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1.
Leuk Lymphoma ; 60(5): 1312-1315, 2019 05.
Article in English | MEDLINE | ID: mdl-30322319

ABSTRACT

We hypothesized that ofatumumab with sequential methylprednisolone - alemtuzumab would be an effective and tolerable regimen for patients with high-risk chronic lymphocytic leukemia (CLL) with TP53 dysfunction. Thirty CLL patients with TP53 dysfunction (15 treatment naive (TN), 15 relapsed/refractory (R/R)) were enrolled in this phase II study. Therapy included ofatumumab with methylprednisolone for 2-4 monthly cycles, then ofatumumab with alemtuzumab for 4-24 weeks, then allogeneic transplantation or maintenance. The rate of overall response, complete response, marrow minimal residual disease (MRD) negativity, 3-year progression-free survival and overall survival were 80, 13, 80, 53, and 66%, respectively, in TN patients and 68, 0, 54, 25, and 53%, respectively, in R/R patients. Notable grade 3/4 toxicities included neutropenia and infection in 43 and 40% of patients, respectively. At median follow-up of 45 months, 13 patients died, and 10 patients are alive posttransplant. Overall, we observed high rates of MRD-negativity and acceptable tolerability in high-risk CLL.


Subject(s)
Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Chromosome Deletion , Chromosomes, Human, Pair 17 , Leukemia, Lymphocytic, Chronic, B-Cell/genetics , Leukemia, Lymphocytic, Chronic, B-Cell/therapy , Sequence Deletion , Tumor Suppressor Protein p53/genetics , Alemtuzumab/administration & dosage , Antibodies, Monoclonal, Humanized/administration & dosage , Antineoplastic Combined Chemotherapy Protocols/adverse effects , Combined Modality Therapy , Hematopoietic Stem Cell Transplantation/adverse effects , Hematopoietic Stem Cell Transplantation/methods , Humans , Leukemia, Lymphocytic, Chronic, B-Cell/diagnosis , Leukemia, Lymphocytic, Chronic, B-Cell/mortality , Methylprednisolone/administration & dosage , Prognosis , Transplantation, Homologous , Treatment Outcome
2.
Br J Haematol ; 184(6): 925-936, 2019 03.
Article in English | MEDLINE | ID: mdl-30537114

ABSTRACT

The L265P somatic mutation in the Myeloid Differentiation Primary Response 88 (MYD88) gene is a recurrent mutation in chronic lymphocytic leukaemia (CLL). This mutation has functional effects in various haematological malignancies but its role in CLL remains to be fully elucidated. Here, we report that MYD88 L265P mutations are associated with mutated immunoglobulin heavy-chain gene (IGHV-M) status and that among IGHV-M patients, the presence of MYD88 L265P is associated with younger age at diagnosis. Using microarray and RNA-Seq gene expression analysis, we further observe that the MYD88 L265P mutation is associated with a distinctive gene expression signature that predicts both failure-free survival and overall survival. This association was validated in an independent cohort of patients. To determine whether MYD88 L265P mutations can be therapeutically exploited in CLL, we treated primary cells with an inhibitor of interleukin 1 receptor-associated kinase 4 (IRAK4), a critical effector of the MYD88 pathway. IRAK4 inhibition decreased downstream nuclear factor-κB signalling and cell viability in CLL cells, indicating the potential of the MYD88 pathway as a therapeutic target in CLL.


Subject(s)
Leukemia, Lymphocytic, Chronic, B-Cell/genetics , Myeloid Differentiation Factor 88/genetics , Adult , Aged , Cohort Studies , Cytokines/biosynthesis , Female , Genes, Immunoglobulin Heavy Chain , Humans , Interleukin-1 Receptor-Associated Kinases/antagonists & inhibitors , Leukemia, Lymphocytic, Chronic, B-Cell/drug therapy , Leukemia, Lymphocytic, Chronic, B-Cell/metabolism , Male , Middle Aged , Molecular Targeted Therapy , Mutation , Myeloid Differentiation Factor 88/metabolism , Prognosis , Signal Transduction , Transcriptome
4.
Clin Cancer Res ; 23(3): 735-745, 2017 Feb 01.
Article in English | MEDLINE | ID: mdl-27503198

