Retinoic acid inducible gene-I(RIG-I)is a caspase recruitment domain(CARD)containing protein that acts as an intracellular RNA receptor and senses virus infection.After binding to double stranded RNA(dsRNA)or 5′-trip...Retinoic acid inducible gene-I(RIG-I)is a caspase recruitment domain(CARD)containing protein that acts as an intracellular RNA receptor and senses virus infection.After binding to double stranded RNA(dsRNA)or 5′-triphosphate single stranded RNA(ssRNA),RIG-I transforms into an open conformation,translocates onto mitochondria,and interacts with the downstream adaptor mitochondrial antiviral signaling(MAVS)to induce the production of type Ⅰ interferon and inflammatory factors via IRF3/7 and NF-κB pathways,respectively.Recently,accumulating evidence suggests that RIG-I could function in non-viral systems and participate in a series of biological events,such as inflammation and inflammation related diseases,cell proliferation,apoptosis and even senescence.Here we review recent advances in antiviral study of RIG-I as well as the functions of RIG-I in other fields.展开更多
Cytosolic retinoic acid-inducible gene I(RIG-I) is an important innate immune RNA sensor and can induce antiviral cytokines, e.g., interferon-β(IFN-β). Innate immune response to hepatitis B virus(HBV) plays a ...Cytosolic retinoic acid-inducible gene I(RIG-I) is an important innate immune RNA sensor and can induce antiviral cytokines, e.g., interferon-β(IFN-β). Innate immune response to hepatitis B virus(HBV) plays a pivotal role in viral clearance and persistence. However, knowledge of the role that RIG-I plays in HBV infection is limited. The woodchuck is a valuable model for studying HBV infection. To characterize the molecular basis of woodchuck RIG-I(w RIG-I), we analyzed the complete coding sequences(CDSs) of w RIG-I, containing 2778 base pairs that encode 925 amino acids. The deduced w RIG-I protein was 106.847 k D with a theoretical isoelectric point(p I) of 6.07, and contained three important functional structures [caspase activation and recruitment domains(CARDs), DEx D/H-box helicases, and a repressor domain(RD)]. In woodchuck fibroblastoma cell line(WH12/6), w RIG-I-targeted small interfering RNA(si RNA) down-regulated RIG-I and its downstrean effector–IFN-β transcripts under RIG-I' ligand, 5'-ppp double stranded RNA(ds RNA) stimulation. We also measured m RNA levels of w RIG-I in different tissues from healthy woodchucks and in the livers from woodchuck hepatitis virus(WHV)-infected woodchucks. The basal expression levels of w RIG-I were abundant in the kidney and liver. Importantly, w RIG-I was significantly up-regulated in acutely infected woodchuck livers, suggesting that RIG-I might be involved in WHV infection. These results may characterize RIG-I in the woodchuck model, providing a strong basis for further study on RIG-I-mediated innate immunity in HBV infection.展开更多
AIM: To investigate the role of retinoic add (RA) and retinaldehyde dehydrogenase-2 (RALDH(2)) of retina and choroid in the guinea pig lens-induced myopic eyes. METHODS: Totally 45 guinea pigs, at age of three weeks, ...AIM: To investigate the role of retinoic add (RA) and retinaldehyde dehydrogenase-2 (RALDH(2)) of retina and choroid in the guinea pig lens-induced myopic eyes. METHODS: Totally 45 guinea pigs, at age of three weeks, were randomly assigned into three groups: the normal control, the lens-induced group and the recovering group. Out of focus was induced by the -6.00D concave lens on the left eye, and lasted for 15 days. All animals underwent biometric measurement (corneal radius of curvature, refraction and axial length). Subsequently, RA content in the retina and RPE/choriod complex was detected by reversed-phase high-performance liquid chromatography. RALDH(2) protein in the retina and RPE/choriod complex was evaluated by the immunohistochemical staining and Western blotting. RESULTS: After wearing -6.00D lens for 15 days, axial length of the lens-induced eye extends and myopia was formed, with RA contents increasing in both the neural retina and RPE/choroid complex. Comparing with the lens-induced group, myopic degree significantly relieved, and its RA contents in both the neural retina and RPE/choroid complex decreased in the recovering group. In the normal control, RALDH(2) protein was expressed positively in the retinal nerve fiber layer (RNFL), inner plexiform layer (IPL) and lateral border of outer nuclear layer (ONL). Retinal RALDH(2) protein increased in the lens-induced group, and was also positive in the outer plexiform layer (OPL). In the recovering group, retinal RALDH(2) protein attenuated the expression in the OPL turns to negative. RALDH(2) protein was not expressed in the choroid of any group. CONCLUSION: RA of retina and chorid participates in the regulation of the lens-induced myopia in guinea pigs, which may be related with retinal RALDH(2) protein.展开更多
