Since the diccovery of neural stem cells(NSCs)in the embryonic and adult mammalian central nerous system,it provided novel ideas forneurogenesis as the potential of proliferation and differentiation of NSCs.One of the...Since the diccovery of neural stem cells(NSCs)in the embryonic and adult mammalian central nerous system,it provided novel ideas forneurogenesis as the potential of proliferation and differentiation of NSCs.One of the ways to promote the clinical application of neural stem cells(NSCs)is searching effective methods which regulate the proliferation and differentiation.This is also a problem urgently to be solved in medical field.Plenty of earlier studies have shown that traditional chinese medicine can promote the proliferation and differentiation of NSCs by regulating the related signaling pathway in vivo and in vitro.The reports of Chinese and foreign literatures on regulating the proliferation and differentiation of neural stem cells in recent ten years and their target and signaling pathways is analyzed in this review.The traditional chinese medicine regulate proliferation and differentiation of NSCs by the signaling pathways of Notch,PI3K/Akt,Wnt/β-catenin,and GFs.And,those signaling pathways have cross-talk in the regulation progress.Moreover,some traditional Chinese medicine,such as astragalus,has a variety of active ingredients to regulate proliferation and differentiation of NSCs through different signaling pathways.However,to accelerate the clinical application of neural stem cells,the studies aboutthe proliferation and differentiation of NSCs and Chinese medicine should be further deepened,the mechanism of multiple targets and the comprehensive regulation function of traditional Chinese medicine should be clarified.展开更多
The experiment was designed to study effects of retinoic acid and ascorbic acid on differentiation of mouse embryonic stem cells to cardiomyocytes. Embryonic bodies （EB） were developed from mESC in suspension cultur...The experiment was designed to study effects of retinoic acid and ascorbic acid on differentiation of mouse embryonic stem cells to cardiomyocytes. Embryonic bodies （EB） were developed from mESC in suspension culture, different levels of concentration of retinoic acid and ascorbic acid were used to determine the optimal conditions for EB formation. The results showed that the optimal concentrations were 10.9 mol. L-1 and 0.1 mg. mL-1 for retinoic acid and ascorbic acids, respectively. 50% of EB which was significantly （p〈0.05） different from the control group developed to cardiomyocytes. In conclusion, rctinoic acid and ascorbic acid had strong ability to promote cardiomyocyte differentiation of mouse embryonic stem cells. 10-9 mol. L-1 retinoic acid and 0.10 mg. mL-1 ascorbic acids were recommended to induce differentiation of mouse ES ceUs toward cardiomyocytes.展开更多
OBJECTIVE To explore the key mechanism of Bazi Bushen capsule(BZBS)in delaying the senescence of mesenchymal stem cells(MSCs)through network pharmacology and in vitro experiments.METHODS Network pharmacology was used ...OBJECTIVE To explore the key mechanism of Bazi Bushen capsule(BZBS)in delaying the senescence of mesenchymal stem cells(MSCs)through network pharmacology and in vitro experiments.METHODS Network pharmacology was used to predict the mechanism targets of BZBS in delaying MSCs senescence.A MSCs senescence model induced by D-galactose(D-gal)was used to investigate the effect and mechanism of BZBS on MSCs senescence in vitro.RESULTS Network pharmacology analy⁃sis showed that BZSB could delay MSCs senes⁃cence.The experiment showed that BZBS could significantly improve the survival activity of the aged MSCs.It significantly reduced the positive rate ofβ-galactosidase staining and p16,p21 expression in aged MSCs,enhanced the ability of adipogenic differentiation and osteogenic differentiation,and increased expression of Nanog,OCT4 and SOX2 in senescent MSCs.CONCLU⁃SIONS Network pharmacology and in vitro cell experiments verified that BZBS could delay MSCs senescence.展开更多
Embryonic stem (ES) cell biology is attracting much attention in cell biology because of their pluripotent behaviors and potential therapeutic applications. However,what maintains ES cell pluripotency and what trigger...Embryonic stem (ES) cell biology is attracting much attention in cell biology because of their pluripotent behaviors and potential therapeutic applications. However,what maintains ES cell pluripotency and what triggers ES cell展开更多
Objective Bone-marrow stem-cell transplantation has been shown to improve cardiac function in patients with AMI, but the safety of intracoronory infusion of autologous peripheral blood stem-cell(PBSCs) in patients wit...Objective Bone-marrow stem-cell transplantation has been shown to improve cardiac function in patients with AMI, but the safety of intracoronory infusion of autologous peripheral blood stem-cell(PBSCs) in patients with AMI is unknown. For this reason, we observe the feasibility and safety of PBSCs transplantation by intracoronory infusion in such patients.Method Fourty one patients with AMI were allocated to receive Granulocyte Colony-Stimulating Factor (G-CSF:Filgrastim,300 μg) with the dose of 300 μg-600 μg/day to mobilize the stem cell, and the duration of applying G-CSF was 5 days . On the sixth day, PBSCs were separated by Baxter CS 3000 blood cell separator into suspend liquid 57 ml. Then the suspend liquid was infused into the infarct related artery (IRA)by occluding the over the wire balloon and infusing artery through balloon center lumen. In the process of the intracoronary infusion of PBSCs, the complications should be observed, which were arrhythmias including of bradycardia, sinus arrest or atrial ventricular block, premature ventricular beats ,ventricular tachycardia, ventricular fibrillation; and hypotention, etc. Results There were total 10 cases with complications during the intracoronary infusion of PBSCs. The incidence of complications was 24.4%(10/41), including bradycardia is 2.4 %(1/41), sinus arrest or atrial ventricular block is 4.9%(2/41), ventricular fibrillation is 2.4 %( 1/41), hypotention is14.6 % (6 /41).Conclusions In patients with AMI, intracoronary infusion of PBSCs is feasible and safe.展开更多