ABSTRACT

PURPOSE: Chronic lymphocytic leukemia (CLL) with 17p deletion typically progresses quickly and is refractory to most conventional therapies. However, some del(17p) patients do not progress for years, suggesting that del(17p) is not the only driving event in CLL progression. We hypothesize that other concomitant genetic abnormalities underlie the clinical heterogeneity of del(17p) CLL. EXPERIMENTAL DESIGN: We profiled the somatic mutations and copy number alterations (CNA) in a large group of del(17p) CLLs as well as wild-type CLL and analyzed the genetic basis of their clinical heterogeneity. RESULTS: We found that increased somatic mutation number associates with poor overall survival independent of 17p deletion (P = 0.003). TP53 mutation was present in 81% of del(17p) CLL, mostly clonal (82%), and clonal mutations with del(17p) exhibit shorter overall survival than subclonal mutations with del(17p) (P = 0.019). Del(17p) CLL has a unique driver mutation profile, including NOTCH1 (15%), RPS15 (12%), DDX3X (8%), and GPS2 (6%). We found that about half of del(17p) CLL cases have recurrent deletions at 3p, 4p, or 9p and that any of these deletions significantly predicts shorter overall survival. In addition, the number of CNAs, but not somatic mutations, predicts shorter time to treatment among patients untreated at sampling. Indolent del(17p) CLLs were characterized by absent or subclonal TP53 mutation and few CNAs, with no difference in somatic mutation number. CONCLUSIONS: We conclude that del(17p) has a unique genomic profile and that clonal TP53 mutations, 3p, 4p, or 9p deletions, and genomic complexity are associated with shorter overall survival. Clin Cancer Res; 23(3); 735-45. ©2016 AACR.


Subject(s)
Chromosome Deletion , Chromosomes, Human, Pair 17/ultrastructure , Leukemia, Lymphocytic, Chronic, B-Cell/genetics , Adult , Aged , Aged, 80 and over , Chromosome Breakage , Chromosomes, Human, Pair 17/genetics , Clone Cells , Disease Progression , Female , Gene Dosage , Germ-Line Mutation , Humans , In Situ Hybridization, Fluorescence , Kaplan-Meier Estimate , Leukemia, Lymphocytic, Chronic, B-Cell/blood , Leukemia, Lymphocytic, Chronic, B-Cell/drug therapy , Leukemia, Lymphocytic, Chronic, B-Cell/mortality , Male , Middle Aged , Mutation , Polymorphism, Single Nucleotide , Saliva/chemistry , Tumor Suppressor Protein p53/genetics , Exome Sequencing
5.
BMC Mol Biol ; 17(1): 20, 2016 08 30.
Article in English | MEDLINE | ID: mdl-27578267

ABSTRACT

BACKGROUND: Cells respond to numerous internal and external stresses, such as heat, cold, oxidative stress, DNA damage, and osmotic pressure changes. In most cases, the primary response to stress is transcriptional induction of genes that assist the cells in tolerating the stress and facilitate the repair of the cellular damage. However, when the transcription machinery itself is stressed, responding by such standard mechanisms may not be possible. RESULTS: In this study, we demonstrate that depletion or inactivation of RNA polymerase II (RNAPII) changes the preferred polyadenylation site usage for several transcripts, and leads to increased transcription of a specific subset of genes. Surprisingly, depletion of RNA polymerase I (RNAPI) also promotes altered polyadenylation site usage, while depletion of RNA polymerase III (RNAPIII) does not appear to have an impact. CONCLUSIONS: Our results demonstrate that stressing the transcription machinery by depleting either RNAPI or RNAPII leads to a novel transcriptional response that results in induction of specific mRNAs and altered polyadenylation of many of the induced transcripts.


Subject(s)
Alternative Splicing , Polyadenylation , RNA Polymerase II/genetics , RNA, Messenger/genetics , Saccharomyces cerevisiae Proteins/genetics , Saccharomyces cerevisiae/genetics , Transcription, Genetic , Gene Deletion
6.
Blood ; 127(25): 3215-24, 2016 06 23.
Article in English | MEDLINE | ID: mdl-27069256

ABSTRACT

BCL2 blunts activation of the mitochondrial pathway to apoptosis, and high-level expression is required for chronic lymphocytic leukemia (CLL) survival. Venetoclax (ABT-199) is a small-molecule selective inhibitor of BCL2 currently in clinical trials for CLL and other malignancies. In conjunction with the phase 1 first-in-human clinical trial of venetoclax in patients with relapsed or refractory CLL (M12-175), we investigated the mechanism of action of venetoclax in vivo, explored whether in vitro sensitivity assays or BH3 profiling correlated with in vivo responses in patients, and determined whether loss of TP53 function affected responses in vitro and in vivo. In all samples tested, venetoclax induced death of CLL cells in vitro at concentrations achievable in vivo, with cell death evident within 4 hours. Apoptotic CLL cells were detected in vivo 6 or 24 hours after a single 20-mg or 50-mg dose in some patients. The extent of mitochondrial depolarization by a BIM BH3 peptide in vitro was correlated with percentage reduction of CLL in the blood and bone marrow in vivo, whereas the half lethal concentration derived from standard cytotoxicity assays was not. CLL cell death in vitro and the depth of clinical responses were independent of deletion of chromosome 17p, TP53 mutation, and TP53 function. These data provide direct evidence that venetoclax kills CLL cells in a TP53-independent fashion by inhibition of BCL2 in patients and support further assessment of BH3 profiling as a predictive biomarker for this drug.


