AIM:To compare the effect of vegetable oils on the uptake of lutein and zeaxanthin by adult retinal pigment epithelial(ARPE)-19 cells in vitro.METHODS:ARPE-19 cells were cultured in Dulbecco’s Modified Eagle Medium-F...AIM:To compare the effect of vegetable oils on the uptake of lutein and zeaxanthin by adult retinal pigment epithelial(ARPE)-19 cells in vitro.METHODS:ARPE-19 cells were cultured in Dulbecco’s Modified Eagle Medium-F-12 supplemented with 10%foetal bovine serum and 1%penicillin–streptomycin in a humidified 5%CO_(2) incubator maintained at 37℃.Cells were treated with 247μmol/L lutein,49μmol/L zeaxanthin and 1%(v/v)of either coconut oil,corn oil,peanut oil,olive oil,sunflower oil,soybean oil,castor oil,or linseed oil for 48h.Lutein and zeaxanthin concentration in the cells were quantified by high performance liquid chromatography.RESULTS:Among the oils tested,the highest lutein and zeaxanthin uptake was observed with coconut oil while the lowest was observed with linseed oil.CONCLUSION:ARPE-19 uptake of lutein and zeaxanthin are found to be dependent on the type of oils.展开更多
AIM:To investigate the effects of nicotinamide(NAM)on bevacizumab(BEV)-induced epithelial-mesenchymal transition(EMT)of human retinal pigment epithelial cells(ARPE-19)and the underling mechanisms.METHODS:ARPE-19 cells...AIM:To investigate the effects of nicotinamide(NAM)on bevacizumab(BEV)-induced epithelial-mesenchymal transition(EMT)of human retinal pigment epithelial cells(ARPE-19)and the underling mechanisms.METHODS:ARPE-19 cells were treated with BEV for 24,48,and 72 h,and the variation degrees of EMTrelated markers(fibronectin,α-SMA,vimentin,and ZO-1)were assessed by Western blotting to select the optimal treatment time point which exhibited the most obvious changes of EMT-related markers for the subsequent experiments.Furthermore,NAM was added to the medium,the m RNA and protein levels of the EMT-related markers were then measured.The accumulation of reactive oxygen species(ROS)and H_(2)O_(2) and the total antioxidant capacity(TAC)of the cells were also measured to evaluate the level of oxidative stress.RESULTS:After being treated with BEV for 72 h,the protein expression levels of EMT-related markers in ARPE-19 cells showed significant changes.Meanwhile the levels of ROS and H_(2)O_(2) were obviously increased,and the TAC of ARPE-19 cells was decreased.Totally 72 h was chosen to be the optimal treatment time point in subsequentexperiments.Furthermore,NAM inhibited BEV-induced EMT by downregulating fibronectin,α-SMA,and vimentin and upregulating ZO-1,decreased the accumulation of ROS and H_(2)O_(2),and enhanced TAC in BEV-treated ARPE-19 cells.CONCLUSION:This study demonstrates that NAM suppressed BEV-induced EMT in ARPE-19 cells by attenuating oxidative stress.Hence,NAM may be a potential therapeutic agent for alleviating neovascular fibrosis of the ocular fundus after anti-vascular endothelial growth factor therapy.展开更多
Objective Oxidative stress plays an important role in retinal pigmental epithelium （RPE） death during aging and the development of age-related macular degeneration.Although early reports indicate that reactive oxyge...Objective Oxidative stress plays an important role in retinal pigmental epithelium （RPE） death during aging and the development of age-related macular degeneration.Although early reports indicate that reactive oxygen species （ROS） including H2O2 can trigger apoptosis at lower concentrations and necrosis at higher concentrations,the exact molecular mechanism of RPE death is still unclear.The purpose of this study was to investigate the molecular pathways involved in RPE death induced by exogenous ROS,especially at higher concentrations.Methods Cultured ARPE-19 cells were treated with H2O2 at different concentrations and cell viability was measured with the MTT assay.Cell death was morphologically studied by microscopy using APOPercentage assay and PI staining.Furthermore,the impact of oxidative stress on ARPE-19 cells was assessed by HO-1 and PARP-1 Western blotting and by the protection of antioxidant EGCG.Calcium influx was determined using the fura-2 calcium indicator and the role of intracellular calcium overload in ARPE-19 cell death was evaluated following cobalt treatment to block calcium effects.Results H2O2 reduced the viability of ARPE-19 cells in a concentration-dependent manner,which was presented as a typical s-shaped curve.Cell death caused by high concentrations of H2O2 was confirmed to be programmed necrosis.Morphologically,dying ARPE-19 cells were extremely swollen and lost the integrity of their plasma membrane,positively detected with APOPercentage assay and PI staining.24-hour treatment with 500 ？mol/L H2O2 induced remarkable up-regulation of HO-1 and PARP-1 in ARPE-19 cells.Moreover,antioxidant treatment using EGCG effectively protected cells from H2O2-induced injury,increasing cell viability from 14.17%±2.31% to 85.77%±4.58%.After H2O2 treatment,intracellular calcium levels were highly elevated with a maximum concentration of 1200nM.Significantly,the calcium channel inhibitor cobalt was able to blunt this calcium influx and blocked the necrotic pathway,rescuing the ARPE-19 cell from H2O2-induced death.Conclusions At high concentrations,H2O2 induces ARPE-19 cell death through a regulated necrotic pathway with calcium overload as a critical step in the cell death program.展开更多
AIM: To observe the protective effect of astaxanthin(AST) against hydroquinone(HQ) mediated cell death in the apoptotic cascade and evaluate intracellular Ca2+ release, caspase-3, and-9 activation, reactive oxygen spe...AIM: To observe the protective effect of astaxanthin(AST) against hydroquinone(HQ) mediated cell death in the apoptotic cascade and evaluate intracellular Ca2+ release, caspase-3, and-9 activation, reactive oxygen species(ROS) production in ARPE-19 cells.METHODS: We cultured ARPE-19 cells in special mediums and performed MTT tests to determine protective effect of AST, before exposing the cells to HQ in an incubator. We analyzed intracellular Ca2+ release experiments, mitochondrial membrane depolarization, glutathione(GSH), glutathione peroxidase(GSH-Px) and ROS experiments, and apoptosis assay.RESULTS: ROS production ranges depend on the amount of cell death. We computed the correlation between ROS ranges and cell death by 20,70-dichlorofluorescein fluorescence, and Ca2+ levels by Fura-2-AM. HQ-induced cell death found out to rise ranges of caspase-3 and-9, and mitochondrial depolarization. These three steps were delayed by AST management.CONCLUSION: ARPE-19 cells are avoided from HQinduced ROS production and caspase-3 and-9 activation by AST. AST may limit the range of caspase synthesis, Ca2+ release and excess production of ROS with antiapoptotic effect. This study proposes a new therapeutic approach for the treatment of age-related macular degeneration.展开更多
