RESUMO
Visualizing gradual changes in neuromelanin distribution within the substantia nigra is an important metric used to monitor the progression of Parkinsonism. This study aimed to identify the origin of the mismatch region between magnetic resonance transverse relaxation times (T2 and T2*) in the substantia nigra and investigate its feasibility and implications for in vivo detection of neuromelanin as a clinical biomarker. The relationships between neuromelanin distribution assessed by histological staining and the area of T2 and T2* mismatch determined by high- and low-resolution magnetic resonance relaxometry at 7T were directly compared in two normal and one depigmented substantia nigra collected at postmortem. In vivo feasibility of assessing T2 and T2* mismatch, clinically, was investigated using 3T magnetic resonance imaging. In the normal postmortem substantia nigra tissue, the T2 and T2* mismatch region exhibiting a linear pattern was strongly colocalized with neuromelanin distribution along the dorsal substantia nigra pars compacta, but a negligible amount of dorsal mismatch was observed in the depigmented brain. The regions of T2 and T2* mismatch from MRI, neuromelanin pigments from histology, and elevated iron signals from mass spectrometry were spatially overlapped for a normal postmortem brain. In preliminary in vivo studies, a similar, linear T2 and T2* mismatch region was observed in the dorsal area of the substantia nigra in eight normal subjects; this mismatch was significantly obscured in eight Parkinson's disease patients. The length of the dorsal linear mismatch line based on the T2*-T2 mask was significantly shorter in the Parkinson's disease patients compared to normal controls; this result was corroborated by reduced striatal uptake of [18F] FP-CIT dopamine transporters assessed by positron emission tomography scans. In conclusion, the measurement of T2 and T2* mismatch could serve as a complementary imaging biomarker to visualize the dorsal region of the substantia nigra pars compacta, which contains large amounts of neuromelanin.
Assuntos
Progressão da Doença , Imageamento por Ressonância Magnética/métodos , Melaninas , Neuroimagem/métodos , Doença de Parkinson/diagnóstico por imagem , Parte Compacta da Substância Negra/diagnóstico por imagem , Idoso , Idoso de 80 Anos ou mais , Biomarcadores , Diagnóstico , Estudos de Viabilidade , Feminino , Humanos , Melaninas/metabolismo , Doença de Parkinson/metabolismo , Doença de Parkinson/patologia , Parte Compacta da Substância Negra/metabolismo , Parte Compacta da Substância Negra/patologiaRESUMO
BACKGROUND: Providing feedback on student performance in the clinical performance assessment (CPA) is meaningful in that it helps students understand their strengths and weaknesses. This study compared students' perception of their CPA scores before and after providing personalized video feedback. METHODS: Two identical online surveys of Year 1 medical students (N = 103) that had undergone CPA were conducted to evaluate students' perceptions about their CPA scores before and after video feedback. Students were given their test scores with assessment analysis reports immediately after completing the CPA. Top-scored students from each station agreed to provide their video-recorded performance to the rest of the students. RESULTS: After comparing their performance video and top-scored video at each station, medical students were more aware of their CPA total score, clinical performance examination (CPX) total score, score of each CPX station, section score for the CPX station, history taking section score, physical examination section score, and doctor-patient relationship section score. Moreover, students became more convinced of their own weaknesses from their history taking and patient education section after viewing video feedback than before. CONCLUSION: The use of the video feedback system might help students recognize their CPA results and identify their strengths and weaknesses.
Assuntos
Competência Clínica/normas , Educação de Graduação em Medicina , Avaliação Educacional/métodos , Estudantes de Medicina/psicologia , Gravação em Vídeo , Atitude do Pessoal de Saúde , Retroalimentação , Humanos , Autoavaliação (Psicologia)RESUMO
Neuromelanin (NM) is an endogenous iron chelating molecule of pigmented neurons in the human substantia nigra (SN). Along with the increase in iron deposition, the reduction in NM-containing dopaminergic neurons and the variation of iron load on NM are generally considered to be important factors participating to pathogenesis of Parkinson's disease (PD). The aim of this study was to non-invasively delineate the spatial distributions of paramagnetic magnetic susceptibility perturbers, such as NM-iron complex and ferric iron in SN. Multiple quantitative MR parameters of T1, T2, T2*, susceptibility weighted image (SWI), quantitative susceptibility map (QSM), and T1 weighted image with magnetization transfer (MT) effects were acquired for six post-mortem SN samples without a history of neurological disease. Co-registered quantitative histological validations were performed to identify and correlate NM pigments, iron deposits, and myelin distributions with respect to associated MR parameters. The regions with NM pigments and iron deposits showed positive magnetic susceptibility (paramagnetic) values, while myelinated areas showed negative magnetic susceptibility (diamagnetic) values from the QSM. The region of reduced T2 values in SN mostly coincided with high iron deposits, but not necessarily with the NM pigments. The correlations between T2*/T2 (or T2*/T22) values and NM pigments were higher than those between T2* values and NM pigments, due to the effective size differences between NM-iron complex and ferric iron. Consequently, separate segmentations of ferric iron from the T2 map and NM-iron complex from the T2*/T2 map (or T2*/T22 map) were possible with the boundary of the SN determined from the T1 weighted image.