Subject(s)
Apoptosis/drug effects , Bridged Bicyclo Compounds, Heterocyclic/pharmacology , Leukemia, Lymphocytic, Chronic, B-Cell/pathology , Sulfonamides/pharmacology , Tumor Suppressor Protein p53/physiology , Animals , Cell Line, Tumor , Drug Resistance, Neoplasm/drug effects , Drug Resistance, Neoplasm/genetics , Gene Expression Regulation, Leukemic/drug effects , Humans , Leukemia, Lymphocytic, Chronic, B-Cell/genetics , Mice , Mice, Knockout , Proto-Oncogene Proteins c-bcl-2/antagonists & inhibitors , Signal Transduction/drug effects , Signal Transduction/genetics , Time Factors , Tumor Suppressor Protein p53/genetics
7.
Mol Cell Biochem ; 412(1-2): 165-71, 2016 Jan.
Article in English | MEDLINE | ID: mdl-26708217

ABSTRACT

Reactive oxygen species are a by-product of aerobic metabolism that can damage lipid, proteins, and nucleic acids. Oxidative damage to DNA is especially critical, because it can lead to cell death or mutagenesis. Previously we reported that the yeast sub1 deletion mutant is sensitive to hydrogen peroxide treatment and that the human SUB1 can complement the sensitivity of the yeast sub1 mutant. In this study, we find that Sub1 protects DNA from oxidative damage in vivo and in vitro. We demonstrate that transcription of SUB1 mRNA is induced by oxidative stress and that the sub1Δ mutant has an increased number of chromosomal DNA strand breaks after peroxide treatment. We further demonstrate that purified Sub1 protein can protect DNA from oxidative damage in vitro, using the metal ion catalyzed oxidation assay.


Subject(s)
DNA Damage , DNA-Binding Proteins/physiology , Oxidative Stress , Transcription Factors/physiology , Humans
8.
Leuk Lymphoma ; 56(12): 3336-42, 2015.
Article in English | MEDLINE | ID: mdl-25971907

ABSTRACT

Obatoclax is a small molecule mimetic of the BH3 domain of BCL-2 family proteins. This phase 1 study combining obatoclax with FR was undertaken in chronic lymphocytic leukemia (CLL) patients relapsed after at least one prior therapy. Obatoclax was given as a 3-h infusion on days 1 and 3 and escalated through three dose levels, with standard dose FR days 1-5. Thirteen patients were enrolled, with a median of two prior therapies. One dose-limiting toxicity (DLT) of a 2-week treatment delay for persistent grade 2-3 neutropenia was observed at the highest obatoclax dose (20 mg/m2), but no maximum tolerated dose (MTD) was reached. The overall response rate (ORR) was 85%, with 15% complete responses (CRs) by NCI-96 criteria and 54% by IWCLL 2008 criteria. Median time to progression was 20 months. It is concluded that obatoclax can be safely administered to relapsed CLL patients in combination with FR and shows promising clinical activity.


Subject(s)
Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Leukemia, Lymphocytic, Chronic, B-Cell/drug therapy , Leukemia, Lymphocytic, Chronic, B-Cell/pathology , Adult , Aged , Antineoplastic Combined Chemotherapy Protocols/adverse effects , Biomarkers , Chromosome Aberrations , Combined Modality Therapy , Disease Progression , Female , Humans , Indoles , Leukemia, Lymphocytic, Chronic, B-Cell/mortality , Male , Middle Aged , Mutation , Neoplasm Staging , Pyrroles/administration & dosage , Recurrence , Retreatment , Rituximab/administration & dosage , Treatment Outcome , Vidarabine/administration & dosage , Vidarabine/analogs & derivatives
9.
J Interferon Cytokine Res ; 35(3): 186-99, 2015 Mar.
Article in English | MEDLINE | ID: mdl-25330068

ABSTRACT

We demonstrate that interferon (IFN)-ß-1b induces an alternative-start transcript containing the C-terminal TLDc domain of nuclear receptor coactivator protein 7 (NCOA7), a member of the OXR family of oxidation resistance proteins. IFN-ß-1b induces NCOA7-AS (alternative start) expression in peripheral blood mononuclear cells (PBMCs) obtained from healthy individuals and multiple sclerosis patients and human fetal brain cells, astrocytoma, neuroblastoma, and fibrosarcoma cells. NCOA7-AS is a previously undocumented IFN-ß-inducible gene that contains only the last 5 exons of full-length NCOA7 plus a unique first exon (exon 10a) that is not found in longer forms of NCOA7. This exon encodes a domain closely related to an important class of bacterial aldo-keto oxido-reductase proteins that play a critical role in regulating redox activity. We demonstrate that NCOA7-AS is induced by IFN and LPS, but not by oxidative stress and exhibits, independently, oxidation resistance activity. We further demonstrate that induction of NCOA7-AS by IFN is dependent on IFN-receptor activation, the Janus kinase-signal transducers and activators of transcription (JAK-STAT) signaling pathway, and a canonical IFN-stimulated response element regulatory sequence upstream of exon 10a. We describe a new role for IFN-ßs involving a mechanism of action that leads to an increase in resistance to inflammation-mediated oxidative stress.