Stem cell-based therapies have been proposed as a potential treatment for neural regeneration following closed head injury.We previously reported that induced neural stem cells exert beneficial effects on neural regen...Stem cell-based therapies have been proposed as a potential treatment for neural regeneration following closed head injury.We previously reported that induced neural stem cells exert beneficial effects on neural regeneration via cell replacement.However,the neural regeneration efficiency of induced neural stem cells remains limited.In this study,we explored differentially expressed genes and long non-coding RNAs to clarify the mechanism underlying the neurogenesis of induced neural stem cells.We found that H19 was the most downregulated neurogenesis-associated lnc RNA in induced neural stem cells compared with induced pluripotent stem cells.Additionally,we demonstrated that H19 levels in induced neural stem cells were markedly lower than those in induced pluripotent stem cells and were substantially higher than those in induced neural stem cell-derived neurons.We predicted the target genes of H19 and discovered that H19 directly interacts with mi R-325-3p,which directly interacts with Ctbp2 in induced pluripotent stem cells and induced neural stem cells.Silencing H19 or Ctbp2 impaired induced neural stem cell proliferation,and mi R-325-3p suppression restored the effect of H19 inhibition but not the effect of Ctbp2 inhibition.Furthermore,H19 silencing substantially promoted the neural differentiation of induced neural stem cells and did not induce apoptosis of induced neural stem cells.Notably,silencing H19 in induced neural stem cell grafts markedly accelerated the neurological recovery of closed head injury mice.Our results reveal that H19 regulates the neurogenesis of induced neural stem cells.H19 inhibition may promote the neural differentiation of induced neural stem cells,which is closely associated with neurological recovery following closed head injury.展开更多
BACKGROUND Accumulating evidence has shown that adipose tissue-derived mesenchymal stem cells(ADSCs)are an effective therapeutic approach for managing coronavirus disease 2019(COVID-19);however,further elucidation is ...BACKGROUND Accumulating evidence has shown that adipose tissue-derived mesenchymal stem cells(ADSCs)are an effective therapeutic approach for managing coronavirus disease 2019(COVID-19);however,further elucidation is required to determine their underlying immunomodulatory effect on the mRNA expression of T helper cell-related transcription factors(TFs)and cytokine release in peripheral blood mononuclear cells(PBMCs).AIM To investigate the impact of ADSCs on the mRNA expression of TFs and cytokine release in PBMCs from colorectal cancer(CRC)patients with severe COVID-19(CRC^(+)patients).METHODS PBMCs from CRC^(+)patients(PBMCs-C+)and age-matched CRC patients(PBMCs-C)were stimulated and cultured in the presence/absence of ADSCs.The mRNA levels of T-box TF TBX21(T-bet),GATA binding protein 3(GATA-3),RAR-related orphan receptor C(RORC),and forkhead box P3(FoxP3)in the PBMCs were determined by reverse transcriptase-polymerase chain reaction.Culture supernatants were evaluated for levels of interferon gamma(IFN-γ),interleukin 4(IL-4),IL-17A,and transforming growth factor beta 1(TGF-β1)using an enzyme-linked immunosorbent assay.RESULTS Compared with PBMCs-C,PBMCs-C+exhibited higher mRNA levels of T-bet and RORC,and increased levels of IFN-γ and IL-17A.Additionally,a significant decrease in FoxP3 mRNA and TGF-β1,as well as an increase in Tbet/GATA-3,RORC/FoxP3,IFN-γ/IL-4,and IL-17A/TGF-β1 ratios were observed in PBMCs-C+.Furthermore,ADSCs significantly induced a functional regulatory T cell(Treg)subset,as evidenced by an increase in FoxP3 mRNA and TGF-β1 release levels.This was accompanied by a significant decrease in the mRNA levels of T-bet and RORC,release of IFN-γ and IL-17A,and T-bet/GATA-3,RORC/FoxP3,IFN-γ/IL-4,and IL-17A/TGF-β1 ratios,compared with the PBMCs-C+alone.CONCLUSION The present in vitro studies showed that ADSCs contributed to the immunosuppressive effects on PBMCs-C+,favoring Treg responses.Thus,ADSC-based cell therapy could be a beneficial approach for patients with severe COVID-19 who fail to respond to conventional therapies.展开更多
Objective Obesity-induced kidney injury contributes to the development of diabetic nephropathy(DN).Here,we identified the functions of ubiquitin-specific peptidase 19(USP19)in HK-2 cells exposed to a combination of hi...Objective Obesity-induced kidney injury contributes to the development of diabetic nephropathy(DN).Here,we identified the functions of ubiquitin-specific peptidase 19(USP19)in HK-2 cells exposed to a combination of high glucose(HG)and free fatty acid(FFA)and determined its association with TGF-beta-activated kinase 1(TAK1).Methods HK-2 cells were exposed to a combination of HG and FFA.USP19 mRNA expression was detected by quantitative RT-PCR(qRT-PCR),and protein analysis was performed by immunoblotting(IB).Cell growth was assessed by Cell Counting Kit-8(CCK-8)viability and 5-ethynyl-2′-deoxyuridine(EdU)proliferation assays.Cell cycle distribution and apoptosis were detected by flow cytometry.The USP19/TAK1 interaction and ubiquitinated TAK1 levels were assayed by coimmunoprecipitation(Co-IP)assays and IB.Results In HG+FFA-challenged HK-2 cells,USP19 was highly expressed.USP19 knockdown attenuated HG+FFA-triggered growth inhibition and apoptosis promotion in HK-2 cells.Moreover,USP19 knockdown alleviated HG+FFA-mediated PTEN-induced putative kinase 1(PINK1)/Parkin pathway inactivation and increased mitochondrial reactive oxygen species(ROS)generation in HK-2 cells.Mechanistically,USP19 stabilized the TAK1 protein through deubiquitination.Importantly,increased TAK1 expression reversed the USP19 knockdown-mediated phenotypic changes and PINK1/Parkin pathway activation in HG+FFA-challenged HK-2 cells.Conclusion The findings revealed that USP19 plays a crucial role in promoting HK-2 cell dysfunction induced by combined stimulation with HG and FFAs by stabilizing TAK1,providing a potential therapeutic strategy for combating DN.展开更多