Assuntos
Ferro/análise , Imageamento por Ressonância Magnética/métodos , Melaninas/análise , Substância Negra/química , Substância Negra/diagnóstico por imagem , Adulto , Idoso , Idoso de 80 Anos ou mais , Autopsia , Feminino , Humanos , Processamento de Imagem Assistida por Computador , Masculino , Pessoa de Meia-IdadeRESUMO
BACKGROUND: Task-specific checklists, holistic rubrics, and analytic rubrics are often used for performance assessments. We examined what factors evaluators consider important in holistic scoring of clinical performance assessment, and compared the usefulness of applying holistic and analytic rubrics respectively, and analytic rubrics in addition to task-specific checklists based on traditional standards. METHODS: We compared the usefulness of a holistic rubric versus an analytic rubric in effectively measuring the clinical skill performances of 126 third-year medical students who participated in a clinical performance assessment conducted by Pusan National University School of Medicine. We conducted a questionnaire survey of 37 evaluators who used all three evaluation methods-holistic rubric, analytic rubric, and task-specific checklist-for each student. The relationship between the scores on the three evaluation methods was analyzed using Pearson's correlation. Inter-rater agreement was analyzed by Kappa index. The effect of holistic and analytic rubric scores on the task-specific checklist score was analyzed using multiple regression analysis. RESULTS: Evaluators perceived accuracy and proficiency to be major factors in objective structured clinical examinations evaluation, and history taking and physical examination to be major factors in clinical performance examinations evaluation. Holistic rubric scores were highly related to the scores of the task-specific checklist and analytic rubric. Relatively low agreement was found in clinical performance examinations compared to objective structured clinical examinations. Meanwhile, the holistic and analytic rubric scores explained 59.1% of the task-specific checklist score in objective structured clinical examinations and 51.6% in clinical performance examinations. CONCLUSION: The results show the usefulness of holistic and analytic rubrics in clinical performance assessment, which can be used in conjunction with task-specific checklists for more efficient evaluation.
Assuntos
Competência Clínica , Avaliação Educacional/métodos , Anamnese , Exame Físico , Estudantes de Medicina , Lista de Checagem , Humanos , Reprodutibilidade dos Testes , Inquéritos e QuestionáriosRESUMO
IL-7 signaling via IL-7Rα and common γ-chain (γc) is necessary for the development and homeostasis of T cells. Although the delicate mechanism in which IL-7Rα downregulation allows the homeostasis of T cell with limited IL-7 has been well known, the exact mechanism behind the interaction between IL-7Rα and γc in the absence or presence of IL-7 remains unclear. Additionally, we are still uncertain as to how only IL-7Rα is separately downregulated by the binding of IL-7 from the IL-7Rα/γc complex. We demonstrate here that 4G3, TUGm2, and 3E12 epitope masking of γc protein are induced in the presence of IL-7, indicating that the epitope alteration is induced by IL-7 binding to the preassembled receptor core. Moreover, the epitope masking of γc protein is inversely correlated with the expression of IL-7Rα upon IL-7 binding, implying that the structural alteration of γc might be involved in the regulation of IL-7Rα expression. The conformational change in γc upon IL-7 binding may contribute not only to forming the functional IL-7 signaling complex but also to optimally regulating the expression of IL-7Rα.