Subject(s)
Immunotherapy/methods , Interferon-beta/metabolism , Leukocytes, Mononuclear/physiology , Multiple Sclerosis/therapy , Nuclear Receptor Coactivators/metabolism , Protein Isoforms/metabolism , Amino Acid Sequence , Cell Line , Humans , Janus Kinases/metabolism , Molecular Sequence Data , Multiple Sclerosis/immunology , Nuclear Receptor Coactivators/genetics , Oxidation-Reduction , Oxidative Stress , Protein Isoforms/genetics , Protein Structure, Tertiary/genetics , Receptors, Interferon/metabolism , Regulatory Elements, Transcriptional/genetics , STAT Transcription Factors/metabolism , Signal Transduction
10.
PLoS One ; 8(3): e58015, 2013.
Article in English | MEDLINE | ID: mdl-23554872

ABSTRACT

Non homologous end joining (NHEJ) is an important process that repairs double strand DNA breaks (DSBs) in eukaryotic cells. Cells defective in NHEJ are unable to join chromosomal breaks. Two different NHEJ assays are typically used to determine the efficiency of NHEJ. One requires NHEJ of linearized plasmid DNA transformed into the test organism; the other requires NHEJ of a single chromosomal break induced either by HO endonuclease or the I-SceI restriction enzyme. These two assays are generally considered equivalent and rely on the same set of NHEJ genes. PC4 is an abundant DNA binding protein that has been suggested to stimulate NHEJ. Here we tested the role of PC4's yeast homolog SUB1 in repair of DNA double strand breaks using different assays. We found SUB1 is required for NHEJ repair of DSBs in plasmid DNA, but not in chromosomal DNA. Our results suggest that these two assays, while similar are not equivalent and that repair of plasmid DNA requires additional factor(s) that are not required for NHEJ repair of chromosomal double-strand DNA breaks. Possible roles for Sub1 proteins in NHEJ of plasmid DNA are discussed.


Subject(s)
Chromosome Breakage , Chromosomes, Fungal/metabolism , DNA Breaks, Double-Stranded , DNA End-Joining Repair/physiology , DNA, Fungal/metabolism , DNA-Binding Proteins/metabolism , Plasmids/metabolism , Saccharomyces cerevisiae Proteins/metabolism , Saccharomyces cerevisiae/metabolism , Transcription Factors/metabolism , Chromosomes, Fungal/genetics , DNA, Fungal/genetics , DNA-Binding Proteins/genetics , Plasmids/genetics , Saccharomyces cerevisiae/genetics , Saccharomyces cerevisiae Proteins/genetics , Transcription Factors/genetics
11.
Nucleic Acids Res ; 41(5): 3104-14, 2013 Mar 01.
Article in English | MEDLINE | ID: mdl-23355614

ABSTRACT

Alternative polyadenylation (APA) is conserved in all eukaryotic cells. Selective use of polyadenylation sites appears to be a highly regulated process and contributes to human pathogenesis. In this article we report that the yeast RPB2 gene is alternatively polyadenylated, producing two mRNAs with different lengths of 3'UTR. In normally growing wild-type cells, polyadenylation preferentially uses the promoter-proximal poly(A) site. After UV damage transcription of RPB2 is initially inhibited. As transcription recovers, the promoter-distal poly(A) site is preferentially used instead, producing more of a longer form of RPB2 mRNA. We show that the relative increase in the long RPB2 mRNA is not caused by increased mRNA stability, supporting the preferential usage of the distal poly(A) site during transcription recovery. We demonstrate that the 3'UTR of RPB2 is sufficient for this UV-induced regulation of APA. We present evidence that while transcription initiation rates do not seem to influence selection of the poly(A) sites of RPB2, the rate of transcription elongation is an important determinant.


Subject(s)
DNA Damage , Polyadenylation/radiation effects , RNA Polymerase II/genetics , Saccharomyces cerevisiae Proteins/genetics , Saccharomyces cerevisiae/genetics , 3' Untranslated Regions , Base Sequence , RNA Polymerase II/metabolism , RNA, Fungal/genetics , RNA, Fungal/metabolism , RNA, Messenger/genetics , RNA, Messenger/metabolism , Saccharomyces cerevisiae/metabolism , Saccharomyces cerevisiae/radiation effects , Saccharomyces cerevisiae Proteins/metabolism , Transcription Elongation, Genetic , Transcription Initiation, Genetic , Ultraviolet Rays
12.
Sheng Wu Yi Xue Gong Cheng Xue Za Zhi ; 29(5): 923-8, 2012 Oct.
Article in Chinese | MEDLINE | ID: mdl-23198436