AIM:To investigate the effect of acetyl-L-carnitine(ALCAR)on cell viability,morphological integrity,and vascular endothelial growth factor(VEGF)expression in human retinal pigment epithelium(ARPE-19)cells using a hypo...AIM:To investigate the effect of acetyl-L-carnitine(ALCAR)on cell viability,morphological integrity,and vascular endothelial growth factor(VEGF)expression in human retinal pigment epithelium(ARPE-19)cells using a hypoxic model.METHODS:In the first set of experiments,the optimal CoCl_(2) dose was determined by exposing ARPE-19 cell cultures to different concentrations.To evaluate the effect of ALCAR on cell viability,five groups of ARPE-19 cell culture were established that included a control group,a sham group(200μM CoCl_(2)),and groups that received 1,10 and 100 mM doses of ALCAR combined with 200μM CoCl_(2),respectively.The cell viability was measured by MTT assay.The morphological characteristics of cells were observed by an inverted phase contrast microscope.The levels of VEGF and HIF-1α secretion by ARPE-19 cells were detected by enzyme linked immunosorbent assay(ELISA)assay.RESULTS:ARPE-19 cells were exposed to different doses of CoCl_(2) in order to create a hypoxia model.Nevertheless,when exposed to a concentration of 200μM CoCl_(2),a notable decrease in viability to 83% was noted.ALCAR was found to increase the cell viability at 1 mM and 10 mM concentrations,while the highest concentration(100 mM)did not have an added effect.The cell viability was found to be significantly higher in the groups treated with a concentration of 1 mM and 10 mM ALCAR compared to the Sham group(P=0.041,P=0.019,respectively).The cell viability and morphology remained unaffected by the greatest dose of ALCAR(100 mM).The administration of 10 mM ALCAR demonstrated a statistically significant reduction in the levels of VEGF and HIF-1α compared with the Sham group(P=0.013,P=0.033,respectively).CONCLUSION:The findings from the current study indicate that ALCAR could represent a viable therapeutic option with the potential to open up novel treatment pathways for retinal diseases,particular relevance for age-related macular degeneration(AMD).However,to fully elucidate ALCAR’s application potential in retinal diseases,additional investigation is necessary to clearly define the exact mechanisms involved.展开更多
Coronavirus disease 2019(COVID-19)is an acute respiratory infection caused by severe acute respiratory syndrome coronavirus 2(SARS-CoV-2).SARS-CoV-2 infection typically presents with fever and respiratory symptoms,whi...Coronavirus disease 2019(COVID-19)is an acute respiratory infection caused by severe acute respiratory syndrome coronavirus 2(SARS-CoV-2).SARS-CoV-2 infection typically presents with fever and respiratory symptoms,which can progress to severe respiratory distress syndrome and multiple organ failure.In severe cases,these complications may even lead to death.One of the causes of COVID-19 deaths is the cytokine storm caused by an overactive immune response.Therefore,suppressing the overactive immune response may be an effective strategy for treating COVID-19.Mesenchymal stem cells(MSCs)and their derived exosomes(MSCs-Exo)have potent homing abilities,immunomodulatory functions,regenerative repair,and antifibrotic effects,promising an effective tool in treating COVID-19.In this paper,we review the main mechanisms and potential roles of MSCs and MSCs-Exo in treating COVID-19.We also summarize relevant recent clinical trials,including the source of cells,the dosage and the efficacy,and the clinical value and problems in this field,providing more theoretical references for the clinical use of MSCs and MSCs-Exo in the treatment of COVID-19.展开更多
Objective:This study aimed to compare the efficacy of anti-CD19 chimeric antigen receptor T cells(CAR-T cells)versus chemotherapy plus donor lymphocyte infusion(chemo-DLI)for treating relapsed CD 19-positive B-cell ac...Objective:This study aimed to compare the efficacy of anti-CD19 chimeric antigen receptor T cells(CAR-T cells)versus chemotherapy plus donor lymphocyte infusion(chemo-DLI)for treating relapsed CD 19-positive B-cell acute lymphoblastic leukemia(B-ALL)after allogeneic hematopoietic stem cell transplantation(allo-HSCT).Methods:Clinical data of 43 patients with B-ALL who relapsed after allo-HSCT were retrospectively analyzed.Twenty-two patients were treated with CAR-T cells(CAR-T group),and 21 with chemotherapy plus DLI(chemo-DLI group).The complete remission(CR)and minimal residual disease(MRD)-negative CR rates,leukemia-free survival(LFS)rate,overall survival(OS)rate,and incidence of acute graft-versus-host disease(aGVHD),cytokine release syndrome(CRS)and immune effector cell-associated neurotoxicity syndrome(ICANS)were compared between the two groups.Results:The CR and MRD-negative CR rates in the CAR-T group(77.3%and 61.5%)were significantly higher than those in the chemo-DLI group(38.1%and 23.8%)(P=0.008 and P=0.003).The 1-and 2-year LFS rates in the CAR-T group were superior to those in the chemo-DLI group:54.5%and 50.0%vs.9.5%and 4.8%(P=0.0001 and P=0.00004).The 1-and 2-year OS rates in the CAR-T versus chemo-DLI group were 59.1%and 54.5%vs.19%and 9.5%(P=0.011 and P=0.003).Six patients(28.6%)with grade 2-4 aGVHD were identified in the chemo-DLI group.Two patients(9.1%)in the CAR-T group developed grade 1-2 aGVHD.Nineteen patients(86.4%)developed CRS in the CAR-T group,comprising grade 1-2 CRS in 13 patients(59.1%)and grade 3 CRS in 6 patients(27.3%).Two patients(9.1%)developed grade 1-2 ICANS.Conclusion:Donor-derived anti-CD19 CAR-T-cell therapy may be better,safer,and more effective than chemo-DLI for B-ALL patients who relapse after allo-HSCT.展开更多
