RÉSUMÉ
Wnt10b, an endogenous inhibitor of adipogenesis, maintains preadipocytes in an undifferentiated state by suppressing adipogenic transcription factors. We have previously demonstrated that Wnt10b transcription during adipogenesis is negatively regulated by X-box-binding protein 1 (XBP1), an important transcription factor of the unfolded protein response. In this report, we demonstrate that XBP1s can directly induce the transcription of microRNA-148a, which in turn mediates the silencing of Wnt10b mRNA during adipogenic differentiation of 3T3-L1 cells. Stability of Wnt10b mRNA was found to be significantly increased by knockdown of XBP1s. Using computational algorithms, a set of microRNAs was predicted to bind Wnt10b mRNA, of which microRNA-148a was selected as a potential target for XBP1s. Our results revealed that microRNA-148a could bind to the 3′UTR of Wnt10b mRNA. Its ectopic expression significantly suppressed both Wnt10b expression and β-catenin activity. When we altered the expression of XBP1 in 3T3-L1 cells, microRNA-148a levels changed accordingly. A potential XBP1 response element was found in the promoter region of microRNA-148a, and XBP1s directly bound to this response element as shown by point mutation analysis and chromatin immunoprecipitation assay. In addition, a microRNA-148a mimic significantly restored adipogenic potential in XBP1-deficient 3T3-L1 cells. These findings provide the first evidence that XBP1s can regulate Wnt10b by a post-transcriptional mechanism through directly inducing microRNA-148a.
Sujet(s)
Cellules 3T3-L1 , Adipogenèse , Immunoprécipitation de la chromatine , Expression génique ectopique , microARN , Mutation ponctuelle , Régions promotrices (génétique) , Éléments de réponse , ARN messager , Facteurs de transcription , Réponse aux protéines mal repliéesRÉSUMÉ
Wnt10b, an endogenous inhibitor of adipogenesis, maintains preadipocytes in an undifferentiated state by suppressing adipogenic transcription factors. We have previously demonstrated that Wnt10b transcription during adipogenesis is negatively regulated by X-box-binding protein 1 (XBP1), an important transcription factor of the unfolded protein response. In this report, we demonstrate that XBP1s can directly induce the transcription of microRNA-148a, which in turn mediates the silencing of Wnt10b mRNA during adipogenic differentiation of 3T3-L1 cells. Stability of Wnt10b mRNA was found to be significantly increased by knockdown of XBP1s. Using computational algorithms, a set of microRNAs was predicted to bind Wnt10b mRNA, of which microRNA-148a was selected as a potential target for XBP1s. Our results revealed that microRNA-148a could bind to the 3′UTR of Wnt10b mRNA. Its ectopic expression significantly suppressed both Wnt10b expression and β-catenin activity. When we altered the expression of XBP1 in 3T3-L1 cells, microRNA-148a levels changed accordingly. A potential XBP1 response element was found in the promoter region of microRNA-148a, and XBP1s directly bound to this response element as shown by point mutation analysis and chromatin immunoprecipitation assay. In addition, a microRNA-148a mimic significantly restored adipogenic potential in XBP1-deficient 3T3-L1 cells. These findings provide the first evidence that XBP1s can regulate Wnt10b by a post-transcriptional mechanism through directly inducing microRNA-148a.
Sujet(s)
Cellules 3T3-L1 , Adipogenèse , Immunoprécipitation de la chromatine , Expression génique ectopique , microARN , Mutation ponctuelle , Régions promotrices (génétique) , Éléments de réponse , ARN messager , Facteurs de transcription , Réponse aux protéines mal repliéesRÉSUMÉ
With the increasing use of culture-expanded mesenchymal stromal cells (MSCs) for cell therapies, factors that regulate the cellular characteristics of MSCs have been of major interest. Oxygen concentration has been shown to influence the functions of MSCs, as well as other normal and malignant stem cells. However, the underlying mechanisms of hypoxic responses and the precise role of hypoxia-inducible factor-1alpha (Hif-1alpha), the master regulatory protein of hypoxia, in MSCs remain unclear, due to the limited span of Hif-1alpha stabilization and the complex network of hypoxic responses. In this study, to further define the significance of Hif-1alpha in MSC function during their self-renewal and terminal differentiation, we established adult bone marrow (BM)-derived MSCs that are able to sustain high level expression of ubiquitin-resistant Hif-1alpha during such long-term biological processes. Using this model, we show that the stabilization of Hif-1alpha proteins exerts a selective influence on colony-forming mesenchymal progenitors promoting their self-renewal and proliferation, without affecting the proliferation of the MSC mass population. Moreover, Hif-1alpha stabilization in MSCs led to the induction of pluripotent genes (oct-4 and klf-4) and the inhibition of their terminal differentiation into osteogenic and adipogenic lineages. These results provide insights into the previously unrecognized roles of Hif-1alpha proteins in maintaining the primitive state of primary MSCs and on the cellular heterogeneities in hypoxic responses among MSC populations.