Assuntos
Epitopos/química , Subunidade gama Comum de Receptores de Interleucina/química , Interleucina-7/metabolismo , Receptores de Interleucina-7/química , Animais , Anticorpos Monoclonais/química , Citocinas/metabolismo , Humanos , Cinética , Camundongos , Camundongos Endogâmicos C57BL , Fosforilação , Ligação Proteica , Domínios Proteicos , Transdução de SinaisRESUMO
Unlike epidermal growth factor-like protein 7 (EGFL7), which is a secreted protein implicated in the regulation of blood vessel formation and cell migration, little is known about the physiological function of EGFL8. Thymic epithelial cells (TECs) play a pivotal role in T-cell development by regulating cellular interactions and expression of growth factors, cytokines, and chemokines. In order to investigate the functional role of EGFL8 in TECs, we transfected TECs with an EGFL8-expressing vector to overexpress EGFL8 protein and with an EGFL8 siRNA to knockdown EGFL8 expression. EGFL8-silenced TECs showed significant increase in the number of adherent thymocytes by enhancing the expression of intercellular adhesion molecule-1 (ICAM-1), while the overexpression of EGFL8 inhibited the adherence of TECs to thymocytes by suppressing ICAM-1 expression. Furthermore, in vitro co-culture study revealed that knockdown of EGFL8 facilitated the maturation of thymocytes to CD4(+) and CD8(+) single-positive populations. These regulatory effects of EGFL8 in T-cell development were further confirmed by the results that knockdown of EGFL8 enhanced the expression of genes involved in thymopoiesis, such as interleukin-7 (IL-7), granulocyte/macrophage-colony stimulating factor (GM-CSF), and thymus-expressed chemokine (TECK). Our data show that EGFL8 exerts inhibitory effects on TECs and thymocytes, suggesting that EGFL8 acts as a negative regulatory molecule in the development of T cells in the mouse thymus.
Assuntos
Proteínas/fisiologia , Linfócitos T/citologia , Timo/citologia , Animais , Linfócitos T CD4-Positivos/citologia , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD8-Positivos/citologia , Linfócitos T CD8-Positivos/metabolismo , Proteínas de Ligação ao Cálcio , Adesão Celular , Linhagem Celular , Família de Proteínas EGF , Células Epiteliais/citologia , Células Epiteliais/metabolismo , Técnicas de Silenciamento de Genes , Camundongos , Camundongos Endogâmicos C57BL , Proteínas/genética , Linfócitos T/metabolismoRESUMO
Cancer stem cells have been hypothesized to drive the growth and metastasis of tumors. Because they need to be targeted for cancer treatment, they have been isolated from many solid cancers. However, cancer stem cells from primary human gastric cancer tissues have not been isolated as yet. For the isolation, we used two cell surface markers: the epithelial cell adhesion molecule (EpCAM) and CD44. When analyzed by flow cytometry, the EpCAM(+)/CD44(+) population accounts for 4.5% of tumor cells. EpCAM(+)/CD44(+) gastric cancer cells formed tumors in immunocompromised mice; however, EpCAM(-)/CD44(-), EpCAM(+)/CD44(-) and EpCAM(-)/CD44(+) cells failed to do so. Xenografts of EpCAM(+)/CD44(+) gastric cancer cells maintained a differentiated phenotype and reproduced the morphological and phenotypical heterogeneity of the original gastric tumor tissues. The tumorigenic subpopulation was serially passaged for several generations without significant phenotypic alterations. Moreover, EpCAM(+)/CD44(+), but not EpCAM(-)/CD44(-), EpCAM(+)/CD44(-) or EpCAM(-)/CD44(+) cells grew exponentially in vitro as cancer spheres in serum-free medium, maintaining the tumorigenicity. Interestingly, a single cancer stem cell generated a cancer sphere that contained various differentiated cells, supporting multi-potency and self-renewal of a cancer stem cell. EpCAM(+)/CD44(+) cells had greater resistance to anti-cancer drugs than other subpopulation cells. The above in vivo and in vitro results suggest that cancer stem cells, which are enriched in the EpCAM(+)/CD44(+) subpopulation of gastric cancer cells, provide an ideal model system for cancer stem cell research.