The cancer cell metastasis is a major death reason for patients with non-small cell lung cancer(NSCLC).Although researchers have disclosed that interleukin 17(IL-17)can increase matrix metalloproteinases(MMPs)inductio...The cancer cell metastasis is a major death reason for patients with non-small cell lung cancer(NSCLC).Although researchers have disclosed that interleukin 17(IL-17)can increase matrix metalloproteinases(MMPs)induction causing NSCLC cell metastasis,the underlying mechanism remains unclear.In the study,we found that IL-17 receptor A(IL-17RA),p300,p-STAT3,Ack-STAT3,and MMP19 were up-regulated both in NSCLC tissues and NSCLC cells stimulated with IL-17.p300,STAT3 and MMP19 overexpression or knockdown could raise or reduce IL-17-induced p-STAT3,Ack-STAT3 and MMP19 level as well as the cell migration and invasion.Mechanism investigation revealed that STAT3 and p300 bound to the same region(−544 to−389 nt)of MMP19 promoter,and p300 could acetylate STAT3-K631 elevating STAT3 transcriptional activity,p-STAT3 or MMP19 expression and the cell mobility exposed to IL-17.Meanwhile,p300-mediated STAT3-K631 acetylation and its Y705-phosphorylation could interact,synergistically facilitating MMP19 gene transcription and enhancing cell migration and invasion.Besides,the animal experiments exhibited that the nude mice inoculated with NSCLC cells by silencing p300,STAT3 or MMP19 gene plus IL-17 treatment,the nodule number,and MMP19,Ack-STAT3,or p-STAT3 production in the lung metastatic nodules were all alleviated.Collectively,these outcomes uncover that IL-17-triggered NSCLC metastasis involves up-regulating MMP19 expression via the interaction of STAT3-K631 acetylation by p300 and its Y705-phosphorylation,which provides a new mechanistic insight and potential strategy for NSCLC metastasis and therapy.展开更多
Objective:To evaluate the efficacy of ponatinib plus gossypol against colorectal cancer HCT-116 and Caco-2 cells.Methods:Cells were treated with ponatinib and/or gossypol at increasing concentrations to evaluate syner...Objective:To evaluate the efficacy of ponatinib plus gossypol against colorectal cancer HCT-116 and Caco-2 cells.Methods:Cells were treated with ponatinib and/or gossypol at increasing concentrations to evaluate synergistic drug interactions by combination index.Cell viability,FGF19/FGFR4,and apoptotic and autophagic cell death were studied.Results:Ponatinib(1.25-40μM)and gossypol(2.5-80μM)monotherapy inhibited HCT-116 and Caco-2 cell viability in a doseand time-dependent manner.The combination of ponatinib and gossypol at a ratio of 1 to 2 significantly decreased cell viability(P<0.05),with a>2-and>4-fold reduction in IC50,respectively,after 24 h and 48 h,as compared to the IC50 of ponatinib.Lower combined concentrations showed greater synergism(combination index<1)with a higher ponatinib dose reduction index.Moreover,ponatinib plus gossypol induced morphological changes in HCT-116and Caco-2 cells,increased beclin-1 and caspase-3,and decreased FGF19,FGFR4,Bcl-2 and p-Akt as compared to treatment with drugs alone.Conclusions:Gossypol enhances ponatinib's anticancer effects against colorectal cancer cells through antiproliferative,apoptotic,and autophagic mechanisms.This may open the way for the future use of ponatinib at lower doses with gossypol as a potentially safer targeted strategy for colorectal cancer treatment.展开更多
BACKGROUND Severe acute respiratory syndrome coronavirus 2 is the virus responsible for coronavirus disease 2019(COVID-19),a disease that has been blamed for inducing or exacerbating symptoms in patients with autoimmu...BACKGROUND Severe acute respiratory syndrome coronavirus 2 is the virus responsible for coronavirus disease 2019(COVID-19),a disease that has been blamed for inducing or exacerbating symptoms in patients with autoimmune diseases.Crohn's disease(CD)is an inflammatory bowel disease that affects genetically susceptible patients who develop an abnormal mucosal immune response to the intestinal microbiota.Patients who underwent hematopoietic stem cell transplantation(HSCT)are considered at risk for COVID-19.AIM To describe for the first time the impact of COVID-19 in CD patients who had undergone autologous,non-myeloablative HSCT.METHODS In this descriptive study a series of 19 patients were diagnosed with positive COVID-19.For two patients there were reports of the occurrence of two infectious episodes.Parameters related to HSCT,such as time elapsed since the procedure,vaccination status,CD status before and after infection,and clinical manifestations resulting from COVID-19,were evaluated.RESULTS Among the patients with COVID-19,three,who underwent Auto HSCT less than six months ago,relapsed and one,in addition to the CD symptoms,started to present thyroid impairment with positive anti-TPO.Only one of the patients required hospitalization for five days to treat COVID-19 and remained in CD clinical remission.Nine patients reported late symptoms that may be related to COVID-19.There were no deaths,and a statistical evaluation of the series of COVID-19 patients compared to those who did not present any infectious episode did not identify significant differences regarding the analyzed parameters.CONCLUSION Despite the change in CD status in three patients and the presence of nine patients with late symptoms,we can conclude that there was no significant adverse impact concerning COVID-19 in the evaluated patients who underwent HSCT to treat CD.展开更多
文摘AIM:To compare the effect of vegetable oils on the uptake of lutein and zeaxanthin by adult retinal pigment epithelial(ARPE)-19 cells in vitro.METHODS:ARPE-19 cells were cultured in Dulbecco’s Modified Eagle Medium-F-12 supplemented with 10%foetal bovine serum and 1%penicillin–streptomycin in a humidified 5%CO_(2) incubator maintained at 37℃.Cells were treated with 247μmol/L lutein,49μmol/L zeaxanthin and 1%(v/v)of either coconut oil,corn oil,peanut oil,olive oil,sunflower oil,soybean oil,castor oil,or linseed oil for 48h.Lutein and zeaxanthin concentration in the cells were quantified by high performance liquid chromatography.RESULTS:Among the oils tested,the highest lutein and zeaxanthin uptake was observed with coconut oil while the lowest was observed with linseed oil.CONCLUSION:ARPE-19 uptake of lutein and zeaxanthin are found to be dependent on the type of oils.