The cervical carcinoma cell line, ME180 cells were treated with tanshinone (Tan) or retinoic acid (RA) in DMSO (final concentration 0.02%, V/V) on 4 successive days. The cells treated with the same concentration of DM...The cervical carcinoma cell line, ME180 cells were treated with tanshinone (Tan) or retinoic acid (RA) in DMSO (final concentration 0.02%, V/V) on 4 successive days. The cells treated with the same concentration of DMSO alone served as control. Morphological studies with light and transmission electron microscopy showed that the cells treated with both Tan and RA became welldifferentiated. The cellular growth and proliferation were suppressed (as revealed by cell counting. [3H]-thymidine uptake and colony-forming assay). The number of nuclear organizer regions(AgNORs) in cells reduced and the distribution type returned nearly to normal type. The tumorigenicity in nude mice was reduced. The cell RNA dot hybridization showed that the expression of c-myc and c-Ha-ras mRNA was inhibited markedly. All above results showed that Tan and RA could reverse some malignant Phenotype and possessed differentiation inducing activity on ME180 cell line. No significant difference was observed between the cells treated with Tan and RA.展开更多
Previous studies have demonstrated that differentiated neural stem cells (NSCs) are more suitable for transplantation than non-differentiated NSCs. In this study, NSCs were expanded in vitro for two passages, induce...Previous studies have demonstrated that differentiated neural stem cells (NSCs) are more suitable for transplantation than non-differentiated NSCs. In this study, NSCs were expanded in vitro for two passages, induced with retinoic acid to differentiate, and harvested between 1 6 days later. They were subsequently cultured in artificial cerebrospinal fluid for an additional 3 days, dudng which their growth and morphology was monitored. NSCs induced for 4 days exhibited a peak rate of cells differentiating into neurons and robust growth. Our results indicate that the optimal time point for transplanting NSCs is following a 4-day period of induced differentiation.展开更多
BACKGROUND: Under induction of retinoic acid (RA), bone marrow stromal cells (BMSCs) can differentiate into nerve cells or neuron-like cells, which do not survive for a long time, so those are restricted to an ap...BACKGROUND: Under induction of retinoic acid (RA), bone marrow stromal cells (BMSCs) can differentiate into nerve cells or neuron-like cells, which do not survive for a long time, so those are restricted to an application. Other neurotrophic factors can also differentiate into neuronal cells through inducing BMSCs; especially, brain-derived neurotrophic factor (BDNF) can delay natural death of neurons and play a key role in survival and growth of neurons. The combination of them is beneficial for differentiation of BMSCs. OBJECTIVE: To investigate the effects of BDNF combining with RA on inducing differentiation of BMSCs to nerve cells of adult rats and compare the results between common medium group and single BDNF group. DESIGN: Randomized controlled animal study SETTING: Department of Neurology, Affiliated Hospital of Xuzhou Medical College MATERIALS: The experiment was carried out in the Clinical Neurological Laboratory of Xuzhou Medical College from September 2003 to April 2005. A total of 24 SD rats, of either gender, 2 months old, weighing 130-150 g, were provided by Experimental Animal Center of Xuzhou Medical College [certification: SYXK (su) 2002-0038]. Materials and reagents: low-glucose DMEM medium, bovine serum, BDNF, RA, trypsin, separating medium of lymphocyte, monoclonal antibody of mouse-anti-nestin, neuro-specific enolase, glial fibrillary acidic protein (GFAP) antibody, SABC kit, and diaminobenzidine (DAB) color agent. All these mentioned above were mainly provided by SIGMA Company, GIBCO Company and Boshide Company. METHODS: Bone marrow of SD rats was selected for density gradient centrifugation. BMSCs were undertaken primary culture and subculture; and then, those cells were induced respectively in various mediums in total of 3 groups, including control group (primary culture), BDNF group (20 μg/L BDNF) and BDNF+RA group (20 μg/L BDNF plus 20 μg/L RA). On the 3^rd and the 7^th days after induction, BMSCs were stained immunocytochemically with nestin (sign of nerve stem cells), neuron-specific enolase (NSE, sign of diagnosing neurons) and GFAP (diagnosing astrocyte), and evaluated cellular property. MAIN OUTCOME MEASURES : Induction and differentiation in vitro of BMSCs in 3 groups RESULTS: (1) Induction and differentiation of BMSCs: Seven days after induction, cells having 2 or more apophyses were observed. Soma shaped like angle or erose form, which were similar to neurons and glial cells having strong refraction. (2) Results of immunocytochemical detection: Three days after induction, rate of positive cells in BDNF+RA group was higher than that in BDNF group and control group [(86.15±4.58)%, (65.43±4.23)%, (4.18±1.09)%, P 〈 0.01]. Seven days after induction, rate of positive cells was lower in BDNF group and BDNF+RA group than that in both groups at 3 days after induction [(31.12±3.18)%, (29.35±2.69)%, P 〈 0.01]; however, amounts of positive cells of NSE and GFAP were higher than those at 3 days after induction (P 〈 0.01); meanwhile, the amount in BDNF+RA group was remarkably higher than that in BDNF group (P 〈 0.01). CONCLUSION: Combination of BDNF and RA can cooperate differentiation of BMSCs into neurons and astrocyte, and the effect is superior to single usage of BDNF.展开更多