Sujet(s)
Humains , Différenciation cellulaire , Prolifération cellulaire , Sous-unité alpha du facteur-1 induit par l'hypoxie/génétique , Facteurs de transcription Krüppel-like/génétique , Cellules souches mésenchymateuses/cytologie , Facteur de transcription Oct-3/génétique , Stabilité protéiqueRÉSUMÉ
Brown adipose tissue is specialized to burn lipids for thermogenesis and energy expenditure. Second-generation antipsychotics (SGA) are the most commonly used drugs for schizophrenia with several advantages over first-line drugs, however, it can cause clinically-significant weight gain. To reveal the involvement of brown adipocytes in SGA-induced weight gain, we compared the effect of clozapine, quetiapine, and ziprasidone, SGA with different propensities to induce weight gain, on the differentiation and the expression of brown fat-specific markers, lipogenic genes and adipokines in a mouse brown preadipocyte cell line. On Oil Red-O staining, the differentiation was inhibited almost completely by clozapine (40 microM) and partially by quetiapine (30 microM). Clozapine significantly down-regulated the brown adipogenesis markers PRDM16, C/EBPbeta, PPARgamma2, UCP-1, PGC-1alpha, and Cidea in dose- and time-dependent manners, whereas quetiapine suppressed PRDM16, PPARgamma2, and UCP-1 much weakly than clozapine. Clozapine also significantly inhibited the mRNA expressions of lipogenic genes ACC, SCD1, GLUT4, aP2, and CD36 as well as adipokines such as resistin, leptin, and adiponectin. In contrast, quetiapine suppressed only resistin and leptin but not those of lipogenic genes and adiponectin. Ziprasidone (10 microM) did not alter the differentiation as well as the gene expression patterns. Our results suggest for the first time that the inhibition of brown adipogenesis may be a possible mechanism to explain weight gain induced by clozapine and quetiapine.
Sujet(s)
Animaux , Humains , Souris , Adipocytes bruns/effets des médicaments et des substances chimiques , Adipogenèse/effets des médicaments et des substances chimiques , Adipokines/métabolisme , Neuroleptiques/administration et posologie , Différenciation cellulaire/effets des médicaments et des substances chimiques , Lignée cellulaire , Survie cellulaire/effets des médicaments et des substances chimiques , Clozapine/administration et posologie , Dibenzothiazépines/administration et posologie , Régulation de l'expression des gènes/effets des médicaments et des substances chimiques , Pipérazines/administration et posologie , Schizophrénie/traitement médicamenteux , Thiazoles/administration et posologie , Prise de poids/effets des médicaments et des substances chimiquesRÉSUMÉ
OBJECTIVE: Haloperidol, a typical antipsychotic, has been the preferred agent for the pharmacological treatment of delirium. Recent studies have shown that atypical antipsychotics can be as effective as haloperidol in managing delirium. However, there are few comparative studies between atypical antipsychotics in the treatment of delirium. We investigated the efficacy and side effects of aripiprazole and quetiapine for the treatment of patients with delirium. METHODS: Forty two inpatients with delirium according to the Diagnostic and Statistical Manual of Mental Disorders, 4th edition. Text Revision and Korean version of Delirium Rating Scale-Revised-98 (K-DRS-98) criteria were included. They were assigned to either aripiprazole or quetiapine groups, with a flexible dosing schedule. K-DRS-98 and Clinical Global Impression-Severity (CGI-S) were used for evaluating the severity of delirium. The degree of sedation was assessed by using the Richmond Agitation-Sedation Scale (RASS) six times per day. The severity of side effect was evaluated with the Drug-Induced ExtraPyramidal Symptoms Scale and the Barnes Akathisia Rating Scale. K-DRS-98 and RASS were conducted daily until the remission of delirium while other measurements were conducted twice at the point of baseline and remission. For statistical analysis, t-test, Fisher's exact test, Mann-Whitney test, analysis of covariance were conducted. RESULTS: The scores of K-DRS-98 in both groups significantly decreased after treatment (p or =-3 (p=0.034). The scores on sleep cycle of K-DRS-98-severity more significantly decreased in the quetiapine group than aripiprazole group (F=4.291, p=0.045). There were no significant side effects both groups including extrapyramidal symptoms. CONCLUSION: These results suggest that both aripiprazole and quetiapine appear to be effective and tolerable in the treatment of delirium. Aripiprazole may be less sedative than quetiapine and it may be more useful than aripiprazole in sleep problem of delirium. To validate our results, further studies with double-blind, placebo-controlled with a large sample will be required.