基金Supported by the National Natural Science Foundation of China(No.81670828)the Shandong Provincial Key Research and Development Program(No.2017GSF18141)+1 种基金the Innovation Project of Shandong Academy of Medical Sciences and the National Science and Technology Major Project of China(No.2017ZX09304-010)supported by the Taishan Scholar Youth Professional Program(No.tspd20150215,No.tsqn20161059)。
文摘AIM:To investigate the effects of nicotinamide(NAM)on bevacizumab(BEV)-induced epithelial-mesenchymal transition(EMT)of human retinal pigment epithelial cells(ARPE-19)and the underling mechanisms.METHODS:ARPE-19 cells were treated with BEV for 24,48,and 72 h,and the variation degrees of EMTrelated markers(fibronectin,α-SMA,vimentin,and ZO-1)were assessed by Western blotting to select the optimal treatment time point which exhibited the most obvious changes of EMT-related markers for the subsequent experiments.Furthermore,NAM was added to the medium,the m RNA and protein levels of the EMT-related markers were then measured.The accumulation of reactive oxygen species(ROS)and H_(2)O_(2) and the total antioxidant capacity(TAC)of the cells were also measured to evaluate the level of oxidative stress.RESULTS:After being treated with BEV for 72 h,the protein expression levels of EMT-related markers in ARPE-19 cells showed significant changes.Meanwhile the levels of ROS and H_(2)O_(2) were obviously increased,and the TAC of ARPE-19 cells was decreased.Totally 72 h was chosen to be the optimal treatment time point in subsequentexperiments.Furthermore,NAM inhibited BEV-induced EMT by downregulating fibronectin,α-SMA,and vimentin and upregulating ZO-1,decreased the accumulation of ROS and H_(2)O_(2),and enhanced TAC in BEV-treated ARPE-19 cells.CONCLUSION:This study demonstrates that NAM suppressed BEV-induced EMT in ARPE-19 cells by attenuating oxidative stress.Hence,NAM may be a potential therapeutic agent for alleviating neovascular fibrosis of the ocular fundus after anti-vascular endothelial growth factor therapy.
基金supported by grants from the Natural Science Foundation of China (30801271)the Changchun Science and Technology Development Fund (08SF39)the International Joint Project from Science and Technology Bureau of Jilin Province (20090746)
文摘Objective Oxidative stress plays an important role in retinal pigmental epithelium （RPE） death during aging and the development of age-related macular degeneration.Although early reports indicate that reactive oxygen species （ROS） including H2O2 can trigger apoptosis at lower concentrations and necrosis at higher concentrations,the exact molecular mechanism of RPE death is still unclear.The purpose of this study was to investigate the molecular pathways involved in RPE death induced by exogenous ROS,especially at higher concentrations.Methods Cultured ARPE-19 cells were treated with H2O2 at different concentrations and cell viability was measured with the MTT assay.Cell death was morphologically studied by microscopy using APOPercentage assay and PI staining.Furthermore,the impact of oxidative stress on ARPE-19 cells was assessed by HO-1 and PARP-1 Western blotting and by the protection of antioxidant EGCG.Calcium influx was determined using the fura-2 calcium indicator and the role of intracellular calcium overload in ARPE-19 cell death was evaluated following cobalt treatment to block calcium effects.Results H2O2 reduced the viability of ARPE-19 cells in a concentration-dependent manner,which was presented as a typical s-shaped curve.Cell death caused by high concentrations of H2O2 was confirmed to be programmed necrosis.Morphologically,dying ARPE-19 cells were extremely swollen and lost the integrity of their plasma membrane,positively detected with APOPercentage assay and PI staining.24-hour treatment with 500 ？mol/L H2O2 induced remarkable up-regulation of HO-1 and PARP-1 in ARPE-19 cells.Moreover,antioxidant treatment using EGCG effectively protected cells from H2O2-induced injury,increasing cell viability from 14.17%±2.31% to 85.77%±4.58%.After H2O2 treatment,intracellular calcium levels were highly elevated with a maximum concentration of 1200nM.Significantly,the calcium channel inhibitor cobalt was able to blunt this calcium influx and blocked the necrotic pathway,rescuing the ARPE-19 cell from H2O2-induced death.Conclusions At high concentrations,H2O2 induces ARPE-19 cell death through a regulated necrotic pathway with calcium overload as a critical step in the cell death program.