Assuntos
Modelos Biológicos , Células-Tronco Neoplásicas/metabolismo , Neoplasias Gástricas/metabolismo , Adulto , Idoso , Animais , Antígenos de Neoplasias/metabolismo , Antineoplásicos/farmacologia , Biomarcadores Tumorais/metabolismo , Moléculas de Adesão Celular/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Molécula de Adesão da Célula Epitelial , Feminino , Humanos , Receptores de Hialuronatos/metabolismo , Masculino , Camundongos , Camundongos Nus , Pessoa de Meia-Idade , Células-Tronco Neoplásicas/citologia , Fenótipo , Pesquisa com Células-Tronco , Neoplasias Gástricas/patologia , Transplante HeterólogoRESUMO
BACKGROUND: Questioning is one of the essential techniques used by lecturers to make lectures more interactive and effective. This study surveyed the perception of questioning techniques by medical school faculty members and analyzed how the questioning technique is used in actual classes. METHODS: Data on the perceptions of the questioning skills used during lectures was collected using a self-questionnaire for faculty members (N = 33) during the second semester of 2008. The questionnaire consisted of 18 items covering the awareness and characteristics of questioning skills. Recorded video tapes were used to observe the faculty members' questioning skills. RESULTS: Most faculty members regarded the questioning technique during classes as being important and expected positive outcomes in terms of the students' participation in class, concentration in class and understanding of the class contents. In the 99 classes analyzed, the median number of questions per class was 1 (0-29). Among them, 40 classes (40.4 %) did not use questioning techniques. The frequency of questioning per lecture was similar regardless of the faculty members' perception. On the other hand, the faculty members perceived that their usual wait time after question was approximately 10 seconds compared to only 2.5 seconds measured from video analysis. More lecture-experienced faculty members tended to ask more questions in class. CONCLUSIONS: There were some discrepancies regarding the questioning technique between the faculty members' perceptions and reality, even though they had positive opinions of the technique. The questioning skills during a lecture need to be emphasized to faculty members.
Assuntos
Educação Médica/métodos , Docentes de Medicina , Humanos , Masculino , Pessoa de Meia-Idade , Faculdades de Medicina , Inquéritos e Questionários , Ensino/métodosRESUMO
Glioblastoma is the most common type of astrocytoma in the brain. Due to its high invasiveness and chemoresistance, patients with advanced stage of glioblastoma have a poor prognosis. SNAI1, an important regulator of epithelial-mesenchymal transition, has been associated with metastasis in various carcinoma cells. However, its roles in glioblastoma cells have been poorly characterized. To examine roles of SNAI1 in glioblastoma cells, we knockdowned SNAI1 expression using siRNA. SNAI1 siRNA increased the expression level of E-cadherin and decreased that of vimentin. In the water-soluble tetrazolium salt (WST-1) assay, SNAI1 siRNA inhibited the proliferation of U87-MG and GBM05 glioblastoma cells. Moreover, in the Boyden chamber assay and Matrigel invasion assay, SNAI1 siRNA inhibited serum-induced migration and invasion of glioblastoma cells. These results suggested that SNAI1 is involved in the proliferation and migration of glioblastoma cells.
Assuntos
Movimento Celular/fisiologia , Proliferação de Células , Glioblastoma/patologia , Glioblastoma/fisiopatologia , Fatores de Transcrição/metabolismo , Caderinas/metabolismo , Linhagem Celular Tumoral , Transição Epitelial-Mesenquimal , Humanos , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Fatores de Transcrição da Família Snail , Fatores de Transcrição/genética , Vimentina/metabolismoRESUMO
The ubiquitin-proteasome system (UPS), lysosomes, and autophagy are essential protein degradation systems for the regulation of a variety of cellular physiological events including the cellular response to injury. It has recently been reported that the UPS and autophagy mediate the axonal degeneration caused by traumatic insults and the retrieval of nerve growth factors. In the peripheral nerves, axonal degeneration after injury is accompanied by myelin degradation, which is tightly related to the reactive changes of Schwann cells called dedifferentiation. In this study, we examined the role of the UPS, lysosomal proteases, and autophagy in the early phase of Wallerian degeneration of injured peripheral nerves. We found that nerve injury induced an increase in the ubiquitin conjugation and lysosomal-associated membrane protein-1 expression within 1 day without any biochemical evidence for autophagy activation. Using an ex vivo explant culture of the sciatic nerve, we observed that inhibiting proteasomes or lysosomal serine proteases prevented myelin degradation, whereas this was not observed when inhibiting autophagy. Interestingly, proteasome inhibition, but not leupeptin, prevented Schwann cells from inducing dedifferentiation markers such as p75 nerve growth factor receptor and glial fibrillary acidic protein in vitro and in vivo. In addition, proteasome inhibitors induced cell cycle arrest and cellular process formation in cultured Schwann cells. Taken together, these findings indicate that the UPS plays a role in the phenotype changes of Schwann cells in response to nerve injury.