ABSTRACT

Fuantai-03(FAT-03), isolated from the Dasyatis akajei, has a strong antiangiogenic activity. The recombinant Fuantai-03 (GST/rFAT-03) fusion protein can be obtained with the DNA recombination technology. In this study, expression conditions of GST/rFAT-03 were optimized by response surface experimental design method. The constructed engineering bacteria containing GST/rFAT-03 plasmid was induced by isopropy-beta-D-thiogalactosid (IPTG), the GST affinity column was used for isolation and purification, and then the effects of different culture time, IPTG concentration, induction temperature and induction time on the amount of soluble GST/rFAT-03 fusion protein were compared. The culture time for optimal expression was 6.13 h, IPTG concentration was 0.36 mmol/L, induction temperature was 19.71 degrees C, and induction time was 13.60 h. The amount of soluble GST/rFAT-03 fusion protein was 7.57 mg/L under above mentioned expression conditions. The results also showed that rFAT-03 significantly inhibited angiogenesis in chicken chorioallantoic membrane in a dose-dependent manner. Moreover, the soluble form of the target protein is useful for further work on purification and on studying its biological function.


Subject(s)
Angiogenesis Inhibitors/biosynthesis , Fish Proteins/biosynthesis , Recombinant Fusion Proteins/biosynthesis , Skates, Fish , Angiogenesis Inhibitors/genetics , Animals , Chickens , Chorioallantoic Membrane/blood supply , Escherichia coli/genetics , Escherichia coli/metabolism , Fish Proteins/genetics , Recombinant Fusion Proteins/genetics
13.
Cancer Chemother Pharmacol ; 62(4): 559-68, 2008 Sep.
Article in English | MEDLINE | ID: mdl-18030471

ABSTRACT

BACKGROUND: Tubeimoside I (TBMS1) was isolated from the tubers of Bolbostemma paniculatum (Maxim.) Franquet. TBMS1 shows potent anti-tumor activity. The present study was conducted to investigate the anti-microtubule role of TBMS1 and its binding site of tubulin. METHODS: Cell growth inhibition was measured by MTT after treatment with TBMS1. Uptake kinetics of TBMS1 by human nasopharyngeal carcinoma CNE-2Z cell line (CNE-2Z) was assayed by HPLC. Microtubule protein (MTP) was prepared from porcine brain through two cycles of polymerization-depolymerization in a high molarity buffer. Inhibition of MTP polymerization induced by TBMS1 was determined by a turbidity measurement and a sedimentation assay; the interactions of TBMS1 with tubulin within CNE-2Z cells were investigated by immunofluorescence microscopy and immunoblotting. TBMS1 was tested for its ability to inhibit binding of known tubulin ligands through competitive binding assay. RESULTS: TBMS1 displayed growth inhibitory activity against CNE-2Z cells with IC(50) value of 16.7 microM for 72 h. HPLC analysis of TBMS1 uptake by CNE-2Z cells displayed the initial slow TBMS1 uptake and then gradually reaching an maximum uptake near 18 h. CNE-2Z cells treated with TBMS1 (25 microM, 3 h) were sufficient to cause the microtubular network disruption. Immunoblot analysis showed that the proportion of cytosolic tubulin of cells treated with TBMS1 increased in a time- and concentration-dependent manner. TBMS1 did not inhibit the binding of vinblastine to tubulin. Colchicine binding to tubulin was inhibited in the presence of TBMS1. CONCLUSIONS: TBMS1 is an anti-microtubule agent, and its binding site of tubulin is the colchicine binding site of tubulin.


Subject(s)
Antineoplastic Agents, Phytogenic/pharmacology , Colchicine/metabolism , Drugs, Chinese Herbal/pharmacology , Saponins/pharmacology , Triterpenes/pharmacology , Tubulin Modulators/pharmacology , Tubulin/metabolism , Animals , Antineoplastic Agents, Phytogenic/metabolism , Binding Sites/drug effects , Carcinoma/drug therapy , Carcinoma/metabolism , Cell Line, Tumor , Cell Proliferation/drug effects , Chromatography, High Pressure Liquid , Dose-Response Relationship, Drug , Drugs, Chinese Herbal/metabolism , Humans , Microtubule Proteins/drug effects , Microtubule Proteins/metabolism , Nasopharyngeal Neoplasms/drug therapy , Nasopharyngeal Neoplasms/metabolism , Polymers/chemistry , Saponins/metabolism , Swine , Triterpenes/metabolism , Tubulin/chemistry , Tubulin Modulators/metabolism
14.
Neurosci Bull ; 23(4): 209-14, 2007 Jul.
Article in English | MEDLINE | ID: mdl-17687395