As to determine the effect of post-remission therapy in prolonging survival and duration of remission after complete remission, 50 patients with APL In complete remission Induced by retinolc acid (RA) were divided int...As to determine the effect of post-remission therapy in prolonging survival and duration of remission after complete remission, 50 patients with APL In complete remission Induced by retinolc acid (RA) were divided into three groups randomly: (A) 30 cases, treated by alternate chemotherapy with RA; (B) 10 cases, with RA alone; (C) 10 cases, only with chemotherapy.The survival curves showed mat Group A had the survival time more than 1 year In 87. 4 %, more than 2 year in 80.7%. 26/30 cases were survival and still in remission, the survival curve tend to be a plateau at 16 months. In Group B more than 1 year in 45. 7%. In Group C, more than 1 year In 50%. (Keplan-Meler x2 = 8. 93 P <0. 01).This result showed that the alternate chemotherapy with RA for post- Induction remission therapy could be useful to Improve long-term survivors and to prolong the duration of remission.展开更多
Objective: To investigate the differentiation-inducing effects of all-trans-retinoic (ATRA) to HPV16 subgenesimmortalized human l:ndocervical cells (H8 cell) in vitro. Methods: HPV16 subgenes-immortalized human...Objective: To investigate the differentiation-inducing effects of all-trans-retinoic (ATRA) to HPV16 subgenesimmortalized human l:ndocervical cells (H8 cell) in vitro. Methods: HPV16 subgenes-immortalized human endocervical cells (H8 cells) were cultured in vitro. After treated with ATRA, the proliferation of immortalized human endocervical cells was measured by MTT assay; morphological changes were observed using M and TEM; cell cycle was analyzed by FCM; expression of Ki67 was tested using immunocytochemistry and the activity of telomerase was tested using PCR-ELISA. Results: ATRA could inhibit proliferation of H8 cells significantly and induce their morphodifferentiation. According to FCM, H8 cells accumulated in G1 phase and expression of Ki67 and activity of telomerase reduced significantly after treatment with ATRA. Conclusion: ATRA could induce the differentiation of H8 cell line obviously, which might be achieved by inhibiting proliferation, blocking cell cycle, and reducing activity of telomerase.展开更多
Objective: To observe the anti-virus effects of andrographolide (AD) on the retinoic acid-inducible gene-I (RIG-I)-Iike receptors (RLRs) signaling pathway when immunological cells were infected with HIN1. Meth...Objective: To observe the anti-virus effects of andrographolide (AD) on the retinoic acid-inducible gene-I (RIG-I)-Iike receptors (RLRs) signaling pathway when immunological cells were infected with HIN1. Methods: Leukomonocyte was obtained from umbilical cord blood by Ficoll density gradient centrifugation, and immunological cells were harvested after cytokines stimulation. Virus infected cell model was established by H1N1 co-cultured with normal human bronchial epithelial cell line (16HBE). The optimal concentration of AD was defined by methyl-thiazolyl-tetrazolium (MTT) assay. After the virus infected cell model was established, AD was added into the medium as a treatment intervention. After 24-h co-culture, cell supernatant was collected for interferon gamma (IFN- ~, ) and interleukin-4 (IL-4) enzyme-linked immunosorbent assay (ELISA) detection while immunological cells for real-time polymerase chain reaction (RT-PCR). Results: The optimal concentration of AD for anti-virus effect was 250 μg/mL. IL-4 and IFN-γ in the supernatant and mRNA levels in RLRs pathway increased when cells was infected by virus, RIG-I, IFN-13 promoter stimulator-1 (IPS-1), interferon regulatory factor (IRF)-7, IRF-3 and nuclear transcription factor K B (NF- K B) mRNA levels increased significantly (P〈0.05). When AD was added into co-culture medium, the levels of IL-4 and IFN-γ were lower than those in the non-interference groups and the mRNA expression levels decreased, RIG-I, IPS-1, IRF-7, IRF-3 and NF- K B decreased significantly in each group with significant statistic differences (P〈0.05). Conclusions: The RLRs mediated viral recognition provided a potential molecular target for acute viral infections and andrographolide could ameliorate H1N1 virus-induced cell mortality. And the antiviral effects might be related to its inhibition of viral-induced activation of the RLRs signaling pathway.展开更多