Sujet(s)
Humains , Neuroleptiques , Rendez-vous et plannings , Délire avec confusion , Diagnostic and stastistical manual of mental disorders (USA) , Dibenzothiazépines , Halopéridol , Patients hospitalisés , Pipérazines , Agitation psychomotrice , QuinolinoneRÉSUMÉ
Golgi SNAP receptor complex 1 (GS28) has been implicated in vesicular transport between intra-Golgi networks and between endoplasmic reticulum (ER) and Golgi. Additional role(s) of GS28 within cells have not been well characterized. We observed decreased expression of GS28 in rat ischemic hippocampus. In this study, we examined the role of GS28 and its molecular mechanisms in neuronal (SK-N-SH) cell death induced by hydrogen peroxide (H2O2). GS28 siRNA-transfected cells treated with H2O2 showed a significant increase in cytotoxicity under glutathione (GSH)-depleted conditions after pretreatment with buthionine sulfoximine, which corresponded to an increase of intracellular reactive oxygen species (ROS) in the cells. Pretreatment of GS28 siRNA-transfected cells with p38 chemical inhibitor significantly inhibited cytotoxicity; we also observed that p38 was activated in the cells by immunoblot analysis. We confirmed the role of p38 MAPK in cotransfected cells with GS28 siRNA and p38 siRNA in the cell viability assay, flow cytometry, and immunoblot. Involvement of apoptotic or autophagic processes in the cells was not shown in the cell viability, flow cytometry, and immunoblot analyses. However, pretreatment of the cells with necrostatin-1 completely inhibited H2O2-induced cytotoxicity, ROS generation, and p38 activation, indicating that the cell death is necroptotic. Collectively these data imply that H2O2 induces necroptotic cell death in the GS28 siRNA-transfected cells and that the necroptotic signals are mediated by sequential activations in RIP1/p38/ROS. Taken together, these results indicate that GS28 has a protective role in H2O2-induced necroptosis via inhibition of p38 MAPK in GSH-depleted neuronal cells.
Sujet(s)
Animaux , Rats , Buthionine sulfoximine , Mort cellulaire , Survie cellulaire , Réticulum endoplasmique , Cytométrie en flux , Glutathion , Hippocampe , Hydrogène , Peroxyde d'hydrogène , Imidazoles , Indoles , Méthionine , Neurones , p38 Mitogen-Activated Protein Kinases , Espèces réactives de l'oxygène , Petit ARN interférent , Protéines SNARERÉSUMÉ
PURPOSE: Topiramate is an antiepileptic drug used widely in the treatment of epilepsy. It has also been reported to reduce body weight in humans and is currently used for eating disorders and obesity; little is known about the mechanism by which this drug induces weight loss. This study was carried out to investigate the effects of topiramate on weight and serum levels of insulin and leptin in young rats fed high fat diet (HFD). METHODS: Forty male Wistar rats (4 weeks old) were randomly divided into four groups (diet, regular diet and high fat diet, treatment, topiramate and placebo). Topiramate (50 mg/kg/day) was orally administered via gastric gavage twice a day for 4 weeks. Food intake and body weight were monitored throughout the study. After 4 weeks, the plasma levels of glucose, triglycerides, free fatty acids, leptin, and insulin were determined. RESULTS: We found significant reduction in body weight gain after topiramate administration in the HFD group (topiramate, 351.6 +/- 28.6g; placebo, 397.6 +/- 28.4g) (P<0.05). Topiramate was able to reduce food intake in both diet groups (P<0.05). Furture, fasting glucose levels were significantly lower in both topiramate groups than placebo groups (P<0.05), and serum leptin levels in the HFD group were decreased (P<0.05). Additionally, there was no significant difference in serum levels of triglycerides, free fatty acids, or insulin between the four groups. CONCLUSION: Topiramate significantly inhibited body weight gain by reducing food intake, especially in the HFD group and reduced serum levels of glucose in both diet groups and of leptin in the HFD group.