Cancer stem cells(CSCs)are the driving force for sustainable tumor growth and metastasis and responsible for drug resistance and cancer relapse.Nanoparticle-based drug delivery has been demonstrated to be effective in...Cancer stem cells(CSCs)are the driving force for sustainable tumor growth and metastasis and responsible for drug resistance and cancer relapse.Nanoparticle-based drug delivery has been demonstrated to be effective in combating tumor growth.However,it has been challenging to selectively eliminate CSCs due to the lack of a general signature for a spectrum of cancers.It is known that CSCs from various types of cancer show lower stiffness compared to non-CSCs.It remains unclear whether low stiffness in CSCs influences cellular uptake in nanoparticle-based drug delivery and thus the chemotherapy efficacy.Graphene quantum dot(GQD)is emerging as a promising carrier material in delivering anti-cancer drugs.We found that breast CSCs were softer than conventional cancer cells,which were further softer compared to healthy breast tissue cells.Importantly,soft CSCs uptook more GQD than conventional cancer cells,while stiff breast cancer cells with relatively low stiffness uptook more GQD than healthy breast cells.Softening cells by pharmacologically inhibiting actomyosin activity using either siRNA or actomyosin inhibitors significantly enhanced the cellular uptake of GQD in breast cancer cells but not CSCs,while stiffening cells by activating actomyosin using CA-MLCK/ROCK or actomyosin activators considerably suppressed the nanoparticle uptake in both cancer cells and CSCs.GQD could specifically target CSC because of low cell stiffness of CSC in breast cancer cell line MCF-7 and MDA-MB-231.Further regulating cell stiffness reflected that decreasing breast cancer cell stiffness by inhibiting actomyosin activity using blebbistatin could promote GQD uptake.Vice versa,stiffening cancer cell by activating actomyosin decreased GQD uptake.The attachment of anti-cancer drug doxorubicin did not alter the trend of GQD uptake in neither soften nor stiffen cancer cells.Actomyosin activity regulates cellular uptake ofGQD might through clathrin and caveolin-mediated endocytosis.Cancer cells are softer than normal cells from the same organ,CSC are softer than non-CSC.Thus we further confirmed that the GQD uptake of normal breast cell line MCF-10 is less than breast cancer cell line MCF-7 and MDA-MB-231.Suggesting the clinical significance that using GQD as drug carrier targeting softer cells could reduce side effects to normal tissue cells.Since CSC are softer than non-CSC,GQD could decreased the percentage of CSC in whole cancer cell population by targeting softer cells.These results suggesting that it would be possible to target cancer cells and CSC by targeting from a perspective of cell mechanical difference.High uptake of nanoparticles in soft cancer cells could not be explained by their differential membrane potentials.Mechanistically,low cell mechanics or inhibiting actomyosin activity activated both clathrin and caveolin-mediated endocytosis signaling pathways,while high cell mechanics or activating actomyosin suppressed these signalings.Pharmacologically inhibiting clathrin or caveolin-mediated endocytosis signaling significantly decreased GQD uptake in CSCs and in conventional breast cancer cells when actomyosin was suppressed.Further,GQD conjugated with doxorubicin could be specifically delivered into CSCs with low stiffness and eliminated more CSCs in the presence of both CSCs and non-CSCs.Taken together,these data reveal the regulatory role of cell mechanics in cellular uptake of nanoparticles and demonstrate that GQD can be utilized to specifically eliminate CSCs,which have important implications in nanoparticle-based drug delivery for cancer therapy.展开更多
OBJECTIVE To establish an in vitro cell model based on patient-specific human neural stem cells to study the pathomechanism of sporadic AD as well as screen candidate drugs.METHODS The peripheral blood cells from spor...OBJECTIVE To establish an in vitro cell model based on patient-specific human neural stem cells to study the pathomechanism of sporadic AD as well as screen candidate drugs.METHODS The peripheral blood cells from sporadic AD patients and cognitive normal controls were repro.grammed into inducedpluripotent stem cells(iPSCs),which were further induced into neural stem cells and neurons.The cell growth curve during the differentiation process was recorded by the IncuCyte ZOOM,and neural stem cells and neurons were identified by immunofluorescence.The apoptosis of neural stem cells and neurons was detected by Click-iT~Plus TUNEL Assay.RESULTS Neural stem cells derived from AD patients and cognitive normal controls can express neural stem cell markers Nes.tin,Sox1,Sox2 and Ki67.TUNEL assay results showed that the number of TUNEL-positive cells in neu.ral stem cells derived from AD patients was significantly higher than that of cognitive normal controls(P<0.01).When neural stem cells were differentiated into neurons,the percentage of MAP2 positive cells in the neural stem cell-derived culture dish of AD patients was significantly higher than the cogni.tive normal controls at day 16 of neuronal differentiation(P<0.01);the TUNEL assay showed that the number of TUNEL-positive cells in AD-derived neurons was significantly greater than that in cognitive normal controls(P<0.01) at day 16 of neuronal differentiation.CONCLUSION Our study revealed that AD-iPSC-derived neural stem cells exhibit premature neuronal differentiation and increased neural apoptosis,which might be relevant to the neuronal loss of AD,thus may provide valuable new tools to screen candidate drugs for the disease and to discover the mechanisms underlying AD pathogenesis.展开更多
OBJECTIVE To establish an in vitro cel model based on patient-specific human microglia to study the pathological mechanism of Alzheimer disease(AD) and to screen candidate drugs.METHODS First,the induced pluripotent s...OBJECTIVE To establish an in vitro cel model based on patient-specific human microglia to study the pathological mechanism of Alzheimer disease(AD) and to screen candidate drugs.METHODS First,the induced pluripotent stem cells(iPSCs) of AD patients and cognitive normal controls(CNC) were induced to hematopoietic progenitor cells(HPCs),and then HPCs were further induced with IL-34,M-CSF,GM-CSF and TGF-β1 for 20 d to obtain microglialike cells(MGLCs).HPCs were isolated by flow cytometry and MGLCs were identified by immunofluorescence.Cell phagocytosis was determined by phagocytosis neutral red experiusing Luminex assay kits,and the cell growth curve during the experiment was recorded by IncuCyte ZOOM.The phagocytic ability and secretion of cytokines of MGLCs were observed under the stimulation of LPS.RESULTS MGLCs from AD patients(AD-MGLCs) and CNC expressed microglia markers