Assuntos
Desdiferenciação Celular/efeitos dos fármacos , Complexo de Endopeptidases do Proteassoma/metabolismo , Células de Schwann/metabolismo , Nervo Isquiático/metabolismo , Ubiquitinação/efeitos dos fármacos , Animais , Autofagia/efeitos dos fármacos , Autofagia/fisiologia , Axotomia , Western Blotting , Ciclo Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Inibidores de Cisteína Proteinase/farmacologia , Imunofluorescência , Processamento de Imagem Assistida por Computador , Leupeptinas/farmacologia , Proteína 1 de Membrana Associada ao Lisossomo/metabolismo , Lisossomos/efeitos dos fármacos , Lisossomos/metabolismo , Camundongos , Fibras Nervosas Mielinizadas/efeitos dos fármacos , Fibras Nervosas Mielinizadas/metabolismo , Células de Schwann/citologia , Células de Schwann/efeitos dos fármacos , Nervo Isquiático/citologia , Nervo Isquiático/efeitos dos fármacos , Nervo Isquiático/lesões , Ubiquitinação/fisiologia , Degeneração Walleriana/metabolismoRESUMO
BACKGROUND: Caffeine is the most commonly consumed psycho-stimulant in the world. The effects of caffeine on the body have been extensively studied; however, its effect on the structure of the brain has not been investigated to date. RESULTS: In the present study we found that the long-term consumption of caffeine can induce ventriculomegaly; this was observed in 40% of the study rats. In the caffeine-treated rats with ventriculomegaly, there was increased production of CSF, associated with the increased expression of Na(+), K(+)-ATPase and increased cerebral blood flow (CBF). In contrast to the chronic effects, acute treatment with caffeine decreased the production of CSF, suggesting 'effect inversion' associated with caffeine, which was mediated by increased expression of the A1 adenosine receptor, in the choroid plexus of rats chronically treated with caffeine. The involvement of the A1 adenosine receptor in the effect inversion of caffeine was further supported by the induction of ventriculomegaly and Na+, K+-ATPase, in A1 agonist-treated rats. CONCLUSION: The results of this study show that long-term consumption of caffeine can induce ventriculomegaly, which is mediated in part by increased production of CSF. Moreover, we also showed that adenosine receptor signaling can regulate the production of CSF by controlling the expression of Na(+), K(+)-ATPase and CBF.
Assuntos
Cafeína/farmacologia , Líquido Cefalorraquidiano/efeitos dos fármacos , Líquido Cefalorraquidiano/fisiologia , Animais , Western Blotting , Cafeína/administração & dosagem , Cafeína/sangue , Ventrículos Cerebrais/patologia , Circulação Cerebrovascular/efeitos dos fármacos , Circulação Cerebrovascular/fisiologia , Plexo Corióideo/metabolismo , Imuno-Histoquímica , Imageamento por Ressonância Magnética , Masculino , Ratos , Ratos Sprague-Dawley , Receptor A1 de Adenosina/metabolismo , ATPase Trocadora de Sódio-Potássio/metabolismo , Teofilina/sangueRESUMO
BACKGROUND: Hedgehog signaling plays critical roles during embryonic development. It is also involved in tissue regeneration and carcinogenesis in various adult tissues. Moreover, it regulates the maintenance of cancer stem cells and adult stem cells. Although hedgehog signaling is important in gastric carcinogenesis, its role in gastric regeneration has not been previously examined. In the present study, we evaluated the expression and roles of hedgehog signaling during gastric regeneration. METHODS: Gastric ulcers were induced by serosal application of an acetic acid solution in mice. Sham-operated mice served as controls. The proliferation of gastric progenitor cells was studied using bromodeoxyuridine (BrdU). The expression of hedgehog signaling molecules and the differentiation of gastric progenitor cells were examined by immunohistochemical staining and Western blotting. RESULTS: One day after the induction of gastric ulcer, the proliferation of gastric progenitor cells increased; however, the expression of hedgehog signaling molecules, including sonic hedgehog (Shh), Indian hedgehog (Ihh), desert hedgehog (Dhh), and patched (Ptch1) decreased at the ulcer margin. From 5 days after the induction of gastric ulcer, newly generated gastric glands and their differentiation were observed at the ulcer margin. The expression of hedgehog signaling molecules gradually increased in the newly generated gastric glands of the ulcer margin. Cyclopamine, a specific inhibitor of hedgehog signaling, significantly inhibited the differentiation of mucous cells and parietal cells during the gastric regeneration process. CONCLUSION: The above results suggest that hedgehog signaling is involved in the differentiation of gastric progenitor cells during the gastric ulcer repair process.