Sujet(s)
Animaux , Humains , Mâle , Rats , Poids , Régime alimentaire , Alimentation riche en graisse , Consommation alimentaire , Troubles de l'alimentation , Épilepsie , Jeûne , Acide gras libre , Fructose , Glucose , Insuline , Leptine , Plasma sanguin , Rat Wistar , Triglycéride , Perte de poidsRÉSUMÉ
When we treated rat bone marrow stromal cells (rBMSCs) with neuronal differentiation induction media, typical unfolded protein response (UPR) was observed. BIP/GRP78 protein expression was time-dependently increased, and three branches of UPR were all activated. ATF6 increased the transcription of XBP1 which was successfully spliced by IRE1. PERK was phosphorylated and it was followed by eIF2alpha phosphorylation. Transcription of two downstream targets of eIF2alpha, ATF4 and CHOP/GADD153, were transiently up-regulated with the peak level at 24 h. Immunocytochemical study showed clear coexpression of BIP and ATF4 with NeuN and Map2, respectively. UPR was also observed during the neuronal differentiation of mouse embryonic stem (mES) cells. Finally, chemical endoplasmic reticulum (ER) stress inducers, thapsigargin, tunicamycin, and brefeldin A, dose-dependently increased both mRNA and protein expressions of NF-L, and, its expression was specific to BIP-positive rBMSCs. Our results showing the induction of UPR during neuronal differentiations of rBMSCs and mES cells as well as NF-L expression by ER stress inducers strongly suggest the potential role of UPR in neuronal differentiation.
Sujet(s)
Animaux , Souris , Rats , Facteur de transcription ATF-4/génétique , Apoptose/effets des médicaments et des substances chimiques , Cellules de la moelle osseuse/cytologie , Différenciation cellulaire , Milieux de culture/pharmacologie , Cellules souches embryonnaires/cytologie , Réticulum endoplasmique/génétique , Expression des gènes/effets des médicaments et des substances chimiques , Protéines du choc thermique/génétique , Protéines associées aux microtubules/génétique , Chaperons moléculaires/génétique , Protéines de tissu nerveux/génétique , Protéines neurofilamenteuses/génétique , Neurones/cytologie , Protéines nucléaires/génétique , Pliage des protéines , Cellules stromalesRÉSUMÉ
Expressions of endoplasmic reticulum stress response (ERSR) genes were examined during the neuronal differentiation of rat fetal cortical precursor cells (rCPC) and rat pheochromocytoma PC12 cells. When rCPC were differentiated into neuronal cells for 7 days, early stem cell marker, nestin, expression was decreased from day 4, and neuronal markers such as neurofilament-L, -M and Tuj1 were increased after day 4. In this condition, expressions of BIP, ATF6, and phosphorylated PERK as well as their down stream signaling molecules such as CHOP, ATF4, XBP1, GADD34, Nrf2 and p58IPK were significantly increased, suggesting the induction of ERSR during neuronal differentiation of rCPC. ERSR was also induced during the differentiation of PC12 cells for 9 days with NGF. Neurofilament-L transcript was time-dependently increased. Both mRNA and protein levels of Tuj1 were increased after the induction, and the significant increase in NeuN was observed at day 9. Similar to the expression patterns of neuronal markers, BIP/GRP78 and CHOP mRNAs were highly increased at day 9, and ATF4 mRNA was also increased from day 7. These results strongly suggest the induction and possible role of ERSR in neuronal differentiation process. Further study to identify targets responsible for neuronal induction will be necessary.