Recent studies have reported that induced pluripotent stem (iPS) cells from mice and humans can differentiate into primordial germ cells. However, whether iPS cells are capable of producing male germ cells is not kn...Recent studies have reported that induced pluripotent stem (iPS) cells from mice and humans can differentiate into primordial germ cells. However, whether iPS cells are capable of producing male germ cells is not known. The objective of this study was to investigate the differentiation potential of mouse iPS cells into spermatogonial stem cells and late-stage male germ cells. We used an approach that combines in vitrodifferentiation and in vivotransplantation. Embryoid bodies (EBs) were obtained from iPS cells using leukaemia inhibitor factor (LIF)-free medium. Quantitative PCR revealed a decrease in Oct4 expression and an increase in StraSand Vasa mRNA in the EBs derived from iPS cells, iPS cell-derived EBs were induced by retinoic acid to differentiate into spermatogonial stem cells (SSCs), as evidenced by their expression of VASA, as well as CDH1 and GFRal, which are markers of SSCs. Furthermore, these germ cells derived from iPS cells were transplanted into recipient testes of mice that had been pre-treated with busulfan. Notably, iPS cell-derived SSCs were able to differentiate into male germ cells ranging from spermatogonia to round spermatids, as shown by VASA and SCP3 expression. This study demonstrates that iPS cells have the potential to differentiate into late-stage male germ cells. The derivation of male germ cells from iPS cells has potential applications in the treatment of male infertility and provides a model for uncovering the molecular mechanisms underlying male germ cell development.展开更多
AIM To investigate the effect of(-)-epigallocatechin-3-gallate(EGCG) on polyinosinic-polycytidylic acid(poly I:C)-triggered intracellular innate immunity against hepatitis C virus(HCV) in hepatocytes. METHODS A cell c...AIM To investigate the effect of(-)-epigallocatechin-3-gallate(EGCG) on polyinosinic-polycytidylic acid(poly I:C)-triggered intracellular innate immunity against hepatitis C virus(HCV) in hepatocytes. METHODS A cell culture model of HCV infection was generated by infecting a hepatoma cell line, Huh7, with HCV JFH-1 strain(JFH-1-Huh7). Poly I:C with a high molecular weight and EGCG were used to stimulate the JFH-1-Huh7 cells. Real-time reverse transcription-polymerase chain reaction was used to detect the expression levels of intracellular m RNAs and of intracellular and extracellular HCV RNA. Enzyme-linked immunosorbent assay was used to evaluate the interferon(IFN)-λ1 protein level in the cell culture supernatant. Immunostaining was used to examine HCV core protein expression in Huh7 cells.RESULTS Our recent study showed that HCV replication could impair poly I:C-triggered intracellular innate immune responses in hepatocytes. In the current study, we showed that EGCG treatment significantly increased the poly I:C-induced expression of Toll-like receptor 3(TLR3), retinoic acid-inducible gene I, and IFN-λ1 in JFH-1-Huh7 cells. In addition, supplementation with EGCG increased the poly I:C-mediated antiviral activity in JFH-1-Huh7 cells at the intracellular and extracellular HCV RNA and protein levels. Further investigation of the mechanisms showed that EGCG treatment significantly enhanced the poly I:C-induced expression of IFN-regulatory factor 9 and several antiviral IFNstimulated genes, including ISG15, ISG56, myxovirus resistance A, and 2'-5'-oligoadenylate synthetase 1, which encode the key antiviral elements in the IFN signaling pathway. CONCLUSION Our observations provide experimental evidence that EGCG has the ability to enhance poly I:C-induced intracellular antiviral innate immunity against HCV replication in hepatocytes.展开更多
Objective The treatment of solid tumors by means of differentiation induction has not yet beengained such broken - through success as all-trans retinoic acid (ATRA) in acute promyelocytic leukemia (APL).To explore mor...Objective The treatment of solid tumors by means of differentiation induction has not yet beengained such broken - through success as all-trans retinoic acid (ATRA) in acute promyelocytic leukemia (APL).To explore more effective inducers for solid tumor therapy, dimethyl sulfoxide (DMSO) is considered as acandidate. Methods In the present study, DMSO was used as inducer to human esophageal cancer cell lines invitro, in compared with classical inducer ATRA, in terms of morphology, cell cycle, growth inhibition, cytokeratin4 expression, gap junction-mediated dye transfer and tumorigenecity in nude mice. Results DMSO as well asATRA induced differentiation of human esophageal carcinoma cells. However, DMSO was confirmed to be moreeffective for induced differentiation of esophageal carcinoma cells than ATRA. Conclusion The results suggestedthat DMSO might have a good prospect in the treatment of solid tumors.展开更多