IBA1,TMEM119,P2 RY12,TREM2 and CD11 B.The results of phagocytosis neutral red experiment showed that under normal conditions,AD-MGLCs had stronger phagocytic ability(P<0.01).Stimulation by LPS resulted in increased phagocytosis of cel s,and the increase in phagocytosis of CNC-MGLCs was higher than AD-MGLCs(P<0.01).Experiments showed that high concentrations of LPS(>2 mg·L^(-1)) resulted in CNC-microglia death(P<0.01),whereas ADMGLCs did not show significant death.The cytokine assay showed that under normal conditions,the concentrations of IFN-γ and IL-2 secreted by AD patients were slightly higher than those of CNC.After LPS stimulation,the secretion of TNF-α,IL-6 and IL-10 was significantly increased.The increased secretion of AD-MGLCs was greater than that CNC-MGLCs(P<0.01).CONCLUSION AD-iPSCs derived MGLCs exhibit significant inflammatory characteristics and are more active than CNC,which may be associated with chronic inflammatory responses caused by microglia in AD,thus may provide valuable new tools to screen candidate drugs for the disease and to discover the mechanisms underlying AD pathogenesis.展开更多
Objective Diabetic patients pose a greater challenge in managing chronic wound healing,leading to a higher amputation risk compared to non-diabetic patients.Due to their paracrine function by secreting various cytokin...Objective Diabetic patients pose a greater challenge in managing chronic wound healing,leading to a higher amputation risk compared to non-diabetic patients.Due to their paracrine function by secreting various cytokines and angiogenic factors,mesenchymal stem cells(MSCs)have been acknowledged to be a potential agent in modulating wound healing process.However,post-transplanted MSCs are vulnerable to death,indicating poor survival and migration ability in the wound site of the host,especially under hyperglycemia.As hyperglycemia induces reactive oxygen species(ROS)generation and cellular apoptosis,improvement of MSCs survival and migration potentials under hyperglycemia could contribute to a more efficient MSCs-based wound healing therapy.Salidroside(Sa),a small-molecule drug derived from Rhodiola plant,has been proved to enhance the paracrine function of skeletal muscle cells,as well as their migration even under hypoxichyperglycemia.Herein,we investigated whether Sa could improve the survival and migration potentials of MSCs,subsequently enhance the wound healing process under hyperglycemia.Methods MSCs were cultured under three conditions:low glucose,high glucose,and high glucose+Sa.qPCR analysis and western blotting were done to examine the mRNA and protein expression level of several factors which are important in upregulating the wound healing process.MTT colorimetric assay,intracellular ROS detection,and flow cytometry assay were employed to examine the effect of Sa in MSCs survival.Transwell chamber assay,scratch assay,and phalloidin staining were done to elucidate the role of Sa in regulating MSCs migration potential.For in vivo experiment,diabetic wound healing mice model was generated to elucidate the effect of Sa-pretreated MSCs transplantation in wound closure rate,as well as re-epithelization status,observed with hematoxylin and eosin staining.The diabetic wound healing mice model were divided into three groups:1)mice injected with PBS,2)mice transplanted with PBS-pretreated MSCs,and 3)mice transplanted with Sa-pretreated MSCs.Results(1)Hyperglycemic condition induced the generation of ROS and suppressed total cell number of MSCs,while Sa treatment into MSCs restored these hyperglycemia-induced alterations.In line with this,total apoptotic cells were also suppressed by treating MSCs with Sa.The expression level of cell survival factor,heme-oxygenase 1(HO-1),was enhanced in Sa-pretreated MSCs.Further treatment of HO-1 inhibitor into Sa-pretreated MSCs nullified the ROS level and total apoptotic cells,indica-ting the importance of HO-1 in mediating the Sa-induced survival of MSCs under hyperglycemia.(2)Transwell chamber and scratch assay results showed that Sa-pretreated MSCs have a higher migration potential under hyperglycemia,supported by higher F-actin polymerization fractal dimension.Fibroblast growth factor 2(FGF2)and hepatocyte growth factor(HGF)expression level,which are essential factors for cell migration,were also improved in Sa-pretreated MSCs under hyperglycemia.(3)In diabetic wound healing mice model,transplantation of Sa-pretreated MSCs resulted in significantly improved wound closure rate and re-epithelization.The protein levels of HO-1,FGF2,and HGF were also enhanced in the tissues obtained from the wound site of diabetic wound healing mice model which were transplanted with Sa-pretreated MSCs.Conclusions Salidroside pretreatment on MSCs could improve their survival and migration potentials,subsequently promoting wound healing process under hyperglycemia.This prospective MSC-based therapy could serve as a novel strategy to improve diabetic wound healing.展开更多
Objective:To investigate the influence of electroacupuncture(EA) combined with repetitive transcranial magnetic stimulation(rTMS) on the temporal profile of nestin expression after induction of focal cerebral ischemia...Objective:To investigate the influence of electroacupuncture(EA) combined with repetitive transcranial magnetic stimulation(rTMS) on the temporal profile of nestin expression after induction of focal cerebral ischemia in adult rats and to explore the mechanism of EA combined with rTMS in treating ischemic brain injury.Method:The model of transient focal ischemia was produced by occlusion of middle cerebral artery.Seventy-five Wistar rats were randomly divided into normal group,model group,EA group,rTMS group and EA +rTMS group.The neurologic impairment rating and ability of learning and memory were observed at the 7th、14th and 28th d after infarction respectively.Meanwhile,Western blotting was used to observe the number of nestin expression positive cells.Result:Nestin-positive cells were found in cortex,subgranular zone(SGZ),subventricular zone(SVZ) of the ipsilateral side at different time points after cerebral ischemia.The number of nestin-positive cells peaked at the 7th d,began to decrease at the 14th d and was significantly higher in EA+rTMS group than that in model group(P< 0.05),then almost reached normal at the 28th d.The improvement of neural motor function deficits as well as the indexes of learning and memory were more obvious in EA+rTMS group compared with model group(P<0.01,P<0.05).These effects were most obvious in EA +rTMS group compared with the EA and rTMS group(P<0.05).Conclusion:EA and rTMS possess the potency of building up and can increase the number of nestin-positive cells in some brain regions after focal cerebral ischemia,which might be one of the important mechanisms of EA combined with rTMS in treating ischemia brain injury.展开更多
