection and decreased to normal level after HCV-1 b clearance, but persistently increased in those with pegylated interferon-α resistance, compared with healthy subjects. Serum mi R-93-5 p expression had an AUC value of 0.8359 in distinguishing patients with pegylated interferon-α resistance from those with pegylated interferon-α sensitivity. HCV-1 b core protein increased mi R-93-5 p expression and induced inactivation of the IFN signaling pathway in Huh7 cells. Furthermore, IFNAR1 was identified as a direct target of mi R-93-5 p, and IFNAR1 restore could rescue mi R-93-5 p-reduced STAT1 phosphorylation, suggesting that the mi R-93-5 p-IFNAR1 axis regulates the IFN signaling pathway.CONCLUSION HCV-1 b core protein-induced mi R-93-5 p up-regulation inhibits the IFN signaling pathway by directly targeting IFNAR1, and the mi R-93-5 p-IFNAR1 axis regulates STAT1 phosphorylation. This axis may be a potential therapeutic target for HCV-1 b infection.
文摘The present study investigated a possible mechanism for endogenous endothelin-1 (ET-1) regulation of atrial natriuretic peptide (ANP) secretion in isolated perfused acute hypoxic rabbit atria. Acute hypoxia significantly enhanced the release of ET-1 and the expression of the ET receptor (ETR) type A and B (ETR<sub>A</sub> and ETR<sub>B</sub>) in atrial tissues, with a concomitant increase in ANP secretion. The ETR<sub>A</sub> or ETR<sub>B</sub> antagonist, BQ123 (0.3 μmol/L) or BQ788 (0.3 μmol/L), respectively attenuated hypoxia-induced ANP secretion. Both antagonists significantly attenuated the levels of hypoxiainduced atrial phosphorylated (p)-extracellular signal-regulated kinase (ERK) and p-protein kinase B (Akt). The ERK and Akt inhibitors, PD098059 (30 μmol/L) and LY294002 (30 μmol/L), respectively mimicked the effect of the ETR antagonists. These results demonstrated that acute hypoxia- mediated atrial ET-1 regulated ANP secretion through ETR and the subsequent mitogenactivated protein kinase (MAPK)/ERK and ETR-phosphatidylinositol-3-kinase (PI3K)/Akt signaling pathways. These pathways may mediate atrial endocrine functions under hypoxic conditions.
文摘A major problem which is poorly understood in the management of bladder cancer is low sensitivity to chemotherapy and high recurrence after transurethral resection. Insulin-like growth factor 1 receptor (IGF-1R) signaling plays a very important role in progression, invasion and metastasis of bladder cancer cells. In this study, we investigated whether IGF-1R was involved in the growth stimulating activity and drug resistance of bladder cancer cells. The results showed: The mRNAs of IGF-1, IGF-2 and IGF-1R were strongly expressed in serum-free cultured T24 cell line, whereas normal urothelial cells did not express these factors/receptors or only in trace levels; T24 cell responded far better to growth stimulation by IGF-1 than did normal urothelial cells; blockage of IGF1R by antisense oligodeoxynucleotide (ODN) significantly inhibited the growth of T24 cell and enhanced sensitivity and apoptosis of T24 cells to mitomycin (MMC). These results suggested that blockage of IGF-IR signaling might potentially contribute to the treatment of bladder cancer cells which are insensitive to chemotherapy.
文摘Early appearance of the serotonergic system in the fetal brain and the various effects of serotonin (5-HT) on brain morphogenesis, have given support to a neurotrophic role of serotonin. This function of serotonin is accomplished through a system of serotonin nerve terminals in the target regions that involves various 5-HT receptors. In visual, auditory and somatosensory cortex an early and intense serotonergic innervation is particularly important. The neuronal somata of these terminals are normally located in the mesencephalon and they have not been observed in the maturing cerebral cortex, neither in the adult brain. By using immunolabeling techniques, fluorescence and confocal microscopy, we observe the presence of both, 5-HT terminals and 5-HT cells in mesencephalon (Me, E17) and in the neopallium (Np, E13-E16) cocultures. Cells immunopositive to 5-HT and to tryptophan-5-hydroxilase are also observed in the Np on day 12 of culture. These results concerning the unexpected presence of serotonergic cells in the fetal cerebral cortex are interesting and may be of importance in corticogenesis. As it happens with other elements of the serotonergic system, the presence of these phenotypically serotonergic cells in the early cerebral cortex may be transitory and probably supporting cortex maturation processes. The molecular signaling path of the 5-HT1A receptor has also been identified.