文摘Retinoic acid inducible gene-I(RIG-I)is a caspase recruitment domain(CARD)containing protein that acts as an intracellular RNA receptor and senses virus infection.After binding to double stranded RNA(dsRNA)or 5′-triphosphate single stranded RNA(ssRNA),RIG-I transforms into an open conformation,translocates onto mitochondria,and interacts with the downstream adaptor mitochondrial antiviral signaling(MAVS)to induce the production of type Ⅰ interferon and inflammatory factors via IRF3/7 and NF-κB pathways,respectively.Recently,accumulating evidence suggests that RIG-I could function in non-viral systems and participate in a series of biological events,such as inflammation and inflammation related diseases,cell proliferation,apoptosis and even senescence.Here we review recent advances in antiviral study of RIG-I as well as the functions of RIG-I in other fields.
基金supported by grants from the National Science and Technology Major Project for Infectious Diseases of China(No.2012ZX10004503)the National Natural Science Foundation of China(No.81101248,No.81261120397 and No.81371828)the Deutsche Forschungs-gemeinschaft(Transregio TRR60)
文摘Cytosolic retinoic acid-inducible gene I(RIG-I) is an important innate immune RNA sensor and can induce antiviral cytokines, e.g., interferon-β(IFN-β). Innate immune response to hepatitis B virus(HBV) plays a pivotal role in viral clearance and persistence. However, knowledge of the role that RIG-I plays in HBV infection is limited. The woodchuck is a valuable model for studying HBV infection. To characterize the molecular basis of woodchuck RIG-I(w RIG-I), we analyzed the complete coding sequences(CDSs) of w RIG-I, containing 2778 base pairs that encode 925 amino acids. The deduced w RIG-I protein was 106.847 k D with a theoretical isoelectric point(p I) of 6.07, and contained three important functional structures [caspase activation and recruitment domains(CARDs), DEx D/H-box helicases, and a repressor domain(RD)]. In woodchuck fibroblastoma cell line(WH12/6), w RIG-I-targeted small interfering RNA(si RNA) down-regulated RIG-I and its downstrean effector–IFN-β transcripts under RIG-I' ligand, 5'-ppp double stranded RNA(ds RNA) stimulation. We also measured m RNA levels of w RIG-I in different tissues from healthy woodchucks and in the livers from woodchuck hepatitis virus(WHV)-infected woodchucks. The basal expression levels of w RIG-I were abundant in the kidney and liver. Importantly, w RIG-I was significantly up-regulated in acutely infected woodchuck livers, suggesting that RIG-I might be involved in WHV infection. These results may characterize RIG-I in the woodchuck model, providing a strong basis for further study on RIG-I-mediated innate immunity in HBV infection.
基金National Natural Science Foundation of China (No.81070752)
文摘AIM: To investigate the role of retinoic add (RA) and retinaldehyde dehydrogenase-2 (RALDH(2)) of retina and choroid in the guinea pig lens-induced myopic eyes. METHODS: Totally 45 guinea pigs, at age of three weeks, were randomly assigned into three groups: the normal control, the lens-induced group and the recovering group. Out of focus was induced by the -6.00D concave lens on the left eye, and lasted for 15 days. All animals underwent biometric measurement (corneal radius of curvature, refraction and axial length). Subsequently, RA content in the retina and RPE/choriod complex was detected by reversed-phase high-performance liquid chromatography. RALDH(2) protein in the retina and RPE/choriod complex was evaluated by the immunohistochemical staining and Western blotting. RESULTS: After wearing -6.00D lens for 15 days, axial length of the lens-induced eye extends and myopia was formed, with RA contents increasing in both the neural retina and RPE/choroid complex. Comparing with the lens-induced group, myopic degree significantly relieved, and its RA contents in both the neural retina and RPE/choroid complex decreased in the recovering group. In the normal control, RALDH(2) protein was expressed positively in the retinal nerve fiber layer (RNFL), inner plexiform layer (IPL) and lateral border of outer nuclear layer (ONL). Retinal RALDH(2) protein increased in the lens-induced group, and was also positive in the outer plexiform layer (OPL). In the recovering group, retinal RALDH(2) protein attenuated the expression in the OPL turns to negative. RALDH(2) protein was not expressed in the choroid of any group. CONCLUSION: RA of retina and chorid participates in the regulation of the lens-induced myopia in guinea pigs, which may be related with retinal RALDH(2) protein.