文摘AIM: To observe the protective effect of astaxanthin(AST) against hydroquinone(HQ) mediated cell death in the apoptotic cascade and evaluate intracellular Ca2+ release, caspase-3, and-9 activation, reactive oxygen species(ROS) production in ARPE-19 cells.METHODS: We cultured ARPE-19 cells in special mediums and performed MTT tests to determine protective effect of AST, before exposing the cells to HQ in an incubator. We analyzed intracellular Ca2+ release experiments, mitochondrial membrane depolarization, glutathione(GSH), glutathione peroxidase(GSH-Px) and ROS experiments, and apoptosis assay.RESULTS: ROS production ranges depend on the amount of cell death. We computed the correlation between ROS ranges and cell death by 20,70-dichlorofluorescein fluorescence, and Ca2+ levels by Fura-2-AM. HQ-induced cell death found out to rise ranges of caspase-3 and-9, and mitochondrial depolarization. These three steps were delayed by AST management.CONCLUSION: ARPE-19 cells are avoided from HQinduced ROS production and caspase-3 and-9 activation by AST. AST may limit the range of caspase synthesis, Ca2+ release and excess production of ROS with antiapoptotic effect. This study proposes a new therapeutic approach for the treatment of age-related macular degeneration.
基金supported by the National Natural Science Foundation of China,Nos.82271397(to MG),82001293(to MG),82171355(to RX),81971295(to RX)and 81671189(to RX)。
文摘Stem cell-based therapies have been proposed as a potential treatment for neural regeneration following closed head injury.We previously reported that induced neural stem cells exert beneficial effects on neural regeneration via cell replacement.However,the neural regeneration efficiency of induced neural stem cells remains limited.In this study,we explored differentially expressed genes and long non-coding RNAs to clarify the mechanism underlying the neurogenesis of induced neural stem cells.We found that H19 was the most downregulated neurogenesis-associated lnc RNA in induced neural stem cells compared with induced pluripotent stem cells.Additionally,we demonstrated that H19 levels in induced neural stem cells were markedly lower than those in induced pluripotent stem cells and were substantially higher than those in induced neural stem cell-derived neurons.We predicted the target genes of H19 and discovered that H19 directly interacts with mi R-325-3p,which directly interacts with Ctbp2 in induced pluripotent stem cells and induced neural stem cells.Silencing H19 or Ctbp2 impaired induced neural stem cell proliferation,and mi R-325-3p suppression restored the effect of H19 inhibition but not the effect of Ctbp2 inhibition.Furthermore,H19 silencing substantially promoted the neural differentiation of induced neural stem cells and did not induce apoptosis of induced neural stem cells.Notably,silencing H19 in induced neural stem cell grafts markedly accelerated the neurological recovery of closed head injury mice.Our results reveal that H19 regulates the neurogenesis of induced neural stem cells.H19 inhibition may promote the neural differentiation of induced neural stem cells,which is closely associated with neurological recovery following closed head injury.
基金Supported by National Natural Science Foundation of China,No.81470982.
文摘BACKGROUND Accumulating evidence has shown that adipose tissue-derived mesenchymal stem cells(ADSCs)are an effective therapeutic approach for managing coronavirus disease 2019(COVID-19);however,further elucidation is required to determine their underlying immunomodulatory effect on the mRNA expression of T helper cell-related transcription factors(TFs)and cytokine release in peripheral blood mononuclear cells(PBMCs).AIM To investigate the impact of ADSCs on the mRNA expression of TFs and cytokine release in PBMCs from colorectal cancer(CRC)patients with severe COVID-19(CRC^(+)patients).METHODS PBMCs from CRC^(+)patients(PBMCs-C+)and age-matched CRC patients(PBMCs-C)were stimulated and cultured in the presence/absence of ADSCs.The mRNA levels of T-box TF TBX21(T-bet),GATA binding protein 3(GATA-3),RAR-related orphan receptor C(RORC),and forkhead box P3(FoxP3)in the PBMCs were determined by reverse transcriptase-polymerase chain reaction.Culture supernatants were evaluated for levels of interferon gamma(IFN-γ),interleukin 4(IL-4),IL-17A,and transforming growth factor beta 1(TGF-β1)using an enzyme-linked immunosorbent assay.RESULTS Compared with PBMCs-C,PBMCs-C+exhibited higher mRNA levels of T-bet and RORC,and increased levels of IFN-γ and IL-17A.Additionally,a significant decrease in FoxP3 mRNA and TGF-β1,as well as an increase in Tbet/GATA-3,RORC/FoxP3,IFN-γ/IL-4,and IL-17A/TGF-β1 ratios were observed in PBMCs-C+.Furthermore,ADSCs significantly induced a functional regulatory T cell(Treg)subset,as evidenced by an increase in FoxP3 mRNA and TGF-β1 release levels.This was accompanied by a significant decrease in the mRNA levels of T-bet and RORC,release of IFN-γ and IL-17A,and T-bet/GATA-3,RORC/FoxP3,IFN-γ/IL-4,and IL-17A/TGF-β1 ratios,compared with the PBMCs-C+alone.CONCLUSION The present in vitro studies showed that ADSCs contributed to the immunosuppressive effects on PBMCs-C+,favoring Treg responses.Thus,ADSC-based cell therapy could be a beneficial approach for patients with severe COVID-19 who fail to respond to conventional therapies.
基金supported by Natural Science Foundation of Shaanxi Province(No.2023-JC-YB-743 and No.2021JQ-905).