Objective:To investigate the effect of SHU555A,a clinically approved iron nanoparticle,labeling on differentiation of bone marrow mesenchymal stem cells(BMSCs) into neurocyte-like cells in vitro.Methods:10 times dilut...Objective:To investigate the effect of SHU555A,a clinically approved iron nanoparticle,labeling on differentiation of bone marrow mesenchymal stem cells(BMSCs) into neurocyte-like cells in vitro.Methods:10 times dilution of 10μl,20μl,40μl and 80μl SHU555A were added to 2ml of culture medium containing rat BMSCs to obtain four experimental groups of SHU555A labeling of BMSCs with ferri ion concentrations of 14μg/ml,28μg/ml,56μg/ml and 112μg/ml,respectively.2ml of culture medium with rat BMSCs did not contain SHU555A served as control group.The BMSCs of all the groups were pre-induced by bFGF,and induced by DMSO/butylated hydroxyanisole(BHA) for six hours,subsequently reverse transcription polymerase chain reaction(RT-PCR) technique was employed to detect mRNA expression of nestin,neuronspecific analase(NSE) and glial fibrillary acid protein(GFAP).Western blot technique was used to detectprotein expression of nestin.Results:Quantitative-PCR revealed high mRNA expression of nestin,NSE and GFAP induced by DMSO/BHA in all the experimental groups,but the difference between the experimental groups and the control group was not significant(P>0.05).Western blot analysis demonstrated there was no statistically significant difference in nestin protein expression between the experimental groups and the control group(P>0.05).Conclusion:SHU555A labeling do not affect differentiation of rat BMSCs into neurocyte-like cells in vitro.展开更多
OBJECTIVE To investigate the inhibitory effect of scutellarin on the self-renewal and differentiation of HT-29 cells-derived cancer stem-like cells(HT-29CSC)in vitro and in vivo,and to explore its mechanism.METHODS Th...OBJECTIVE To investigate the inhibitory effect of scutellarin on the self-renewal and differentiation of HT-29 cells-derived cancer stem-like cells(HT-29CSC)in vitro and in vivo,and to explore its mechanism.METHODS The effect of scutellarin on the growth of HT-29CSC was determined by 3D Culture assay.The effect of scutellarin on growth and transformation of HT-29CSC was probed by soft agar colony formation assay.The effect of scutellarin on the differentiation of HT-29CSC was determined by serum induction differentiation assay in vitro.The effects of scutellarin on the expressions of marker gene Lgr5,target gene c-Myc,proliferation gene CK20 and Nanog gene were measured by quantitative real-time RT-PCR.Investigate the effect of scutellarin on the expression of c-Myc,Gli1,and Lgr5 protein by Western blotting.A subcutaneous xenograft model of colon cancer in nude mice was established and administered by intraperitoneal injection.The change of body weight and tumor size of nude mice were observed every two days.Investi⁃gate the effects of scutellarin on the growth of xenograft tumors in nude mice.The expression of CD133,Lgr5,Gli1,Ptch1,c-Myc,Ki67,CK20,Nanog gene in tumors were measured by quantitative real-time RT-PCR.The expression of c-Myc,Gli1,Lgr5,CD133,Ki67 protein were measured by Western blotting.RESULTS Scutellarin can inhibit the growth of HT-29CSC in 3D culture.Compared with the solvent control group,scutellarin can significantly inhibit the growth and transformation and differentiation of HT-29CSC in vitro(P<0.01).The expression levels of marker genes Lgr5,target gene c-Myc,proliferation gene CK20 and Nanog in HT-29CSC were down-regulated by scutellarin.Scutellarin can reduce the expression of c-Myc,Gli1,and Lgr5 protein in HT-29CSC.Scutellarin can inhibit the growth of colon cancer xenografts,lower CD133,Lgr5,Gli1,Ptch1,c-Myc,Ki67,CK20,and Nanog mRNA level of xenograft tumors,reduce the expression of c-Myc,Gli1,Lgr5,CD133,and Ki67 protein of xenograft tumors in nude mice.CONCLUSION Scutellarin,which is the main component of scutellaria barbata,can inhibit the differentiation of HT-29CSC and the mechanism is to inhibit the activity of Hedgehog signaling pathway.展开更多
Non-alcoholic fatty liver disease(NAFLD)is the most common chronic liver disease,defined by several phases,ranging from benign fat accumulation to non-alcoholic steatohepatitis(NASH),which can lead to liver cancer and...Non-alcoholic fatty liver disease(NAFLD)is the most common chronic liver disease,defined by several phases,ranging from benign fat accumulation to non-alcoholic steatohepatitis(NASH),which can lead to liver cancer and cirrhosis.Although NAFLD is a disease of disordered metabolism,it also involves several immune cell-mediated inflammatory processes,either promoting and/or suppressing hepatocyte inflammation through the secretion of pro-inflammatory and/or anti-inflammatory factors to influence the NAFLD process.However,the underlying disease mechanism and the role of immune cells in NAFLD are still under investigation,leaving many open-ended questions.In this review,we presented the recent concepts about the interplay of immune cells in the onset and pathogenesis of NAFLD.We also highlighted the specific non-immune cells exhibiting immunological properties of therapeutic significance in NAFLD.We hope that this review will help guide the development of future NAFLD therapeutics.展开更多
OBJECTIVE The eradication of cancer stem cells(CSCs) is signifcant for cancer therapy and prevention.METHODS In this study,we evaluated WM130,a novel derivative of matrine,for its effect on CSCs using human hepatocell...OBJECTIVE The eradication of cancer stem cells(CSCs) is signifcant for cancer therapy and prevention.METHODS In this study,we evaluated WM130,a novel derivative of matrine,for its effect on CSCs using human hepatocellular carcinoma(HCC) cell lines,their sphere cells,and sorted EpCAM+cells.RESULTS We revealed that WM130 could not only inhibit proliferation and colony formation of HCC cells,but also suppress the expression of some stemness-related genes and up-regulate some mature hepatocyte marker genes,indicating a promotion of differentiation from CSCs to hepatocytes.WM130 also suppressed the proliferation of doxorubicin-resistant hepatoma cells,and markedly reduced the cells with CSC biomarker EpCAM.Moreover,WM130 suppressed HCC spheres,not only primary spheres but also subsequent spheres,indicating an inhibitory effect on self-renewal capability of CSCs.Interestingly,WM130 exhibiteda remarkable inhibitory preference on HCC spheres and EpCAM+cells rather than their parental HCC cells and EpCAM-cells respectively.In vivo,WM130 inhibited HCC xenograft growth,decreased the number of sphere-forming cells,and remarkably decreased the levels of EpCAM mRNA and protein in tumor xenografts.Better inhibitory effect was achieved by WM130 in combination with doxorubicin.Further mechanism study revealed that WM130 inhibited AKT/GSK3β/β-catenin signaling pathway.CONCLUSION Collectively,our results suggest that WM130 remark.ably inhibits hepatic CSCs,and this effect may via the down-regulation of the AKT/GSK3β/β-catenin pathway.These findings provide a strong rationale for the use of WM130 as a novel drug candidate in HCC therapy.展开更多