Sujet(s)
Animaux , Rats , Stress du réticulum endoplasmique , Réticulum endoplasmique , Facteur de croissance nerveuse , Nestine , Neurones , Cellules PC12 , Phéochromocytome , Rivières , ARN messager , Cellules souchesRÉSUMÉ
We have previously isolated a novel protein "B/K" that contains two C2-like domains. Here, we report the isolatioin and mRNA distribution of a human B/K isoform, and protein kinase A (PKA)-dependent phosphorylation of the B/K protein. The 1.5 kb human B/K cDNA clone exhibits 89% and 97% identities with rat B/K in the sequences of nucleotide and amino acid, respectively. Human B/K isoform encodes a 474 amino acid protein and shows structural features similar to the rat counterpart including two C2 domains, three consensus sequences for PKA, absence of a transmembrane region, and conservation of the N-terminal cysteine cluster. On Northern and dot blot analyses, a 3.0 kb B/K transcript was abundantly present in human brain, kidney, and prostate. Among the brain regions, strong signals were observed in the frontal and temporal lobes, the hippocampus, the hypothalamus, the amygdala, the substantia nigra, and the pituitary. Recombinant B/K proteins containing three consensus sites for PKA was very efficiently phosphorylated in vitro by PKA catalytic subunit. B/K protein which was overexpressed in LLC-PK1 cells was also strongly phosphorylated in vivo by vasopressin analog DDAVP, and PKA-specific inhibitor H89 as well as type 2 vasopressin receptor antagonist specifically suppressed DDAVP-induced B/K phosphorylation. These results suggest that B/K proteins play a role as potential substrates for PKA in the area where they are expressed.
Sujet(s)
Rats , Souris , Mâle , Humains , Femelle , Animaux , Adulte , Similitude de séquences d'acides aminés , Analyse de séquence d'ADN , Isoformes de protéines/génétique , Phosphorylation , Phosphoprotéines/génétique , Données de séquences moléculaires , Analyse de profil d'expression de gènes , ADN complémentaire/composition chimique , Cyclic AMP-Dependent Protein Kinases/physiologie , Clonage moléculaire , Lignée cellulaire , Séquence nucléotidique , Séquence d'acides aminésRÉSUMÉ
BACKGROUND: Nerve growth factor (NGF) promotes the survival and differentiation of vertebrate neurons, and their actions are mediated by two classes of cell surface receptors: tyrosine kinase A receptor (TrkA) and p75 neurotrophic receptor (p75NTR). We evaluated the role of NGF receptors in neuronal survival and the physical interactions between them. METHODS: Organotypic hippocampal slices were obtained from 5 to 7-day-old rat pups and were grown for 14 days in vitro. The expression of the TrkA and p75NTR was evaluated by the western blot and immunohistochemical methods. The neuroprotective effect of NGF on the blocking of antibody-induced neuronal cell death was tested by the application of NGF (0, 50 and 150 ng/ml) to the culture media in the presence of 200 ng/ml of blocking antibodies against TrkA and p75NTR. Functional interactions between the two receptors were examined using the immunoprecipitation method. RESULTS: TrkA and p75NTR were co-expressed in the principal neurons of the hippocampal slice culture, and the expression level was increased time dependently until 14 days of culture. The blocking antibody against each receptor induced neuronal damage in time and dose-dependent manners. NFG delayed or prevented the blocking antibody from inducing neuronal damage. Results from the immunoprecipitation experiment showed physical interactions between the two NGF receptors. CONCLUSIONS: Our results indicate that the co-expressed NGF receptors, TrkA and p75NTR, might have protective roles in the survival of neuronal cells through the cooperative interactions between them.
Sujet(s)
Animaux , Rats , Anticorps bloquants , Technique de Western , Mort cellulaire , Milieux de culture , Immunoprécipitation , Facteur de croissance nerveuse , Neurones , Neuroprotecteurs , Protein-tyrosine kinases , Récepteur facteur croissance nerf , Récepteurs de surface cellulaire , Récepteurs facteur croissance nerf , VertébrésRÉSUMÉ
Nursing the aged is yet a familiar system in Korea. However, it is a social system in urgent need that requires series of preliminary arrangements to cope with the aging society. In addition, a full-fledged preparedness is imperative, lest the nursing system should follow precedent failures of the National Medical Insurance System and the separation of dispensary from medical practice. The concept of nursing the aged exceeds the simple care (so called 'Subal') of inconveniences caused by the process of aging. One should keep in mind that 'nursing the aged' outcomes from the illness as well as the aftereffect thereof. These physical and mental inconveniences should be overcome by means of medical treatments and care that are oriented to proper physical conditions. All sectors of specialties relevant to elderly nursing care should be systematically working together, so as to set up a sound master plan for nursing the aged. Both medical and para-medical human resources shall be secured and, they must be ready to work in good harmony. Medical approaches as to the home-cares and, construction of the infrastructures relevant thereto shall be properly taken place during the pre-testing period.