文摘The cervical carcinoma cell line, ME180 cells were treated with tanshinone (Tan) or retinoic acid (RA) in DMSO (final concentration 0.02%, V/V) on 4 successive days. The cells treated with the same concentration of DMSO alone served as control. Morphological studies with light and transmission electron microscopy showed that the cells treated with both Tan and RA became welldifferentiated. The cellular growth and proliferation were suppressed (as revealed by cell counting. [3H]-thymidine uptake and colony-forming assay). The number of nuclear organizer regions(AgNORs) in cells reduced and the distribution type returned nearly to normal type. The tumorigenicity in nude mice was reduced. The cell RNA dot hybridization showed that the expression of c-myc and c-Ha-ras mRNA was inhibited markedly. All above results showed that Tan and RA could reverse some malignant Phenotype and possessed differentiation inducing activity on ME180 cell line. No significant difference was observed between the cells treated with Tan and RA.
基金the National Natural Science Foundation of China, No. 30570628, 30770751
文摘Previous studies have demonstrated that differentiated neural stem cells (NSCs) are more suitable for transplantation than non-differentiated NSCs. In this study, NSCs were expanded in vitro for two passages, induced with retinoic acid to differentiate, and harvested between 1 6 days later. They were subsequently cultured in artificial cerebrospinal fluid for an additional 3 days, dudng which their growth and morphology was monitored. NSCs induced for 4 days exhibited a peak rate of cells differentiating into neurons and robust growth. Our results indicate that the optimal time point for transplanting NSCs is following a 4-day period of induced differentiation.
文摘BACKGROUND: Under induction of retinoic acid (RA), bone marrow stromal cells (BMSCs) can differentiate into nerve cells or neuron-like cells, which do not survive for a long time, so those are restricted to an application. Other neurotrophic factors can also differentiate into neuronal cells through inducing BMSCs; especially, brain-derived neurotrophic factor (BDNF) can delay natural death of neurons and play a key role in survival and growth of neurons. The combination of them is beneficial for differentiation of BMSCs. OBJECTIVE: To investigate the effects of BDNF combining with RA on inducing differentiation of BMSCs to nerve cells of adult rats and compare the results between common medium group and single BDNF group. DESIGN: Randomized controlled animal study SETTING: Department of Neurology, Affiliated Hospital of Xuzhou Medical College MATERIALS: The experiment was carried out in the Clinical Neurological Laboratory of Xuzhou Medical College from September 2003 to April 2005. A total of 24 SD rats, of either gender, 2 months old, weighing 130-150 g, were provided by Experimental Animal Center of Xuzhou Medical College [certification: SYXK (su) 2002-0038]. Materials and reagents: low-glucose DMEM medium, bovine serum, BDNF, RA, trypsin, separating medium of lymphocyte, monoclonal antibody of mouse-anti-nestin, neuro-specific enolase, glial fibrillary acidic protein (GFAP) antibody, SABC kit, and diaminobenzidine (DAB) color agent. All these mentioned above were mainly provided by SIGMA Company, GIBCO Company and Boshide Company. METHODS: Bone marrow of SD rats was selected for density gradient centrifugation. BMSCs were undertaken primary culture and subculture; and then, those cells were induced respectively in various mediums in total of 3 groups, including control group (primary culture), BDNF group (20 μg/L BDNF) and BDNF+RA group (20 μg/L BDNF plus 20 μg/L RA). On the 3^rd and the 7^th days after induction, BMSCs were stained immunocytochemically with nestin (sign of nerve stem cells), neuron-specific enolase (NSE, sign of diagnosing neurons) and GFAP (diagnosing astrocyte), and evaluated cellular property. MAIN OUTCOME MEASURES : Induction and differentiation in vitro of BMSCs in 3 groups RESULTS: (1) Induction and differentiation of BMSCs: Seven days after induction, cells having 2 or more apophyses were observed. Soma shaped like angle or erose form, which were similar to neurons and glial cells having strong refraction. (2) Results of immunocytochemical detection: Three days after induction, rate of positive cells in BDNF+RA group was higher than that in BDNF group and control group [(86.15±4.58)%, (65.43±4.23)%, (4.18±1.09)%, P 〈 0.01]. Seven days after induction, rate of positive cells was lower in BDNF group and BDNF+RA group than that in both groups at 3 days after induction [(31.12±3.18)%, (29.35±2.69)%, P 〈 0.01]; however, amounts of positive cells of NSE and GFAP were higher than those at 3 days after induction (P 〈 0.01); meanwhile, the amount in BDNF+RA group was remarkably higher than that in BDNF group (P 〈 0.01). CONCLUSION: Combination of BDNF and RA can cooperate differentiation of BMSCs into neurons and astrocyte, and the effect is superior to single usage of BDNF.