基金supported by the National Natural Science Foundation of China,No.81971125(to ZYH).
文摘Oxidized low-density lipoprotein receptor 1(OLR1)is upregulated in neurons and participates in hypertension-induced neuronal apoptosis.OLR1 deletion exerts protective effects on cerebral damage induced by hypertensive-induced stroke.Therefore,OLR1 is likely involved in the progress of intracerebral hemorrhage.In this study,we examined the potential role of OLR1 in intracerebral hemorrhage using a rat model.OLR1 small interfering RNA(10μL;50 pmol/μL)was injected into the right basal ganglia to knock down OLR1.Twenty-four hours later,0.5 U collagenase type VII was injected to induce intracerebral hemorrhage.We found that knockdown of OLR1 attenuated neurological behavior impairment in rats with intracerebral hemorrhage and reduced hematoma,neuron loss,inflammatory reaction,and oxidative stress in rat brain tissue.We also found that silencing of OLR1 suppressed ferroptosis induced by intracerebral hemorrhage and the p38 signaling pathway.Therefore,silencing OLR1 exhibits protective effects against secondary injury of intracerebral hemorrhage.These findings suggest that OLR1 may be a novel potential therapeutic target for intracerebral hemorrhage.
文摘In the past,contraction-induced production of reactive oxygen species(ROS)has been implicated in oxidative stress to skeletal muscle.As research advances,clear evidence has revealed a more complete role of ROS under both physiologic and pathologic conditions.Central to the role of ROS is the redox signaling pathways that control exercise-induced major physiologic and cellular responses and adaptations,such as mitochondrial biogenesis,mitophagy,mitochondrial morphologic dynamics,antioxidant defense,and inflammation.The current review focuses on how muscle contraction and immobilization may activate or inhibit redox signalings and their impact on muscle mitochondrial homeostasis and physiologic implications.
基金supported by a grant from Army Medical Research Program of China(No.08G168)
文摘Many researchers employed mammalian expression system to artificially express cannabinoid receptors, but immunoblot data that directly prove efficient protein expression can hardly be seen in related research reports. In present study, we demonstrated cannabinoid receptor protein was not able to be properly expressed with routine mammalian expression system. This inefficient expression was rescued by endowing an exogenous signal peptide ahead of cannabinoid receptor peptide. In addition, the artificially synthesized cannabinoid receptor was found to aggregate under routine sample denaturing temperatures (i.e.,≥95°C), forming a large molecular weight band when analyzed by immuno-blotting. Only denaturing temperatures ≤75°C yielded a clear band at the predicted molecular weight. Collectively, we showed that efficient mammalian expression of cannabinoid receptors need a signal peptide sequence, and described the requirement for a low sample denaturing temperature in immuno-blot analysis. These findings provide very useful information for efficient mammalian expression and immuno-blotting of membrane receptors.
基金Supported by National Institutes of Health RO1 grants,No.EY011500National Institutes of Health R35 grants,No.GM122491Cornelius Vanderbilt Endowed Chair(Vanderbilt University),No.NS065868(to Gurevich VV)and No.DA030103(to Gurevich EV)
文摘The activation of the mitogen-activated protein(MAP) kinases extracellular signal-regulated kinase(ERK)1/2 was traditionally used as a readout of signaling of G protein-coupled receptors(GPCRs) via arrestins, as opposed to conventional GPCR signaling via G proteins. Several recent studies using HEK293 cells where all G proteins were genetically ablated or inactivated, or both non-visual arrestins were knocked out, demonstrated that ERK1/2 phosphorylation requires G protein activity, but does not necessarily require the presence of non-visual arrestins. This appears to contradict the prevailing paradigm. Here we discuss these results along with the recent data on gene edited cells and arrestinmediated signaling. We suggest that there is no real controversy. G proteins might be involved in the activation of the upstream-most MAP3Ks, although in vivo most MAP3K activation is independent of heterotrimeric G proteins, being initiated by receptor tyrosine kinases and/or integrins. As far as MAP kinases are concerned, the best-established role of arrestins is scaffolding of the three-tiered cascades(MAP3K-MAP2 K-MAPK). Thus, it seems likely that arrestins, GPCRbound and free, facilitate the propagation of signals in these cascades, whereas signal initiation via MAP3K activation may be independent of arrestins. Different MAP3Ks are activated by various inputs, some of which are mediated by G proteins, particularly in cell culture, where we artificially prevent signaling by receptor tyrosine kinases and integrins, thereby favoring GPCR-induced signaling. Thus, there is no reason to change the paradigm: Arrestins and G proteins play distinct non-overlapping roles in cell signaling.