Sujet(s)
Sujet âgé , Humains , Vieillissement , Services de soins à domicile , Assurance , Corée , Soins de longue durée , Soins , Soins infirmiersRÉSUMÉ
B/K protein is a novel protein containing double C2-like domains. We examined the specific signaling pathway that regulates the transcription of B/K in PC12 cells. When the cells were treated with forskolin (50microM), B/K mRNA and protein levels were time-dependently decreased, reaching the lowest level at 3 or 4 hr, and thereafter returning to the control level. Chemicals such as dibutyryl-cAMP, cell- permeable cyclic AMP (cAMP) analogue and CGS21680, adenosine receptor A2A agonist, also repressed the B/K transcription. However, 1, 9-dideoxyforskolin did not show inhibitory effect on B/K transcription, suggesting direct involvement of cAMP in the forskolin-induced inhibition of B/K transcription. Effect of forskolin, dibutyryl cAMP and CGS21680 was significantly reduced in PKA-deficient PC12 cell line (PC12-123.7). One cAMP-response element (CRE) -like sequence (B/K CLS) was found in the promoter region of B/K DNA, and electrophoretic mobility shift assay indicated its binding to CREM and CREB. Forskolin significantly suppressed the promoter activity in CHO-K1 cells transfected with the constructs containing B/K CLS, but not with the construct in which B/K CLS was mutated (AC: TG). Taken together, we suggest that the transcription of B/K gene in PC12 cells may be regulated by PKA-dependent mechanism.
Sujet(s)
Animaux , Colforsine , AMP cyclique , Cyclic AMP-Dependent Protein Kinases , ADN , Test de retard de migration électrophorétique , Cellules PC12 , Régions promotrices (génétique) , Récepteurs purinergiques P1 , ARN messagerRÉSUMÉ
Liver cirrhosis is one of the major complications of hepatitis C virus (HCV) infection, but the mechanisms underlying HCV-related fibrogenesis are still not clear. Although the roles of HCV core protein remain poorly understood, it is supposed to play an important role in the regulation of cellular growth and hepatocarcinogenesis. The aim of this study was to examine the role of HCV core protein on the hepatic fibrogenesis. We established an in vitro co-culture system with primary hepatic stellate cell (HSC) isolated from rats, and a stable HepG2-HCV core cell line which had been transfected with HCV core gene. The expressions of fibrosis-related molecules transforming growth factor beta1 (TGF-beta1), transforming growth factor b receptor II (TGF beta RII), alpha-smooth muscle actin (alpha-SMA) and connective tissue growth factor (CTGF) were analyzed via histological or molecular methods. In addition, the expression levels of matrix metaloprotinase-2 (MMP-2) and collagen type I (Col I) from the co-cultured media were measured by zymogram and ELISA, respectively. The expressions of alpha-SMA, TGF-beta1, Col I, TGF beta RII and MMP-2 were significantly increased in the co-culture of stable HepG2-HCV core with HSC. Moreover, the significant increases of CTGF and TGF-beta1 in the HCV core-expressing cells were observed by either Northern or Western blot analysis. These results suggest that HCV core protein may contribute to the hepatic fibrogenesis via up-regulation of CTGF and TGF-beta1.
Sujet(s)
Animaux , Mâle , Rats , Actines/métabolisme , Lignée cellulaire tumorale , Cellules cultivées , Techniques de coculture , Collagène de type I/métabolisme , Matrix metalloproteinase 2/métabolisme , Protéines précoces immédiates/biosynthèse , Protéines et peptides de signalisation intercellulaire/biosynthèse , Foie/métabolisme , Cirrhose du foie/métabolisme , Rat Sprague-Dawley , Récepteurs TGF-bêta/métabolisme , Facteur de croissance transformant bêta/métabolisme , Régulation positive , Protéines du core viral/génétiqueRÉSUMÉ
BACKGROUND: Mesenchymal stem cells (MSC) can be defined by their extensive in vitro self renewal capacity and multi-lineage differentiation potentiality. These cells possess in vitro immunosuppressive properties that appear not to be major histocompatibility complex (MHC) restricted. This study evaluated the immune suppressive effect of mouse MSC on mixed lymphocyte reaction (MLR), and the mechanisms were investigated. METHODS: MSC were obtained from BALB/c bone marrow and cultured in low-glucose DMEM media. The expression of surface antigens and cell cycle were analyzed by flow cytometry. The MSC-induced suppression was assessed by MLR and transwell culture. RESULTS: The BALB/c MSC constitutively expressed MHC class I and CD54 (ICAM-1) antigens but were negative for MHC class II, CD40, CD80 (B7-1) and CD106 (VCAM-1) antigens. MSC suppressed allogeneic C57BL/6 T lymphocytes proliferation by adding them to MLR in which C3H spleen cells were used as a stimulator. This inhibition was dependent on the dose of BALB/c MSC but independent of MHC. C57BL/6 T lymphocytes proliferation was still inhibited when BALB/c MSC were added in culture 3 days after starting of MLR. When MSC were separated from C57BL/6 T cells by using the transwell membrane, the suppression of immune response wasn't observed, which suggested that the suppressive effect was dependent on cell-cell contact between BALB/c MSC and C57BL/6 T cells. When C57BL/6 T lymphocytes were cultured with MSC, the percentage of C57BL/6 T cells in G0 phase increased from 51.8+/-7.66% to 77.2+/-7.39% compared with the case that only C57BL/6 T cells were cultured. When the C57BL/6 T cells were cultured with C3H spleen cells, most of C57BL/6 T cells were in G2/M (96.38+/-3.33%). But by the addition of MSC to MLR, the percentage of T cells in G2/M decreased to 33.0+/-9.66% while that of T cells in G0 increased to 66.2+/-7.46%. CONCLUSIONS: We concluded that the cell cycle of responder T lymphocytes in MLR is arrested at G0 phase by MSC.