ABSTRACT

OBJECTIVE: To investigate a possibility of repairing damaged brain by intracerebroventricular transplantation of neural stem cells (NSCs) in the adult mice subjected to glutamate-induced excitotoxic injury. METHODS: Mouse NSCs were isolated from the brains of embryos at 15-day postcoitum (dpc). The expression of nestin, a special antigen for NSC, was detected by immunocytochemistry. Immunofluorescence staining was carried out to observe the survival and location of transplanted NSCs. The animals in the MSG + NSCs group received intracerebroventricular transplantation of NSCs (approximately 1.0 x 10(5) cells) separately on day 1 and day 10 after 10-d MSG exposure (4.0 g/kg per day). The mice in control and MSG groups received intracerebroventricular injection of Dulbecco's minimum essential medium (DMEM) instead of NSCs. On day 11 after the last NSC transplantation, the test of Y-maze discrimination learning was performed, and then the histopathology of the animal brains was studied to analyze the MSG-induced functional and morphological changes of brain and the effects of intracerebroventricular transplantation of NSCs on the brain repair. RESULTS: The isolated cells were Nestin-positive. The grafted NSCs in the host brain were region-specifically survived at 10-d post-transplantation. Intracerebroventricular transplantation of NSCs obviously facilitated the brain recovery from glutamate-induced behavioral disturbances and histopathological impairs in adult mice. CONCLUSION: Intracerebroventricular transplantation of NSCs may be feasible in repairing diseased or damaged brain tissue.


Subject(s)
Glutamic Acid/toxicity , Neurons/physiology , Neurotoxicity Syndromes , Stem Cell Transplantation/methods , Stem Cells/physiology , Animals , Cell Count , Disease Models, Animal , Embryo, Mammalian , Injections, Intraventricular/methods , Intermediate Filament Proteins/metabolism , Mice , Mice, Inbred Strains , Nerve Tissue Proteins/metabolism , Nestin , Neurotoxicity Syndromes/etiology , Neurotoxicity Syndromes/pathology , Neurotoxicity Syndromes/surgery , Time Factors
15.
Nan Fang Yi Ke Da Xue Xue Bao ; 27(5): 679-81, 2007 May.
Article in Chinese | MEDLINE | ID: mdl-17545088

ABSTRACT

OBJECTIVE: To observe the ultrastructural changes of HeLa cells in response to tubeimoside I (TBMS1) treatment and the protective effect of cyclosporine A (CsA), and explore the role of intrinsic apoptosis pathway in TBMS1-induced HeLa cell apoptosis. METHODS: HeLa cells were treated with TBMS1 (10-50 micromo/L) alone or in combination with 2 micromol/L CsA for 12 and 24 h and observed with transmission electron microscope (TEM) for the ultrastructural changes of the cells. RESULTS: TBMS1 induced apoptosis of HeLa cells in a concentration- and time-dependant manner. Under TEM, the treated cells progressively shrunk and the intercellular space widened with loss of microvillus, mitochondrial swelling, rough endoplasmic reticulum enlargement, chromatin condensation, nuclear shrinkage and nuclear pyknosis as TBMS1 concentration increased. At low concentrations, CsA offered partial protection of the mitochondria from TBMS1-induced damage whereas high-concentration CsA did not. CONCLUSION: TBMS1 induces ultrastructural changes typical for apoptosis of the HeLa cells, which provides morphological evidence for the role of intrinsic apoptosis pathway in TBMS1-induced apoptosis.


Subject(s)
Apoptosis/drug effects , Cyclosporine/pharmacology , Saponins/pharmacology , Triterpenes/pharmacology , Cell Nucleus/drug effects , Cell Nucleus/ultrastructure , Dose-Response Relationship, Drug , Endoplasmic Reticulum, Rough/drug effects , Endoplasmic Reticulum, Rough/ultrastructure , Female , HeLa Cells , Humans , Immunosuppressive Agents/pharmacology , Microscopy, Electron, Transmission , Mitochondrial Swelling/drug effects , Time Factors , Uterine Cervical Neoplasms/pathology , Uterine Cervical Neoplasms/ultrastructure
16.
Nan Fang Yi Ke Da Xue Xue Bao ; 26(8): 1132-5, 2006 Aug.
Article in Chinese | MEDLINE | ID: mdl-16939901

ABSTRACT

OBJECTIVE: To study the effect of saikosaponins, the active ingredients of Bupleurum chinense DC. on glutamate and GABA expressions in the hippocampus of slow kindling rats induced by pentetrazole. METHODS: Forty-eight healthy Sprague-Dawley rats were randomly divided into 6 equal groups, namely the blank control group (group A), normal saline (NS) group (group B), sodium valproate group (group C), and 3 saikosaponins groups of high, medium and small doses (groups D, E, and F, respectively). The rats in each group other than group A were given corresponding treatments after slow kindling by pentetrazole. After 4 weeks of treatment, the rats were sacrifices and the brain tissues were sampled, sliced and stained by immunohistochemically, and the results were analyzed according to the positive cell number and gray scale. RESULTS: In CA1 region, the glutamate-positive cell number and gray scale of group B was significantly different from the other groups (P<0.05), but such difference was not observed in the CA2 and DG (P>0.05); In CA1, CA2 and DG of the hippocampus, the GABA-positive cell number of group B was significantly greater but the gray scale lower than those of the other groups (P<0.05). In CA1 and CA2 regions of the hippocampus, the glutamate- and GABA-positive cell ratio of group B was significantly lower than that of the other groups (P<0.05), but in CA1, CA2, and DG region of the hippocampus, the ratio of gray scale between glutamate- and GABA-positive cells was comparable between the groups (P>0.05). CONCLUSION: The expression of glutamate and GABA, especially the latter, increased in chronic kindling rat hippocampus. Saikosaponins intervene in such changes of glutamate and GABA to contain their expressions within normal range, which may be one of the mechanisms of saikosaponins to inhibit slow kindling induced by pentetrazole.