基金supported by National Natural Science Foundation of China(81473549)Fundamental Research Funds for Central Universities(XDJK2017E158)
文摘Since the diccovery of neural stem cells(NSCs)in the embryonic and adult mammalian central nerous system,it provided novel ideas forneurogenesis as the potential of proliferation and differentiation of NSCs.One of the ways to promote the clinical application of neural stem cells(NSCs)is searching effective methods which regulate the proliferation and differentiation.This is also a problem urgently to be solved in medical field.Plenty of earlier studies have shown that traditional chinese medicine can promote the proliferation and differentiation of NSCs by regulating the related signaling pathway in vivo and in vitro.The reports of Chinese and foreign literatures on regulating the proliferation and differentiation of neural stem cells in recent ten years and their target and signaling pathways is analyzed in this review.The traditional chinese medicine regulate proliferation and differentiation of NSCs by the signaling pathways of Notch,PI3K/Akt,Wnt/β-catenin,and GFs.And,those signaling pathways have cross-talk in the regulation progress.Moreover,some traditional Chinese medicine,such as astragalus,has a variety of active ingredients to regulate proliferation and differentiation of NSCs through different signaling pathways.However,to accelerate the clinical application of neural stem cells,the studies aboutthe proliferation and differentiation of NSCs and Chinese medicine should be further deepened,the mechanism of multiple targets and the comprehensive regulation function of traditional Chinese medicine should be clarified.
基金Supported by the Scientifi c Research Foundation for Doctors of Northeast Agricultural University(2012RCB27)Open Projects of Key Laboratory of Animal Genetics,Breeding and Reproduction,College of Heilongjiang Province(GXZDSYS-2012-07)
文摘The experiment was designed to study effects of retinoic acid and ascorbic acid on differentiation of mouse embryonic stem cells to cardiomyocytes. Embryonic bodies （EB） were developed from mESC in suspension culture, different levels of concentration of retinoic acid and ascorbic acid were used to determine the optimal conditions for EB formation. The results showed that the optimal concentrations were 10.9 mol. L-1 and 0.1 mg. mL-1 for retinoic acid and ascorbic acids, respectively. 50% of EB which was significantly （p〈0.05） different from the control group developed to cardiomyocytes. In conclusion, rctinoic acid and ascorbic acid had strong ability to promote cardiomyocyte differentiation of mouse embryonic stem cells. 10-9 mol. L-1 retinoic acid and 0.10 mg. mL-1 ascorbic acids were recommended to induce differentiation of mouse ES ceUs toward cardiomyocytes.
基金Natural Science Foundation of Hebei Province(H2022106065)Scientific Research Program of Hebei Provincial Administration of Traditional Chinese Medicine(2023172)。
文摘OBJECTIVE To explore the key mechanism of Bazi Bushen capsule(BZBS)in delaying the senescence of mesenchymal stem cells(MSCs)through network pharmacology and in vitro experiments.METHODS Network pharmacology was used to predict the mechanism targets of BZBS in delaying MSCs senescence.A MSCs senescence model induced by D-galactose(D-gal)was used to investigate the effect and mechanism of BZBS on MSCs senescence in vitro.RESULTS Network pharmacology analy⁃sis showed that BZSB could delay MSCs senes⁃cence.The experiment showed that BZBS could significantly improve the survival activity of the aged MSCs.It significantly reduced the positive rate ofβ-galactosidase staining and p16,p21 expression in aged MSCs,enhanced the ability of adipogenic differentiation and osteogenic differentiation,and increased expression of Nanog,OCT4 and SOX2 in senescent MSCs.CONCLU⁃SIONS Network pharmacology and in vitro cell experiments verified that BZBS could delay MSCs senescence.
文摘Embryonic stem (ES) cell biology is attracting much attention in cell biology because of their pluripotent behaviors and potential therapeutic applications. However,what maintains ES cell pluripotency and what triggers ES cell
文摘Objective Bone-marrow stem-cell transplantation has been shown to improve cardiac function in patients with AMI, but the safety of intracoronory infusion of autologous peripheral blood stem-cell(PBSCs) in patients with AMI is unknown. For this reason, we observe the feasibility and safety of PBSCs transplantation by intracoronory infusion in such patients.Method Fourty one patients with AMI were allocated to receive Granulocyte Colony-Stimulating Factor (G-CSF:Filgrastim,300 μg) with the dose of 300 μg-600 μg/day to mobilize the stem cell, and the duration of applying G-CSF was 5 days . On the sixth day, PBSCs were separated by Baxter CS 3000 blood cell separator into suspend liquid 57 ml. Then the suspend liquid was infused into the infarct related artery (IRA)by occluding the over the wire balloon and infusing artery through balloon center lumen. In the process of the intracoronary infusion of PBSCs, the complications should be observed, which were arrhythmias including of bradycardia, sinus arrest or atrial ventricular block, premature ventricular beats ,ventricular tachycardia, ventricular fibrillation; and hypotention, etc. Results There were total 10 cases with complications during the intracoronary infusion of PBSCs. The incidence of complications was 24.4%(10/41), including bradycardia is 2.4 %(1/41), sinus arrest or atrial ventricular block is 4.9%(2/41), ventricular fibrillation is 2.4 %( 1/41), hypotention is14.6 % (6 /41).Conclusions In patients with AMI, intracoronary infusion of PBSCs is feasible and safe.