Sujet(s)
Animaux , Souris , Antigènes de surface , Moelle osseuse , Cycle cellulaire , Cytométrie en flux , Phase G0 , Test de culture lymphocytaire mixte , Lymphocytes , Complexe majeur d'histocompatibilité , Membranes , Cellules souches mésenchymateuses , Rate , Lymphocytes TRÉSUMÉ
delta12-Prostaglandin (PG) J2 is known to elicit an anti-neoplastic effects via apoptosis induction. Previous study showed delta12-PGJ2-induced apoptosis utilized caspase cascade through cytochrome c-dependent pathways in HeLa cells. In this study, the cellular mechanism of delta12-PGJ2- induced apoptosis in HeLa cells, specifically, the role of two mitochondrial factors; bcl-2 and apoptosis-inducing factor (AIF) was investigated. Bcl-2 attenuated delta12-PGJ2-induced caspase activation, loss of mitochondrial transmembrane potential (delta psi m), nuclear fragmentation, DNA laddering, and growth curve inhibition for approximately 24 h, but not for longer time. AIF was not released from mitochondria, even if the delta psi m was dissipated. One of the earliest events observed in delta12-PGJ2-induced apoptotic events was dissipation of delta psi m, the process known to be inhibited by bcl-2. Pre-treatment of z-VAD- fmk, the pan-caspase inhibitor, resulted in the attenuation of delta psi m depolarization in delta12-PGJ2- induced apoptosis. Up-regulation of Sox-4 protein by delta12-PGJ2 was observed in HeLa and bcl-2 overexpressing HeLa B4 cell lines. Bcl-2 overexpression did not attenuate the expression of Sox-4 and its expression coincided with other apoptotic events. These results suggest that delta12-PGJ2 induced Sox-4 expression may activate another upstream caspases excluding the caspase 9-caspase 3 cascade of mitochondrial pathway. These and previous findings together suggest that delta12-PGJ2-induced apoptosis in HeLa cells is caspase-dependent, AIF-independent events which may be affected by Sox-4 protein expression up-regulated by delta12-PGJ2.
Sujet(s)
Femelle , Humains , Chlorométhyl cétones d'acides aminés/pharmacologie , Antinéoplasiques/pharmacologie , Apoptose/effets des médicaments et des substances chimiques , Caspases/physiologie , Cytochromes c/physiologie , Flavoprotéines/métabolisme , Cellules HeLa , Protéines HMG/physiologie , Protéines membranaires/métabolisme , Mitochondries/métabolisme , Prostaglandine D2/pharmacologie , Transport des protéines/physiologie , Protéines proto-oncogènes c-bcl-2/biosynthèse , Activation de la transcription , Transactivateurs/physiologieRÉSUMÉ
OBJECTIVE: Serotonin is a neurotransmitter which might play an important role in the pathophysiology of schizophrenia. As atypical antipsychotics, which antagonize serotonin receptors, exert effects on negative symptoms of schizophrenia, perturbation of serotonin system has been thought as a good indicator to probe the pathogenesis of schizophrenia. Serotonin transporter is crucial for regulating serotonergic neurotransmission and has a deletion/insertion polymorphism of the gene's transcriptional initiation site. This study was designed to examine the association of schizophrenia and polymorphism of serotonin transporter gene (5-HTTLPR). METHODS: Genomic DNA analysis with polymerase chain reaction (PCR) was used for genetic typing of polymorphism of 5-HTTLPR. It was carried out among a total of 111 patients with schizophrenia and 208 normal controls. The significances of genetic association of the polymorphisms with the disease and with clinical variables were estimated by chi-square test and analysis of variances. RESULTS: The results were as follows: 1) There were no significant differences in allelic or genotype frequencies between the group of patients with schizophrenia and controls. 2) There were no significant differences in positive syndrome scale score of positive and negative syndrome scale (PANSS), duration of illness and number of admission according to 5-HTTLPR genotypes. 3) There was a difference in age at onset according to 5-HTTLPR genotypes. 4) There was a significant difference in negative syndrome scale score and general psychopathology score of PANSS according to 5-HTTLPR genotypes. CONCLUSION: These results suggest that polymorphism of serotonin transporter gene might be related to the pathophysiology of schizophrenia.
