目的:观察MAPK4基因敲除(KO)对小鼠脾脏免疫细胞组成的影响并探讨其意义。方法:利用基因同源重组对小鼠MAPK4基因进行敲除,PCR法鉴定MAPK4^-/-小鼠;Western blot检测小鼠脾脏中MAPK4蛋白表达;计算MAPK4^-/-小鼠脾脏的脏器指数和总细胞数...目的:观察MAPK4基因敲除(KO)对小鼠脾脏免疫细胞组成的影响并探讨其意义。方法:利用基因同源重组对小鼠MAPK4基因进行敲除,PCR法鉴定MAPK4^-/-小鼠;Western blot检测小鼠脾脏中MAPK4蛋白表达;计算MAPK4^-/-小鼠脾脏的脏器指数和总细胞数;HE染色观察脾脏的病理学变化;流式细胞术(FCM)分析小鼠脾脏中CD19^+B细胞和CD3^+T细胞的比例并计算细胞绝对数,并检测CD4^+T和CD8^+T细胞的比例及其活化相关膜分子CD62L和CD69的表达;最后,FCM分析脾脏中固有免疫细胞DC细胞、NK细胞、巨噬细胞和γδT细胞的比例并计算细胞绝对数。结果:与野生型(WT)小鼠相比,MAPK4^-/-小鼠的MAPK4蛋白表达水平明显降低( P <0.05);脏器指数无明显改变( P >0.05),脾脏细胞总数明显降低( P <0.05),脾小结增大,结构紊乱( P <0.05);CD19^+B细胞和CD3 ^+T细胞的细胞绝对数都明显降低( P <0.001);CD4 +T细胞的比例和绝对数均明显减少( P <0.01),同时,CD8 +T细胞的绝对数明显减少( P <0.001);固有免疫细胞中DC细胞、NK细胞、巨噬细胞和γδT细胞的细胞绝对数均明显降低( P <0.001)。结论: MAPK4敲除明显影响小鼠脾脏中免疫细胞的组成,提示其在机体免疫细胞发育和功能中具有重要作用。展开更多
Background:Angiogenesis plays an important role in the occurrence and development of non-small cell lung cancer(NSCLC).The atypical mitogen-activated protein kinase 4(MAPK4)has been shown to be involved in the pathoge...Background:Angiogenesis plays an important role in the occurrence and development of non-small cell lung cancer(NSCLC).The atypical mitogen-activated protein kinase 4(MAPK4)has been shown to be involved in the pathogenesis of various diseases.However,the potential role of MAPK4 in the tumor angiogenesis of NSCLC remains unclear.Methods:Adult male C57BL/6 wild-type mice were randomly divided into the control group and p-siMAPK4 intervention group,respectively.The cell proliferation was analyzed with flow cytometry and immunofluorescence staining.The vascular density in tumor mass was analyzed by immuno-fluorescence staining.The expressions of MAPK4 and related signaling molecules were detected by western blot analysis and immunofluorescence staining,and so on.Results:We found that the expression of MAPK4,which was dominantly expressed in local endothelial cells(ECs),was correlated with tumor angiogenesis of NSCLC.Furthermore,MAPK4 silencing inhibited the proliferation and migration abilities of human umbilical vein ECs(HUVECs).QKHZC-2020-4Y156,QKH-JC-2018-1428,QKH-RC-2019-5612;Collaborative Innovation Center of Chinese Ministry of Education,Grant/Award Number:2020-39;Program for Science and Technology Joint Fund Project in Zunyi Science and Technology Bureau and Zunyi Medical University,Grant/Award Numbers:ZSKH-RPT-2020-6,ZSKH-HZ-2021-193;Graduate Research Fund of Zunyi Medical University,Grant/Award Number:2023-ZYK-171 Global gene analysis showed that MAPK4 silencing altered the expression of multiple genes related to cell cycle and angiogenesis pathways,and that MAPK4 silencing increased transduction of the extracellular regulated protein kinases 1/2(ERK1/2)pathway but not Akt and c-Jun n-terminal kinase pathways.Further analysis showed that MAPK4 silencing inhibited the proliferation and migration abilities of HUVECs cultured in tumor cell supernatant,which was accompanied with increased transduction of the ERK1/2 pathway.Clinical data analysis suggested that the higher expression of MAPK4 and CD34 were associated with poor prognosis of patients with NSCLC.Targeted silencing of MAPK4 in ECs using small interfering RNA driven by the CD34 promoter effectively inhibited tumor angiogenesis and growth of NSCLC in vivo.Conclusion:Our results reveal that MAPK4 plays an important role in the angiogenesis and development of NSCLC.MAPK4 may thus represent a new target for NSCLC.展开更多