文摘Objective Obesity-induced kidney injury contributes to the development of diabetic nephropathy(DN).Here,we identified the functions of ubiquitin-specific peptidase 19(USP19)in HK-2 cells exposed to a combination of high glucose(HG)and free fatty acid(FFA)and determined its association with TGF-beta-activated kinase 1(TAK1).Methods HK-2 cells were exposed to a combination of HG and FFA.USP19 mRNA expression was detected by quantitative RT-PCR(qRT-PCR),and protein analysis was performed by immunoblotting(IB).Cell growth was assessed by Cell Counting Kit-8(CCK-8)viability and 5-ethynyl-2′-deoxyuridine(EdU)proliferation assays.Cell cycle distribution and apoptosis were detected by flow cytometry.The USP19/TAK1 interaction and ubiquitinated TAK1 levels were assayed by coimmunoprecipitation(Co-IP)assays and IB.Results In HG+FFA-challenged HK-2 cells,USP19 was highly expressed.USP19 knockdown attenuated HG+FFA-triggered growth inhibition and apoptosis promotion in HK-2 cells.Moreover,USP19 knockdown alleviated HG+FFA-mediated PTEN-induced putative kinase 1(PINK1)/Parkin pathway inactivation and increased mitochondrial reactive oxygen species(ROS)generation in HK-2 cells.Mechanistically,USP19 stabilized the TAK1 protein through deubiquitination.Importantly,increased TAK1 expression reversed the USP19 knockdown-mediated phenotypic changes and PINK1/Parkin pathway activation in HG+FFA-challenged HK-2 cells.Conclusion The findings revealed that USP19 plays a crucial role in promoting HK-2 cell dysfunction induced by combined stimulation with HG and FFAs by stabilizing TAK1,providing a potential therapeutic strategy for combating DN.
文摘AIM:To investigate the effect of acetyl-L-carnitine(ALCAR)on cell viability,morphological integrity,and vascular endothelial growth factor(VEGF)expression in human retinal pigment epithelium(ARPE-19)cells using a hypoxic model.METHODS:In the first set of experiments,the optimal CoCl_(2) dose was determined by exposing ARPE-19 cell cultures to different concentrations.To evaluate the effect of ALCAR on cell viability,five groups of ARPE-19 cell culture were established that included a control group,a sham group(200μM CoCl_(2)),and groups that received 1,10 and 100 mM doses of ALCAR combined with 200μM CoCl_(2),respectively.The cell viability was measured by MTT assay.The morphological characteristics of cells were observed by an inverted phase contrast microscope.The levels of VEGF and HIF-1α secretion by ARPE-19 cells were detected by enzyme linked immunosorbent assay(ELISA)assay.RESULTS:ARPE-19 cells were exposed to different doses of CoCl_(2) in order to create a hypoxia model.Nevertheless,when exposed to a concentration of 200μM CoCl_(2),a notable decrease in viability to 83% was noted.ALCAR was found to increase the cell viability at 1 mM and 10 mM concentrations,while the highest concentration(100 mM)did not have an added effect.The cell viability was found to be significantly higher in the groups treated with a concentration of 1 mM and 10 mM ALCAR compared to the Sham group(P=0.041,P=0.019,respectively).The cell viability and morphology remained unaffected by the greatest dose of ALCAR(100 mM).The administration of 10 mM ALCAR demonstrated a statistically significant reduction in the levels of VEGF and HIF-1α compared with the Sham group(P=0.013,P=0.033,respectively).CONCLUSION:The findings from the current study indicate that ALCAR could represent a viable therapeutic option with the potential to open up novel treatment pathways for retinal diseases,particular relevance for age-related macular degeneration(AMD).However,to fully elucidate ALCAR’s application potential in retinal diseases,additional investigation is necessary to clearly define the exact mechanisms involved.
基金Supported by Science and Technology Department Project of Jilin Province,China,No.20230101163JCthe Outstanding Youth Fund Project of Jilin Provincial Department of Education,China,No.JJKH20241324KJ.
文摘Coronavirus disease 2019(COVID-19)is an acute respiratory infection caused by severe acute respiratory syndrome coronavirus 2(SARS-CoV-2).SARS-CoV-2 infection typically presents with fever and respiratory symptoms,which can progress to severe respiratory distress syndrome and multiple organ failure.In severe cases,these complications may even lead to death.One of the causes of COVID-19 deaths is the cytokine storm caused by an overactive immune response.Therefore,suppressing the overactive immune response may be an effective strategy for treating COVID-19.Mesenchymal stem cells(MSCs)and their derived exosomes(MSCs-Exo)have potent homing abilities,immunomodulatory functions,regenerative repair,and antifibrotic effects,promising an effective tool in treating COVID-19.In this paper,we review the main mechanisms and potential roles of MSCs and MSCs-Exo in treating COVID-19.We also summarize relevant recent clinical trials,including the source of cells,the dosage and the efficacy,and the clinical value and problems in this field,providing more theoretical references for the clinical use of MSCs and MSCs-Exo in the treatment of COVID-19.
基金supported by grants from the National Natural Science Foundation of China(No.82020108004)the Hospital-level Clinical Innovation Military-Civilian Special Project of Army Medical University(No.2018JSLC0020)+1 种基金Chongqing Science and Technology Innovation Leading Talent(No.CSTCCXLJRC201718)Natural Science Foundation of Chongqing Innovation Group Science Program(No.cstc2021jcyj-cxttX0001).
文摘Objective:This study aimed to compare the efficacy of anti-CD19 chimeric antigen receptor T cells(CAR-T cells)versus chemotherapy plus donor lymphocyte infusion(chemo-DLI)for treating relapsed CD 19-positive B-cell acute lymphoblastic leukemia(B-ALL)after allogeneic hematopoietic stem cell transplantation(allo-HSCT).Methods:Clinical data of 43 patients with B-ALL who relapsed after allo-HSCT were retrospectively analyzed.Twenty-two patients were treated with CAR-T cells(CAR-T group),and 21 with chemotherapy plus DLI(chemo-DLI group).The complete remission(CR)and minimal residual disease(MRD)-negative CR rates,leukemia-free survival(LFS)rate,overall survival(OS)rate,and incidence of acute graft-versus-host disease(aGVHD),cytokine release syndrome(CRS)and immune effector cell-associated neurotoxicity syndrome(ICANS)were compared between the two groups.Results:The CR and MRD-negative CR rates in the CAR-T group(77.3%and 61.5%)were significantly higher than those in the chemo-DLI group(38.1%and 23.8%)(P=0.008 and P=0.003).The 1-and 2-year LFS rates in the CAR-T group were superior to those in the chemo-DLI group:54.5%and 50.0%vs.9.5%and 4.8%(P=0.0001 and P=0.00004).The 1-and 2-year OS rates in the CAR-T versus chemo-DLI group were 59.1%and 54.5%vs.19%and 9.5%(P=0.011 and P=0.003).Six patients(28.6%)with grade 2-4 aGVHD were identified in the chemo-DLI group.Two patients(9.1%)in the CAR-T group developed grade 1-2 aGVHD.Nineteen patients(86.4%)developed CRS in the CAR-T group,comprising grade 1-2 CRS in 13 patients(59.1%)and grade 3 CRS in 6 patients(27.3%).Two patients(9.1%)developed grade 1-2 ICANS.Conclusion:Donor-derived anti-CD19 CAR-T-cell therapy may be better,safer,and more effective than chemo-DLI for B-ALL patients who relapse after allo-HSCT.