Sujet(s)
Humains , Neuroleptiques , ADN , Génotype , Agents neuromédiateurs , Réaction de polymérisation en chaîne , Psychopathologie , Récepteurs sérotoninergiques , Schizophrénie , Transporteurs de la sérotonine , Sérotonine , Transmission synaptiqueRÉSUMÉ
OBJECTIVES: Contemporary understanding of schizophrenia has evolved over the last century, yet its pathogenesis is not clear. Environmental stresses in early gestational period, which in turn, can cause neurodevelopmental abnormalities, is one possible etiologic factors in the development of schizophrenia. Heat shock protein 70(HSP70), which is thought to be a protective factor against environmental stresses in a cell, might be involved in the development of schizophrenia. Abnormal immunoreactivity to HSP70 has been identified in patients with schizophrenia. Therefore, genes for HSP70 might be candidates that affect susceptibility to schizophrenia. Three genes encoding HSP70 such as HSP70-1, HSP70-hom and HSP70-2 have been identified in the MHC class III region and they have been known to have genetic polymorphisms. We examined the association of schizophrenia and polymorphisms of HSP70-1, HSP70-hom and HSP70-2 genes in this study. METHODS: We investigated 161 patients with schizophrenia and 165 controls. DNA analysis with polymerase chain reaction(PCR) followed by enzyme restriction was used for the allelic typing of each polymorphism of HSP70-1, HSP70-hom and HSP70-2. The significances of genetic association of the polymorphisms with the disease and with clinical variables were estimated by chi-square test and analysis of variances. RESULTS: 1) There were no significant differences in allelic or genotype frequencies of HSP70-1 and HSP70-hom between the group of patients with schizophrenia and controls. 2) There was a tendency of difference in genotype frequency of HSP70-2, and a significant difference in allelic frequency of HSP70-2 between the group of patients with schizophrenia and controls. 3) There were no significant differences in terms of severity of symptoms and age at onset among the three HSP70 genotypes in the group of patients with schizophrenia. CONCLUSION: These results suggest that polymorphism of HSP70-2 might be related to the pathogenesis of schizophrenia.
Sujet(s)
Humains , ADN , Génotype , Protéines du choc thermique , Température élevée , Protéines du choc thermique HSP70 , Polymorphisme génétique , SchizophrénieRÉSUMÉ
The completion of the rough draft of the human genome is a remarkable achievement. It provides the overall structures of huge DNA molecules that constitute the genome and an outline of the information needed to create a human being This paper reviewed new ideas, projects, and scientific advances made by the Human Genome Project. We also discussed the future of medicine and biomedical research in postgenomic era.
Sujet(s)
Humains , Humains , ADN , Génome , Génome humain , Génomique , Projet génome humain , Protéome , ProtéomiqueRÉSUMÉ
Over the past years, university administrators have known how hard it is to transform into the modern university. Rigid in-bred research system, narrow interest, unworkable graduate programs are complicatedly woven into a network of academic fraction. Cronyism and protectionism flood various laboratories and research institutes affiliated with the university. Until recently, the department structure of medical school has steadfastly guarded its territory and refused to allow non-medical undergraduate students to apply for the graduate schools of medical science. The graduate schools in medical science are considered just extra appendages because most of graduate students should be engaged in hard work position such as junior faculty or residentship training course of university hospital. In the present environment of graduate program, medical schools are consequently not able to bring in full-time young researchers, but only recently has its door been open for others. It should be time to reorganize the medical school graduate course into large multidisciplinary research group by expanding graduate programs.