Aralia taibaiensi,widely distributed in western China,particularly in the Qinba Mountains,has been utilized as a folk medicine for treating diabetes,gastropathy,rheumatism,and cardiovascular diseases.Saponins from A.t...Aralia taibaiensi,widely distributed in western China,particularly in the Qinba Mountains,has been utilized as a folk medicine for treating diabetes,gastropathy,rheumatism,and cardiovascular diseases.Saponins from A.taibaiensis(sAT)have demonstrated protective effects against oxidative stress and mitochondrial dysfunction induced by ischemia/reperfusion(I/R).However,the underlying mechanisms remain unclear.In vivo,middle cerebral artery occlusion/reperfusion(MCAO/R)induced inflammatory infiltration,neuronal injury,cell apoptosis,mitochondrial dysfunction,and oxidative stress in the ischaemic penumbra,which were effectively mitigated by sAT.sAT increased the mRNA and protein expression levels of apelin and its receptor apelin/apelin receptors(ARs)both in vivo and in vitro.(Ala13)-Apelin-13(F13A)and small interfering RNA(siRNA)abolished the regulatory effects of sAT on neuroprotection mediated by adenosine 5'-monophosphate(AMP)-activated protein kinase(AMPK)/protein kinase B(Akt).Furthermore,sAT induced apelin/AR expression by simultaneously inhibiting P38 mitogen-activated protein kinase(P38 MAPK)/activating transcription factor 4(ATF4)and upregulating hypoxia-inducible factor-1α(HIF-1α).Our findings indicate that sAT regulates apelin/AR/AMPK by inhibiting P38 MAPK/ATF4 and upregulating HIF-1a,thereby suppressing oxidative stress and mitochondrial dysfunction.展开更多
AIM:To investigate the effects of a Chinese medicine formula“Qingxuan Runmu Yin”(QRY)on ocular surface inflammation in a rat model of dry eye,and its mechanism via the toll-like receptor 4(TLR4)/transforming growth ...AIM:To investigate the effects of a Chinese medicine formula“Qingxuan Runmu Yin”(QRY)on ocular surface inflammation in a rat model of dry eye,and its mechanism via the toll-like receptor 4(TLR4)/transforming growth factor kinase 1(TAK1)/p38 mitogen-activated protein kinase(p38MAPK)signaling pathway.METHODS:Seventy-two Sprague-Dawley rats were randomly divided into six groups(n=12 each):the control group,model group,3 groups of QRY(with low-,medium-,and high-doses),and SB203580 group.Dry eye was induced using benzalkonium chloride.Schirmer’s test(SIT)and corneal fluorescein staining(CFS)were performed every 14d throughout the experiment.Histopathological changes in corneal and conjunctival tissues were observed using hematoxylin and eosin(HE)and periodic acid-Schiff(PAS)staining.Protein expression levels of key inflammatory markers and signaling pathway targets were assessed via immunohistochemistry,ELISA,and Western blotting.RESULTS:Compared to the control group,the model group showed significant reductions in SIT and increases in CFS scores,alongside structural disorganization of corneal/conjunctival tissues,decreased conjunctival goblet cell(CGC)numbers,and elevated expression of inflammatory markers[interleukin(IL)-1β,IL-6,tumor necrosis factoralpha(TNF-α),matrix metalloproteinase-9(MMP9)]and pathway proteins(TLR4,p-TAK1,p-p38MAPK;P<0.05).Treatment with QRY(low,medium,and high doses)and SB203580 significantly improved SIT scores,reduced CFS scores,restored corneoconjunctival structure,increased CGC numbers,and decreased expression levels of IL-1β,IL-6,TNF-α,MMP9,TLR4,p-TAK1,and p-p38MAPK proteins compared to the model group(P<0.05).CONCLUSION:QRY may alleviate ocular surface inflammation associated with dry eye by inhibiting the TLR4/TAK1/p38MAPK signaling pathway,highlighting its potential therapeutic efficacy for dry eye.展开更多