基金National Natural Science Foundation of China(Grants Numbers 81902878 and 81971468).
文摘The cancer cell metastasis is a major death reason for patients with non-small cell lung cancer(NSCLC).Although researchers have disclosed that interleukin 17(IL-17)can increase matrix metalloproteinases(MMPs)induction causing NSCLC cell metastasis,the underlying mechanism remains unclear.In the study,we found that IL-17 receptor A(IL-17RA),p300,p-STAT3,Ack-STAT3,and MMP19 were up-regulated both in NSCLC tissues and NSCLC cells stimulated with IL-17.p300,STAT3 and MMP19 overexpression or knockdown could raise or reduce IL-17-induced p-STAT3,Ack-STAT3 and MMP19 level as well as the cell migration and invasion.Mechanism investigation revealed that STAT3 and p300 bound to the same region(−544 to−389 nt)of MMP19 promoter,and p300 could acetylate STAT3-K631 elevating STAT3 transcriptional activity,p-STAT3 or MMP19 expression and the cell mobility exposed to IL-17.Meanwhile,p300-mediated STAT3-K631 acetylation and its Y705-phosphorylation could interact,synergistically facilitating MMP19 gene transcription and enhancing cell migration and invasion.Besides,the animal experiments exhibited that the nude mice inoculated with NSCLC cells by silencing p300,STAT3 or MMP19 gene plus IL-17 treatment,the nodule number,and MMP19,Ack-STAT3,or p-STAT3 production in the lung metastatic nodules were all alleviated.Collectively,these outcomes uncover that IL-17-triggered NSCLC metastasis involves up-regulating MMP19 expression via the interaction of STAT3-K631 acetylation by p300 and its Y705-phosphorylation,which provides a new mechanistic insight and potential strategy for NSCLC metastasis and therapy.
基金financial support from the Theodor Bilharz Research InstituteWarrak El-Hadar+1 种基金ImbabaGiza 12411,Egypt。
文摘Objective:To evaluate the efficacy of ponatinib plus gossypol against colorectal cancer HCT-116 and Caco-2 cells.Methods:Cells were treated with ponatinib and/or gossypol at increasing concentrations to evaluate synergistic drug interactions by combination index.Cell viability,FGF19/FGFR4,and apoptotic and autophagic cell death were studied.Results:Ponatinib(1.25-40μM)and gossypol(2.5-80μM)monotherapy inhibited HCT-116 and Caco-2 cell viability in a doseand time-dependent manner.The combination of ponatinib and gossypol at a ratio of 1 to 2 significantly decreased cell viability(P<0.05),with a>2-and>4-fold reduction in IC50,respectively,after 24 h and 48 h,as compared to the IC50 of ponatinib.Lower combined concentrations showed greater synergism(combination index<1)with a higher ponatinib dose reduction index.Moreover,ponatinib plus gossypol induced morphological changes in HCT-116and Caco-2 cells,increased beclin-1 and caspase-3,and decreased FGF19,FGFR4,Bcl-2 and p-Akt as compared to treatment with drugs alone.Conclusions:Gossypol enhances ponatinib's anticancer effects against colorectal cancer cells through antiproliferative,apoptotic,and autophagic mechanisms.This may open the way for the future use of ponatinib at lower doses with gossypol as a potentially safer targeted strategy for colorectal cancer treatment.
文摘BACKGROUND Severe acute respiratory syndrome coronavirus 2 is the virus responsible for coronavirus disease 2019(COVID-19),a disease that has been blamed for inducing or exacerbating symptoms in patients with autoimmune diseases.Crohn's disease(CD)is an inflammatory bowel disease that affects genetically susceptible patients who develop an abnormal mucosal immune response to the intestinal microbiota.Patients who underwent hematopoietic stem cell transplantation(HSCT)are considered at risk for COVID-19.AIM To describe for the first time the impact of COVID-19 in CD patients who had undergone autologous,non-myeloablative HSCT.METHODS In this descriptive study a series of 19 patients were diagnosed with positive COVID-19.For two patients there were reports of the occurrence of two infectious episodes.Parameters related to HSCT,such as time elapsed since the procedure,vaccination status,CD status before and after infection,and clinical manifestations resulting from COVID-19,were evaluated.RESULTS Among the patients with COVID-19,three,who underwent Auto HSCT less than six months ago,relapsed and one,in addition to the CD symptoms,started to present thyroid impairment with positive anti-TPO.Only one of the patients required hospitalization for five days to treat COVID-19 and remained in CD clinical remission.Nine patients reported late symptoms that may be related to COVID-19.There were no deaths,and a statistical evaluation of the series of COVID-19 patients compared to those who did not present any infectious episode did not identify significant differences regarding the analyzed parameters.CONCLUSION Despite the change in CD status in three patients and the presence of nine patients with late symptoms,we can conclude that there was no significant adverse impact concerning COVID-19 in the evaluated patients who underwent HSCT to treat CD.