文摘Cancer stem cells(CSCs)are the driving force for sustainable tumor growth and metastasis and responsible for drug resistance and cancer relapse.Nanoparticle-based drug delivery has been demonstrated to be effective in combating tumor growth.However,it has been challenging to selectively eliminate CSCs due to the lack of a general signature for a spectrum of cancers.It is known that CSCs from various types of cancer show lower stiffness compared to non-CSCs.It remains unclear whether low stiffness in CSCs influences cellular uptake in nanoparticle-based drug delivery and thus the chemotherapy efficacy.Graphene quantum dot(GQD)is emerging as a promising carrier material in delivering anti-cancer drugs.We found that breast CSCs were softer than conventional cancer cells,which were further softer compared to healthy breast tissue cells.Importantly,soft CSCs uptook more GQD than conventional cancer cells,while stiff breast cancer cells with relatively low stiffness uptook more GQD than healthy breast cells.Softening cells by pharmacologically inhibiting actomyosin activity using either siRNA or actomyosin inhibitors significantly enhanced the cellular uptake of GQD in breast cancer cells but not CSCs,while stiffening cells by activating actomyosin using CA-MLCK/ROCK or actomyosin activators considerably suppressed the nanoparticle uptake in both cancer cells and CSCs.GQD could specifically target CSC because of low cell stiffness of CSC in breast cancer cell line MCF-7 and MDA-MB-231.Further regulating cell stiffness reflected that decreasing breast cancer cell stiffness by inhibiting actomyosin activity using blebbistatin could promote GQD uptake.Vice versa,stiffening cancer cell by activating actomyosin decreased GQD uptake.The attachment of anti-cancer drug doxorubicin did not alter the trend of GQD uptake in neither soften nor stiffen cancer cells.Actomyosin activity regulates cellular uptake ofGQD might through clathrin and caveolin-mediated endocytosis.Cancer cells are softer than normal cells from the same organ,CSC are softer than non-CSC.Thus we further confirmed that the GQD uptake of normal breast cell line MCF-10 is less than breast cancer cell line MCF-7 and MDA-MB-231.Suggesting the clinical significance that using GQD as drug carrier targeting softer cells could reduce side effects to normal tissue cells.Since CSC are softer than non-CSC,GQD could decreased the percentage of CSC in whole cancer cell population by targeting softer cells.These results suggesting that it would be possible to target cancer cells and CSC by targeting from a perspective of cell mechanical difference.High uptake of nanoparticles in soft cancer cells could not be explained by their differential membrane potentials.Mechanistically,low cell mechanics or inhibiting actomyosin activity activated both clathrin and caveolin-mediated endocytosis signaling pathways,while high cell mechanics or activating actomyosin suppressed these signalings.Pharmacologically inhibiting clathrin or caveolin-mediated endocytosis signaling significantly decreased GQD uptake in CSCs and in conventional breast cancer cells when actomyosin was suppressed.Further,GQD conjugated with doxorubicin could be specifically delivered into CSCs with low stiffness and eliminated more CSCs in the presence of both CSCs and non-CSCs.Taken together,these data reveal the regulatory role of cell mechanics in cellular uptake of nanoparticles and demonstrate that GQD can be utilized to specifically eliminate CSCs,which have important implications in nanoparticle-based drug delivery for cancer therapy.
基金supported by National key research and development program(2016YFC1306300)
文摘OBJECTIVE To establish an in vitro cell model based on patient-specific human neural stem cells to study the pathomechanism of sporadic AD as well as screen candidate drugs.METHODS The peripheral blood cells from sporadic AD patients and cognitive normal controls were repro.grammed into inducedpluripotent stem cells(iPSCs),which were further induced into neural stem cells and neurons.The cell growth curve during the differentiation process was recorded by the IncuCyte ZOOM,and neural stem cells and neurons were identified by immunofluorescence.The apoptosis of neural stem cells and neurons was detected by Click-iT~Plus TUNEL Assay.RESULTS Neural stem cells derived from AD patients and cognitive normal controls can express neural stem cell markers Nes.tin,Sox1,Sox2 and Ki67.TUNEL assay results showed that the number of TUNEL-positive cells in neu.ral stem cells derived from AD patients was significantly higher than that of cognitive normal controls(P<0.01).When neural stem cells were differentiated into neurons,the percentage of MAP2 positive cells in the neural stem cell-derived culture dish of AD patients was significantly higher than the cogni.tive normal controls at day 16 of neuronal differentiation(P<0.01);the TUNEL assay showed that the number of TUNEL-positive cells in AD-derived neurons was significantly greater than that in cognitive normal controls(P<0.01) at day 16 of neuronal differentiation.CONCLUSION Our study revealed that AD-iPSC-derived neural stem cells exhibit premature neuronal differentiation and increased neural apoptosis,which might be relevant to the neuronal loss of AD,thus may provide valuable new tools to screen candidate drugs for the disease and to discover the mechanisms underlying AD pathogenesis.
基金National Key Research and Development Program(2016YFC1306301).
文摘OBJECTIVE To establish an in vitro cel model based on patient-specific human microglia to study the pathological mechanism of Alzheimer disease(AD) and to screen candidate drugs.METHODS First,the induced pluripotent stem cells(iPSCs) of AD patients and cognitive normal controls(CNC) were induced to hematopoietic progenitor cells(HPCs),and then HPCs were further induced with IL-34,M-CSF,GM-CSF and TGF-β1 for 20 d to obtain microglialike cells(MGLCs).HPCs were isolated by flow cytometry and MGLCs were identified by immunofluorescence.Cell phagocytosis was determined by phagocytosis neutral red experiusing Luminex assay kits,and the cell growth curve during the experiment was recorded by IncuCyte ZOOM.The phagocytic ability and secretion of cytokines of MGLCs were observed under the stimulation of LPS.RESULTS MGLCs from AD patients(AD-MGLCs) and CNC expressed microglia markers IBA1,TMEM119,P2 RY12,TREM2 and CD11 B.The results of phagocytosis neutral red experiment showed that under normal conditions,AD-MGLCs had stronger phagocytic ability(P<0.01).Stimulation by LPS resulted in increased phagocytosis of cel s,and the increase in phagocytosis of CNC-MGLCs was higher than AD-MGLCs(P<0.01).Experiments showed that high concentrations of LPS(>2 mg·L^(-1)) resulted in CNC-microglia death(P<0.01),whereas ADMGLCs did not show significant death.The cytokine assay showed that under normal conditions,the concentrations of IFN-γ and IL-2 secreted by AD patients were slightly higher than those of CNC.After LPS stimulation,the secretion of TNF-α,IL-6 and IL-10 was significantly increased.The increased secretion of AD-MGLCs was greater than that CNC-MGLCs(P<0.01).CONCLUSION AD-iPSCs derived MGLCs exhibit significant inflammatory characteristics and are more active than CNC,which may be associated with chronic inflammatory responses caused by microglia in AD,thus may provide valuable new tools to screen candidate drugs for the disease and to discover the mechanisms underlying AD pathogenesis.