文摘目的:观察MAPK4基因敲除(KO)对小鼠脾脏免疫细胞组成的影响并探讨其意义。方法:利用基因同源重组对小鼠MAPK4基因进行敲除,PCR法鉴定MAPK4^-/-小鼠;Western blot检测小鼠脾脏中MAPK4蛋白表达;计算MAPK4^-/-小鼠脾脏的脏器指数和总细胞数;HE染色观察脾脏的病理学变化;流式细胞术(FCM)分析小鼠脾脏中CD19^+B细胞和CD3^+T细胞的比例并计算细胞绝对数,并检测CD4^+T和CD8^+T细胞的比例及其活化相关膜分子CD62L和CD69的表达;最后,FCM分析脾脏中固有免疫细胞DC细胞、NK细胞、巨噬细胞和γδT细胞的比例并计算细胞绝对数。结果:与野生型(WT)小鼠相比,MAPK4^-/-小鼠的MAPK4蛋白表达水平明显降低( P <0.05);脏器指数无明显改变( P >0.05),脾脏细胞总数明显降低( P <0.05),脾小结增大,结构紊乱( P <0.05);CD19^+B细胞和CD3 ^+T细胞的细胞绝对数都明显降低( P <0.001);CD4 +T细胞的比例和绝对数均明显减少( P <0.01),同时,CD8 +T细胞的绝对数明显减少( P <0.001);固有免疫细胞中DC细胞、NK细胞、巨噬细胞和γδT细胞的细胞绝对数均明显降低( P <0.001)。结论: MAPK4敲除明显影响小鼠脾脏中免疫细胞的组成,提示其在机体免疫细胞发育和功能中具有重要作用。
基金National Natural Science Foundation of China,Grant/Award Numbers:82272812,81960509,32160178Program for High Level Innovative Talents in Guizhou Province,Grant/Award Number:QKH-RC-2016-4031+4 种基金Program for Excellent Young Talents of Zunyi Medical University,Grant/Award Number:15ZY-001Project of Guizhou Provincial Department of Science and Technology,Grant/Award Numbers:QKHZC-2020-4Y156,QKH-JC-2018-1428,QKH-RC-2019-5612Collaborative Innovation Center of Chinese Ministry of Education,Grant/Award Number:2020-39Program for Science and Technology Joint Fund Project in Zunyi Science and Technology Bureau and Zunyi Medical University,Grant/Award Numbers:ZSKH-RPT-2020-6,ZSKH-HZ-2021-193Graduate Research Fund of Zunyi Medical University,Grant/Award Number:2023-ZYK-171。
文摘Background:Angiogenesis plays an important role in the occurrence and development of non-small cell lung cancer(NSCLC).The atypical mitogen-activated protein kinase 4(MAPK4)has been shown to be involved in the pathogenesis of various diseases.However,the potential role of MAPK4 in the tumor angiogenesis of NSCLC remains unclear.Methods:Adult male C57BL/6 wild-type mice were randomly divided into the control group and p-siMAPK4 intervention group,respectively.The cell proliferation was analyzed with flow cytometry and immunofluorescence staining.The vascular density in tumor mass was analyzed by immuno-fluorescence staining.The expressions of MAPK4 and related signaling molecules were detected by western blot analysis and immunofluorescence staining,and so on.Results:We found that the expression of MAPK4,which was dominantly expressed in local endothelial cells(ECs),was correlated with tumor angiogenesis of NSCLC.Furthermore,MAPK4 silencing inhibited the proliferation and migration abilities of human umbilical vein ECs(HUVECs).QKHZC-2020-4Y156,QKH-JC-2018-1428,QKH-RC-2019-5612;Collaborative Innovation Center of Chinese Ministry of Education,Grant/Award Number:2020-39;Program for Science and Technology Joint Fund Project in Zunyi Science and Technology Bureau and Zunyi Medical University,Grant/Award Numbers:ZSKH-RPT-2020-6,ZSKH-HZ-2021-193;Graduate Research Fund of Zunyi Medical University,Grant/Award Number:2023-ZYK-171 Global gene analysis showed that MAPK4 silencing altered the expression of multiple genes related to cell cycle and angiogenesis pathways,and that MAPK4 silencing increased transduction of the extracellular regulated protein kinases 1/2(ERK1/2)pathway but not Akt and c-Jun n-terminal kinase pathways.Further analysis showed that MAPK4 silencing inhibited the proliferation and migration abilities of HUVECs cultured in tumor cell supernatant,which was accompanied with increased transduction of the ERK1/2 pathway.Clinical data analysis suggested that the higher expression of MAPK4 and CD34 were associated with poor prognosis of patients with NSCLC.Targeted silencing of MAPK4 in ECs using small interfering RNA driven by the CD34 promoter effectively inhibited tumor angiogenesis and growth of NSCLC in vivo.Conclusion:Our results reveal that MAPK4 plays an important role in the angiogenesis and development of NSCLC.MAPK4 may thus represent a new target for NSCLC.