文摘As to determine the effect of post-remission therapy in prolonging survival and duration of remission after complete remission, 50 patients with APL In complete remission Induced by retinolc acid (RA) were divided into three groups randomly: (A) 30 cases, treated by alternate chemotherapy with RA; (B) 10 cases, with RA alone; (C) 10 cases, only with chemotherapy.The survival curves showed mat Group A had the survival time more than 1 year In 87. 4 %, more than 2 year in 80.7%. 26/30 cases were survival and still in remission, the survival curve tend to be a plateau at 16 months. In Group B more than 1 year in 45. 7%. In Group C, more than 1 year In 50%. (Keplan-Meler x2 = 8. 93 P <0. 01).This result showed that the alternate chemotherapy with RA for post- Induction remission therapy could be useful to Improve long-term survivors and to prolong the duration of remission.
基金this work was supported by a grant from the Key Project of Chongqing Science & Technology Committee. (No. 2004-27-17).
文摘Objective: To investigate the differentiation-inducing effects of all-trans-retinoic (ATRA) to HPV16 subgenesimmortalized human l:ndocervical cells (H8 cell) in vitro. Methods: HPV16 subgenes-immortalized human endocervical cells (H8 cells) were cultured in vitro. After treated with ATRA, the proliferation of immortalized human endocervical cells was measured by MTT assay; morphological changes were observed using M and TEM; cell cycle was analyzed by FCM; expression of Ki67 was tested using immunocytochemistry and the activity of telomerase was tested using PCR-ELISA. Results: ATRA could inhibit proliferation of H8 cells significantly and induce their morphodifferentiation. According to FCM, H8 cells accumulated in G1 phase and expression of Ki67 and activity of telomerase reduced significantly after treatment with ATRA. Conclusion: ATRA could induce the differentiation of H8 cell line obviously, which might be achieved by inhibiting proliferation, blocking cell cycle, and reducing activity of telomerase.
基金Supported by the National Natural Science Foundation of China(No.81273616 and 30973693)the Doctoral Fund of the Ministryof Education 2010(No.20104401110003)the Natural Science Foundation of Guangdong Province(No.S2013010013434)
文摘Objective: To observe the anti-virus effects of andrographolide (AD) on the retinoic acid-inducible gene-I (RIG-I)-Iike receptors (RLRs) signaling pathway when immunological cells were infected with HIN1. Methods: Leukomonocyte was obtained from umbilical cord blood by Ficoll density gradient centrifugation, and immunological cells were harvested after cytokines stimulation. Virus infected cell model was established by H1N1 co-cultured with normal human bronchial epithelial cell line (16HBE). The optimal concentration of AD was defined by methyl-thiazolyl-tetrazolium (MTT) assay. After the virus infected cell model was established, AD was added into the medium as a treatment intervention. After 24-h co-culture, cell supernatant was collected for interferon gamma (IFN- ~, ) and interleukin-4 (IL-4) enzyme-linked immunosorbent assay (ELISA) detection while immunological cells for real-time polymerase chain reaction (RT-PCR). Results: The optimal concentration of AD for anti-virus effect was 250 μg/mL. IL-4 and IFN-γ in the supernatant and mRNA levels in RLRs pathway increased when cells was infected by virus, RIG-I, IFN-13 promoter stimulator-1 (IPS-1), interferon regulatory factor (IRF)-7, IRF-3 and nuclear transcription factor K B (NF- K B) mRNA levels increased significantly (P〈0.05). When AD was added into co-culture medium, the levels of IL-4 and IFN-γ were lower than those in the non-interference groups and the mRNA expression levels decreased, RIG-I, IPS-1, IRF-7, IRF-3 and NF- K B decreased significantly in each group with significant statistic differences (P〈0.05). Conclusions: The RLRs mediated viral recognition provided a potential molecular target for acute viral infections and andrographolide could ameliorate H1N1 virus-induced cell mortality. And the antiviral effects might be related to its inhibition of viral-induced activation of the RLRs signaling pathway.