基金Supported by grants from the National Natural Science Foundation of China ( 81372202,81872273, 31871367)
文摘Objective Diabetic patients pose a greater challenge in managing chronic wound healing,leading to a higher amputation risk compared to non-diabetic patients.Due to their paracrine function by secreting various cytokines and angiogenic factors,mesenchymal stem cells(MSCs)have been acknowledged to be a potential agent in modulating wound healing process.However,post-transplanted MSCs are vulnerable to death,indicating poor survival and migration ability in the wound site of the host,especially under hyperglycemia.As hyperglycemia induces reactive oxygen species(ROS)generation and cellular apoptosis,improvement of MSCs survival and migration potentials under hyperglycemia could contribute to a more efficient MSCs-based wound healing therapy.Salidroside(Sa),a small-molecule drug derived from Rhodiola plant,has been proved to enhance the paracrine function of skeletal muscle cells,as well as their migration even under hypoxichyperglycemia.Herein,we investigated whether Sa could improve the survival and migration potentials of MSCs,subsequently enhance the wound healing process under hyperglycemia.Methods MSCs were cultured under three conditions:low glucose,high glucose,and high glucose+Sa.qPCR analysis and western blotting were done to examine the mRNA and protein expression level of several factors which are important in upregulating the wound healing process.MTT colorimetric assay,intracellular ROS detection,and flow cytometry assay were employed to examine the effect of Sa in MSCs survival.Transwell chamber assay,scratch assay,and phalloidin staining were done to elucidate the role of Sa in regulating MSCs migration potential.For in vivo experiment,diabetic wound healing mice model was generated to elucidate the effect of Sa-pretreated MSCs transplantation in wound closure rate,as well as re-epithelization status,observed with hematoxylin and eosin staining.The diabetic wound healing mice model were divided into three groups:1)mice injected with PBS,2)mice transplanted with PBS-pretreated MSCs,and 3)mice transplanted with Sa-pretreated MSCs.Results(1)Hyperglycemic condition induced the generation of ROS and suppressed total cell number of MSCs,while Sa treatment into MSCs restored these hyperglycemia-induced alterations.In line with this,total apoptotic cells were also suppressed by treating MSCs with Sa.The expression level of cell survival factor,heme-oxygenase 1(HO-1),was enhanced in Sa-pretreated MSCs.Further treatment of HO-1 inhibitor into Sa-pretreated MSCs nullified the ROS level and total apoptotic cells,indica-ting the importance of HO-1 in mediating the Sa-induced survival of MSCs under hyperglycemia.(2)Transwell chamber and scratch assay results showed that Sa-pretreated MSCs have a higher migration potential under hyperglycemia,supported by higher F-actin polymerization fractal dimension.Fibroblast growth factor 2(FGF2)and hepatocyte growth factor(HGF)expression level,which are essential factors for cell migration,were also improved in Sa-pretreated MSCs under hyperglycemia.(3)In diabetic wound healing mice model,transplantation of Sa-pretreated MSCs resulted in significantly improved wound closure rate and re-epithelization.The protein levels of HO-1,FGF2,and HGF were also enhanced in the tissues obtained from the wound site of diabetic wound healing mice model which were transplanted with Sa-pretreated MSCs.Conclusions Salidroside pretreatment on MSCs could improve their survival and migration potentials,subsequently promoting wound healing process under hyperglycemia.This prospective MSC-based therapy could serve as a novel strategy to improve diabetic wound healing.
基金Supported by the National Nature Science Foundation of China(No.30672216)Project of Wuhan Hygiene Bureau(No.WX08A01,No.WZ08B02)
文摘Objective:To investigate the influence of electroacupuncture(EA) combined with repetitive transcranial magnetic stimulation(rTMS) on the temporal profile of nestin expression after induction of focal cerebral ischemia in adult rats and to explore the mechanism of EA combined with rTMS in treating ischemic brain injury.Method:The model of transient focal ischemia was produced by occlusion of middle cerebral artery.Seventy-five Wistar rats were randomly divided into normal group,model group,EA group,rTMS group and EA +rTMS group.The neurologic impairment rating and ability of learning and memory were observed at the 7th、14th and 28th d after infarction respectively.Meanwhile,Western blotting was used to observe the number of nestin expression positive cells.Result:Nestin-positive cells were found in cortex,subgranular zone(SGZ),subventricular zone(SVZ) of the ipsilateral side at different time points after cerebral ischemia.The number of nestin-positive cells peaked at the 7th d,began to decrease at the 14th d and was significantly higher in EA+rTMS group than that in model group(P< 0.05),then almost reached normal at the 28th d.The improvement of neural motor function deficits as well as the indexes of learning and memory were more obvious in EA+rTMS group compared with model group(P<0.01,P<0.05).These effects were most obvious in EA +rTMS group compared with the EA and rTMS group(P<0.05).Conclusion:EA and rTMS possess the potency of building up and can increase the number of nestin-positive cells in some brain regions after focal cerebral ischemia,which might be one of the important mechanisms of EA combined with rTMS in treating ischemia brain injury.