Subject(s)
Glutamic Acid/biosynthesis , Hippocampus/drug effects , Kindling, Neurologic/metabolism , Oleanolic Acid/analogs & derivatives , Saponins/pharmacology , gamma-Aminobutyric Acid/biosynthesis , Animals , Female , Hippocampus/metabolism , Immunohistochemistry , Male , Oleanolic Acid/pharmacology , Pentylenetetrazole , Random Allocation , Rats , Rats, Sprague-Dawley
17.
Nan Fang Yi Ke Da Xue Xue Bao ; 26(4): 452-5, 2006 Apr.
Article in Chinese | MEDLINE | ID: mdl-16624750

ABSTRACT

OBJECTIVE: To study the effect of saikosaponins, the active ingredients of Bupleurum chinense DC, on glial fibrillary acidic protein (GFAP) expression in hippocampal astrocytes of chronic kindling rats induced by pentetrazole (PTZ). METHODS: Forty-eight healthy Sprague-Dawley rats were randomized into 6 equal groups, namely the blank control group (Group A), normal saline group (Group B), sodium valproate group (Group C), and 3 saikosaponins groups of high, medium and small doses (Groups D, E, and F, respectively). The rats (except those in Group A) received intraperitoneal injection of PTZ to induce chronic kindling 1 h after the respective agents as indicated were administered intragastrically on a daily basis for 4 consecutive weeks. Upon completion of the treatment course, the rats were sacrificed and the brain tissues were sampled, sliced and stained for immunohistochemical examination. The results were analyzed to calculate the positive cell count, cross-sectional area of the cells and the gray scale. RESULTS: In group B, the positive cell population and cross-sectional area of the positive cells were the greatest among the groups (P<0.01), but the positive cell gray scale of the CA1 and CA2 regions and the dentate gyrus (DG) of the hippocampus was the lowest. The CA1 region of Group B was significantly different from that of groups A, C and D (P<0.01), and the CA2 region different from groups A, C, D and E (P<0.05), while the DG different from group F (P<0.05) and groups A, C, D and E (P<0.01). CONCLUSION: In chronic kindling rats induced by PTZ, GFAP overexpression can be inhibited by saikosaponins, which suppress the abnormal activation of hippocampal astrocyte of the kindling rats.


Subject(s)
Glial Fibrillary Acidic Protein/biosynthesis , Hippocampus/metabolism , Kindling, Neurologic/metabolism , Oleanolic Acid/analogs & derivatives , Pentylenetetrazole , Saponins/pharmacology , Animals , Astrocytes/metabolism , Bupleurum/chemistry , Depression, Chemical , Female , Glial Fibrillary Acidic Protein/genetics , Male , Oleanolic Acid/pharmacology , Random Allocation , Rats , Rats, Sprague-Dawley
18.
Nan Fang Yi Ke Da Xue Xue Bao ; 26(2): 177-80, 2006 Feb.
Article in Chinese | MEDLINE | ID: mdl-16503523

ABSTRACT

OBJECTIVE: To study the effect of saikosaponins, the active ingredients of Bupleurum chinense DC, on epileptic seizure and EEG of pentetrazole (PTZ)-induced chronic kindling rats. METHODS: Forty-eight healthy Sprague-Dawley rats were randomly divided into 6 equal groups, namely the blank control group, normal saline (NS) group, sodium valproate (VPA) group, and 3 saikosaponins groups of high, medium and small doses. Except those in the blank control group, the rats in the other groups were all given different treatments as specified prior to intraperitoneal PTZ injection to induce kindling on a daily basis for 4 consecutive weeks. Epileptic seizures of the rats were recorded during the treatment and EEG recorded at the end of the treatments. RESULTS: Seizure frequency in the 3 saikosaponins groups decreased 2 weeks later, which was especially obvious in the high-dose group (P<0.05). The kindling rate was significantly lower in high-dose saikosaponins group than in the other treatment groups after 4 weeks of the treatment (P<0.05), with also less intense seizure onset (P<0.01) and differences in the wave form of EEG. CONCLUSION: Saikosaponins can inhibit PTZ-induced epileptic seizure in kindling rats and antagonize the kindling effect of PTZ.