基金supported by the National Science Foundation of China(No.81903832),the Key Research and Development Plan of Shaanxi Province(No.2022SF-182),the Fundamental Research Funds for the Central Universities(No.D5000210799)Shaanxi Provincial Traditional Chinese Medicine Administration"Double Chain Integration"Middle Youth Research and Innovation Team Project(No.2022-SLRH-LJ-007)Shanghai Science and Technology Innovation Action Plan(No.22S21902600).
文摘Aralia taibaiensi,widely distributed in western China,particularly in the Qinba Mountains,has been utilized as a folk medicine for treating diabetes,gastropathy,rheumatism,and cardiovascular diseases.Saponins from A.taibaiensis(sAT)have demonstrated protective effects against oxidative stress and mitochondrial dysfunction induced by ischemia/reperfusion(I/R).However,the underlying mechanisms remain unclear.In vivo,middle cerebral artery occlusion/reperfusion(MCAO/R)induced inflammatory infiltration,neuronal injury,cell apoptosis,mitochondrial dysfunction,and oxidative stress in the ischaemic penumbra,which were effectively mitigated by sAT.sAT increased the mRNA and protein expression levels of apelin and its receptor apelin/apelin receptors(ARs)both in vivo and in vitro.(Ala13)-Apelin-13(F13A)and small interfering RNA(siRNA)abolished the regulatory effects of sAT on neuroprotection mediated by adenosine 5'-monophosphate(AMP)-activated protein kinase(AMPK)/protein kinase B(Akt).Furthermore,sAT induced apelin/AR expression by simultaneously inhibiting P38 mitogen-activated protein kinase(P38 MAPK)/activating transcription factor 4(ATF4)and upregulating hypoxia-inducible factor-1α(HIF-1α).Our findings indicate that sAT regulates apelin/AR/AMPK by inhibiting P38 MAPK/ATF4 and upregulating HIF-1a,thereby suppressing oxidative stress and mitochondrial dysfunction.
基金Supported by National Natural Science Foundation of China(No.81973908)Natural Science Foundation of Heilongjiang Province(No.PL2024H224)Postgraduate Innovation Fund of Heilongjiang University of Chinese Medicine(No.2023yjscx018).
文摘AIM:To investigate the effects of a Chinese medicine formula“Qingxuan Runmu Yin”(QRY)on ocular surface inflammation in a rat model of dry eye,and its mechanism via the toll-like receptor 4(TLR4)/transforming growth factor kinase 1(TAK1)/p38 mitogen-activated protein kinase(p38MAPK)signaling pathway.METHODS:Seventy-two Sprague-Dawley rats were randomly divided into six groups(n=12 each):the control group,model group,3 groups of QRY(with low-,medium-,and high-doses),and SB203580 group.Dry eye was induced using benzalkonium chloride.Schirmer’s test(SIT)and corneal fluorescein staining(CFS)were performed every 14d throughout the experiment.Histopathological changes in corneal and conjunctival tissues were observed using hematoxylin and eosin(HE)and periodic acid-Schiff(PAS)staining.Protein expression levels of key inflammatory markers and signaling pathway targets were assessed via immunohistochemistry,ELISA,and Western blotting.RESULTS:Compared to the control group,the model group showed significant reductions in SIT and increases in CFS scores,alongside structural disorganization of corneal/conjunctival tissues,decreased conjunctival goblet cell(CGC)numbers,and elevated expression of inflammatory markers[interleukin(IL)-1β,IL-6,tumor necrosis factoralpha(TNF-α),matrix metalloproteinase-9(MMP9)]and pathway proteins(TLR4,p-TAK1,p-p38MAPK;P<0.05).Treatment with QRY(low,medium,and high doses)and SB203580 significantly improved SIT scores,reduced CFS scores,restored corneoconjunctival structure,increased CGC numbers,and decreased expression levels of IL-1β,IL-6,TNF-α,MMP9,TLR4,p-TAK1,and p-p38MAPK proteins compared to the model group(P<0.05).CONCLUSION:QRY may alleviate ocular surface inflammation associated with dry eye by inhibiting the TLR4/TAK1/p38MAPK signaling pathway,highlighting its potential therapeutic efficacy for dry eye.