基金Henan ontstanding talent program(084200510012)zhou research programs(083SGYS33262-5)zhou university 2011 project,third constraction projection:basic and clinical research of stem cell
文摘Objective:To investigate the effect of SHU555A,a clinically approved iron nanoparticle,labeling on differentiation of bone marrow mesenchymal stem cells(BMSCs) into neurocyte-like cells in vitro.Methods:10 times dilution of 10μl,20μl,40μl and 80μl SHU555A were added to 2ml of culture medium containing rat BMSCs to obtain four experimental groups of SHU555A labeling of BMSCs with ferri ion concentrations of 14μg/ml,28μg/ml,56μg/ml and 112μg/ml,respectively.2ml of culture medium with rat BMSCs did not contain SHU555A served as control group.The BMSCs of all the groups were pre-induced by bFGF,and induced by DMSO/butylated hydroxyanisole(BHA) for six hours,subsequently reverse transcription polymerase chain reaction(RT-PCR) technique was employed to detect mRNA expression of nestin,neuronspecific analase(NSE) and glial fibrillary acid protein(GFAP).Western blot technique was used to detectprotein expression of nestin.Results:Quantitative-PCR revealed high mRNA expression of nestin,NSE and GFAP induced by DMSO/BHA in all the experimental groups,but the difference between the experimental groups and the control group was not significant(P>0.05).Western blot analysis demonstrated there was no statistically significant difference in nestin protein expression between the experimental groups and the control group(P>0.05).Conclusion:SHU555A labeling do not affect differentiation of rat BMSCs into neurocyte-like cells in vitro.
基金National Natural Science Foundation of China(8157381381173598)+1 种基金Excellent Talent Program of Chengdu University of Traditional Chinese Medicine(YXRC2019002)Fund of Scientific Research Innovation Team Construction in Sichuan Provincial University(18TD0017)
文摘OBJECTIVE To investigate the inhibitory effect of scutellarin on the self-renewal and differentiation of HT-29 cells-derived cancer stem-like cells(HT-29CSC)in vitro and in vivo,and to explore its mechanism.METHODS The effect of scutellarin on the growth of HT-29CSC was determined by 3D Culture assay.The effect of scutellarin on growth and transformation of HT-29CSC was probed by soft agar colony formation assay.The effect of scutellarin on the differentiation of HT-29CSC was determined by serum induction differentiation assay in vitro.The effects of scutellarin on the expressions of marker gene Lgr5,target gene c-Myc,proliferation gene CK20 and Nanog gene were measured by quantitative real-time RT-PCR.Investigate the effect of scutellarin on the expression of c-Myc,Gli1,and Lgr5 protein by Western blotting.A subcutaneous xenograft model of colon cancer in nude mice was established and administered by intraperitoneal injection.The change of body weight and tumor size of nude mice were observed every two days.Investi⁃gate the effects of scutellarin on the growth of xenograft tumors in nude mice.The expression of CD133,Lgr5,Gli1,Ptch1,c-Myc,Ki67,CK20,Nanog gene in tumors were measured by quantitative real-time RT-PCR.The expression of c-Myc,Gli1,Lgr5,CD133,Ki67 protein were measured by Western blotting.RESULTS Scutellarin can inhibit the growth of HT-29CSC in 3D culture.Compared with the solvent control group,scutellarin can significantly inhibit the growth and transformation and differentiation of HT-29CSC in vitro(P<0.01).The expression levels of marker genes Lgr5,target gene c-Myc,proliferation gene CK20 and Nanog in HT-29CSC were down-regulated by scutellarin.Scutellarin can reduce the expression of c-Myc,Gli1,and Lgr5 protein in HT-29CSC.Scutellarin can inhibit the growth of colon cancer xenografts,lower CD133,Lgr5,Gli1,Ptch1,c-Myc,Ki67,CK20,and Nanog mRNA level of xenograft tumors,reduce the expression of c-Myc,Gli1,Lgr5,CD133,and Ki67 protein of xenograft tumors in nude mice.CONCLUSION Scutellarin,which is the main component of scutellaria barbata,can inhibit the differentiation of HT-29CSC and the mechanism is to inhibit the activity of Hedgehog signaling pathway.
文摘Non-alcoholic fatty liver disease(NAFLD)is the most common chronic liver disease,defined by several phases,ranging from benign fat accumulation to non-alcoholic steatohepatitis(NASH),which can lead to liver cancer and cirrhosis.Although NAFLD is a disease of disordered metabolism,it also involves several immune cell-mediated inflammatory processes,either promoting and/or suppressing hepatocyte inflammation through the secretion of pro-inflammatory and/or anti-inflammatory factors to influence the NAFLD process.However,the underlying disease mechanism and the role of immune cells in NAFLD are still under investigation,leaving many open-ended questions.In this review,we presented the recent concepts about the interplay of immune cells in the onset and pathogenesis of NAFLD.We also highlighted the specific non-immune cells exhibiting immunological properties of therapeutic significance in NAFLD.We hope that this review will help guide the development of future NAFLD therapeutics.
基金This study was partially supported by a grant from the ministry of HealthL abor and Welfare of Japan+1 种基金Hum an Genom e and Regenerative Medicine Project (ChairpersonHidehiko Saito)
基金supported by National Natural Science Foundation of China(81270508)National Major Special Science and Technology Project(2012ZX09103101-043)
文摘OBJECTIVE The eradication of cancer stem cells(CSCs) is signifcant for cancer therapy and prevention.METHODS In this study,we evaluated WM130,a novel derivative of matrine,for its effect on CSCs using human hepatocellular carcinoma(HCC) cell lines,their sphere cells,and sorted EpCAM+cells.RESULTS We revealed that WM130 could not only inhibit proliferation and colony formation of HCC cells,but also suppress the expression of some stemness-related genes and up-regulate some mature hepatocyte marker genes,indicating a promotion of differentiation from CSCs to hepatocytes.WM130 also suppressed the proliferation of doxorubicin-resistant hepatoma cells,and markedly reduced the cells with CSC biomarker EpCAM.Moreover,WM130 suppressed HCC spheres,not only primary spheres but also subsequent spheres,indicating an inhibitory effect on self-renewal capability of CSCs.Interestingly,WM130 exhibiteda remarkable inhibitory preference on HCC spheres and EpCAM+cells rather than their parental HCC cells and EpCAM-cells respectively.In vivo,WM130 inhibited HCC xenograft growth,decreased the number of sphere-forming cells,and remarkably decreased the levels of EpCAM mRNA and protein in tumor xenografts.Better inhibitory effect was achieved by WM130 in combination with doxorubicin.Further mechanism study revealed that WM130 inhibited AKT/GSK3β/β-catenin signaling pathway.CONCLUSION Collectively,our results suggest that WM130 remark.ably inhibits hepatic CSCs,and this effect may via the down-regulation of the AKT/GSK3β/β-catenin pathway.These findings provide a strong rationale for the use of WM130 as a novel drug candidate in HCC therapy.