Subject(s)
Electroencephalography/drug effects , Epilepsy/physiopathology , Kindling, Neurologic/drug effects , Oleanolic Acid/analogs & derivatives , Saponins/pharmacology , Animals , Anticonvulsants/pharmacology , Epilepsy/chemically induced , Female , Male , Oleanolic Acid/pharmacology , Pentylenetetrazole , Random Allocation , Rats , Rats, Sprague-Dawley
19.
Cancer Chemother Pharmacol ; 57(3): 389-99, 2006 Feb.
Article in English | MEDLINE | ID: mdl-16172906

ABSTRACT

BACKGROUND: Tubeimoside I (TBMS1), a triterpenoid saponin, isolated from the tubers of Bolbostemma paniculatum, showed potent antitumor and antitumor-promoting effects. The objective of this study is to investigate the role of mitochondria and mitochondria cytochrome c in TBMS1-mediated apoptosis of human cervical carcinoma HeLa cell line. METHODS: Viability of HeLa cells was measured by MTT assay. Apoptotic induction by TBMS1 was determined by fluorescence microscopy, flow cytometry and gel electrophoresis of fragmented DNA. Mitochondrial transmembrane potential (Deltapsim) was assayed by flow cytometry. Cytochrome c (Cyt c) was detected by Western blotting. RESULTS: The results showed that Cyclosporin A (CsA) partly protected HeLa cells from growth inhibitory effect of TBMS1, and partly countered the ability of TBMS1 to rapidly induce apoptosis in HeLa cells, and that TBMS1 decreased Deltapsim and induced Cyt c release by a mechanism inhibited by CsA, and that TBMS1 induced apoptosis of HeLa cells dose-dependently in accordance with increase of cytosolic Cyt c. CONCLUSIONS: TBMS1 opens the permeability transition (PT) pore, thereby decreasing Deltapsim, releasing Cyt c from mitochondria, and further causing a series of events consistent with established mechanistic models of apoptosis.


Subject(s)
Apoptosis/drug effects , Cytochromes c/physiology , Mitochondria/physiology , Saponins/pharmacology , Triterpenes/pharmacology , Acridine Orange , Annexin A5 , Apoptosis/physiology , Blotting, Western , Cell Membrane/chemistry , Cell Membrane/drug effects , Cell Nucleus/drug effects , Cell Proliferation/drug effects , Chromatin Assembly and Disassembly/drug effects , Cucurbitaceae/chemistry , Cyclosporine/pharmacology , Cytochromes c/metabolism , DNA Fragmentation/drug effects , Dose-Response Relationship, Drug , Drugs, Chinese Herbal/chemistry , Drugs, Chinese Herbal/pharmacology , Ethidium , HeLa Cells , Humans , Microscopy, Electron , Microscopy, Fluorescence/methods , Mitochondria/drug effects , Mitochondria/ultrastructure , Molecular Structure , Propidium , Saponins/chemistry , Triterpenes/chemistry
20.
Eur Neuropsychopharmacol ; 16(3): 170-7, 2006 Apr.
Article in English | MEDLINE | ID: mdl-16257184

ABSTRACT

The present study was conducted to investigate a possible protection of ferulic acid against excitotoxic effects of maternal intragastric (ig) administration of monosodium glutamate (MSG) at a late stage of pregnancy on developing mouse fetal brain. [(3)H]-labeled glutamate was used as radiotracer to study the effect of ferulic acid on distribution of MSG in mouse fetal brain. MSG dissolved in distilled water (2.0 g/kg body weight, 640 kBq of [(3)H]glutamate/mouse, ig) or/and sodium ferulate (SF) (20, 40, 80 mg/kg body weight, ip), was given to pregnant mice at 17-19 days; the distribution of [(3)H] glutamate in the mouse fetal brains was measured at 30, 60, 90, 120 min after administration of MSG or/and SF. Maternal mice were given MSG (1.0, 2.0, 4.0 g/kg body weight, ig) or/and SF (20, 40, 80 mg/kg body weight, ip) simultaneously at 17-19 days of pregnancy, and then behavioral tests and histopathological observations were used to analyze glutamate-induced functional and morphological changes of the brains of their offspring, and Western blot analysis was performed for examining expressions of bcl-2 and caspase-3. The results showed that SF obviously inhibited the uptake of labeled glutamate in fetal brain. In addition, SF countered the effects of MSG on behavior, histopathology, genetic toxicity, and expression of apoptosis-related gene. The results suggest that ferulic acid is a novel competitive N-methyl-D-aspartate (NMDA) receptor antagonist and neuroprotector. In conclusion, maternal administration of ferulic acid has potent protective effects against glutamate-induced neurotoxicity in their filial mice.


Subject(s)
Brain/growth & development , Coumaric Acids/pharmacology , Food Additives/toxicity , Free Radical Scavengers/pharmacology , Sodium Glutamate/antagonists & inhibitors , Sodium Glutamate/toxicity , Animals , Apoptosis/genetics , Behavior, Animal/drug effects , Blotting, Western , Brain/drug effects , Brain/metabolism , Brain/pathology , Discrimination Learning/drug effects , Female , Glutamic Acid/metabolism , Intubation, Gastrointestinal , Male , Maze Learning/drug effects , Mice , Molecular Conformation , Motor Activity/drug effects , Pregnancy , Sodium Glutamate/administration & dosage
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