文摘Recent studies have reported that induced pluripotent stem (iPS) cells from mice and humans can differentiate into primordial germ cells. However, whether iPS cells are capable of producing male germ cells is not known. The objective of this study was to investigate the differentiation potential of mouse iPS cells into spermatogonial stem cells and late-stage male germ cells. We used an approach that combines in vitrodifferentiation and in vivotransplantation. Embryoid bodies (EBs) were obtained from iPS cells using leukaemia inhibitor factor (LIF)-free medium. Quantitative PCR revealed a decrease in Oct4 expression and an increase in StraSand Vasa mRNA in the EBs derived from iPS cells, iPS cell-derived EBs were induced by retinoic acid to differentiate into spermatogonial stem cells (SSCs), as evidenced by their expression of VASA, as well as CDH1 and GFRal, which are markers of SSCs. Furthermore, these germ cells derived from iPS cells were transplanted into recipient testes of mice that had been pre-treated with busulfan. Notably, iPS cell-derived SSCs were able to differentiate into male germ cells ranging from spermatogonia to round spermatids, as shown by VASA and SCP3 expression. This study demonstrates that iPS cells have the potential to differentiate into late-stage male germ cells. The derivation of male germ cells from iPS cells has potential applications in the treatment of male infertility and provides a model for uncovering the molecular mechanisms underlying male germ cell development.
基金Supported by the National Natural Science Foundation of China,No.81500449the Natural Science Foundation of Shanghai,No.14ZR1434200+2 种基金Shanghai Municipal Commission of Health and Family Planning,No.20144Y0175the Scientific Research Foundation for the Returned Overseas Chinese Scholarsthe State Education Ministry of China,No.20150909-6
文摘AIM To investigate the effect of(-)-epigallocatechin-3-gallate(EGCG) on polyinosinic-polycytidylic acid(poly I:C)-triggered intracellular innate immunity against hepatitis C virus(HCV) in hepatocytes. METHODS A cell culture model of HCV infection was generated by infecting a hepatoma cell line, Huh7, with HCV JFH-1 strain(JFH-1-Huh7). Poly I:C with a high molecular weight and EGCG were used to stimulate the JFH-1-Huh7 cells. Real-time reverse transcription-polymerase chain reaction was used to detect the expression levels of intracellular m RNAs and of intracellular and extracellular HCV RNA. Enzyme-linked immunosorbent assay was used to evaluate the interferon(IFN)-λ1 protein level in the cell culture supernatant. Immunostaining was used to examine HCV core protein expression in Huh7 cells.RESULTS Our recent study showed that HCV replication could impair poly I:C-triggered intracellular innate immune responses in hepatocytes. In the current study, we showed that EGCG treatment significantly increased the poly I:C-induced expression of Toll-like receptor 3(TLR3), retinoic acid-inducible gene I, and IFN-λ1 in JFH-1-Huh7 cells. In addition, supplementation with EGCG increased the poly I:C-mediated antiviral activity in JFH-1-Huh7 cells at the intracellular and extracellular HCV RNA and protein levels. Further investigation of the mechanisms showed that EGCG treatment significantly enhanced the poly I:C-induced expression of IFN-regulatory factor 9 and several antiviral IFNstimulated genes, including ISG15, ISG56, myxovirus resistance A, and 2'-5'-oligoadenylate synthetase 1, which encode the key antiviral elements in the IFN signaling pathway. CONCLUSION Our observations provide experimental evidence that EGCG has the ability to enhance poly I:C-induced intracellular antiviral innate immunity against HCV replication in hepatocytes.
文摘Objective The treatment of solid tumors by means of differentiation induction has not yet beengained such broken - through success as all-trans retinoic acid (ATRA) in acute promyelocytic leukemia (APL).To explore more effective inducers for solid tumor therapy, dimethyl sulfoxide (DMSO) is considered as acandidate. Methods In the present study, DMSO was used as inducer to human esophageal cancer cell lines invitro, in compared with classical inducer ATRA, in terms of morphology, cell cycle, growth inhibition, cytokeratin4 expression, gap junction-mediated dye transfer and tumorigenecity in nude mice. Results DMSO as well asATRA induced differentiation of human esophageal carcinoma cells. However, DMSO was confirmed to be moreeffective for induced differentiation of esophageal carcinoma cells than ATRA. Conclusion The results suggestedthat DMSO might have a good prospect in the treatment of solid tumors.