Assuntos
Mucosa Gástrica/metabolismo , Proteínas Hedgehog/metabolismo , Regeneração/fisiologia , Transdução de Sinais , Células-Tronco/metabolismo , Animais , Western Blotting , Diferenciação Celular , Proliferação de Células , Mucosa Gástrica/citologia , Mucosa Gástrica/fisiologia , Proteínas Hedgehog/antagonistas & inibidores , Imuno-Histoquímica , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Receptores Acoplados a Proteínas G/metabolismo , Receptor Smoothened , Células-Tronco/fisiologia , Úlcera Gástrica/metabolismo , Úlcera Gástrica/patologia , Úlcera Gástrica/fisiopatologia , Alcaloides de Veratrum/farmacologiaRESUMO
Ischemia-induced cerebral injury evolves over a longer period than previously believed through post-ischemic inflammation. Retinoic acid (RA) has been shown to exert cytoprotective effects on several cells, but its effects on ischemia-induced cerebral injury have been poorly characterized. The aim of the present study was to examine the effects of all-trans-RA on ischemia-induced cerebral injury and elucidate the underlying mechanism. All-trans-RA treatment reduced the size of the ischemia-induced cerebral infarct. To elucidate the underlying mechanism, ischemia-induced cerebral inflammation was studied by examination of expressions of interleukin 1beta (IL-1beta) and ED-1. RA treatment significantly reduced the cerebral inflammation. Moreover, cerebral ischemic induction of cyclooxygenase-2 (COX-2) and CCAAT/enhancer binding protein beta (C/EBPbeta), which binds to the COX-2 promoter, was also inhibited by RA. These results suggest that RA can reduce ischemia-induced cerebral injury by an anti-inflammatory action, which may be effected via inhibition of C/EBPbeta-mediated COX-2 induction.
Assuntos
Isquemia Encefálica/tratamento farmacológico , Encéfalo/efeitos dos fármacos , Tretinoína/uso terapêutico , Análise de Variância , Animais , Anti-Inflamatórios/uso terapêutico , Western Blotting , Encéfalo/metabolismo , Isquemia Encefálica/metabolismo , Proteína beta Intensificadora de Ligação a CCAAT/metabolismo , Ciclo-Oxigenase 2/metabolismo , Ectodisplasinas/metabolismo , Imuno-Histoquímica , Interleucina-1beta/metabolismo , Masculino , Microscopia Confocal , Neurônios/metabolismo , Fotomicrografia , Ratos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Gastric cancer is the second most common cause of cancer deaths worldwide. The underlying molecular mechanisms of its carcinogenesis are relatively poorly characterized. Hedgehog (Hh) signaling, which is critical for development of various organs including the gastrointestinal tract, has been associated with gastric cancer. The present study was undertaken to reveal the underlying mechanism by which Hh signaling controls gastric cancer cell proliferation. Treatment of gastric cancer cells with cyclopamine, a specific inhibitor of Hh signaling pathway, reduced proliferation and induced apoptosis of gastric cancer cells. Cyclopamine treatment induced cytochrome c release from mitochondria and cleavage of caspase 9. Moreover, Bcl-2 expression was significantly reduced by cyclopamine treatment. These results suggest that Hh signaling regulates the survival of gastric cancer cells by regulating the expression of Bcl-2.
Assuntos
Regulação Neoplásica da Expressão Gênica , Proteínas Hedgehog/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Neoplasias Gástricas/metabolismo , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Proteínas Hedgehog/genética , Humanos , Proteínas Proto-Oncogênicas c-bcl-2/genética , Transdução de Sinais/efeitos dos fármacos , Neoplasias Gástricas/genética , Alcaloides de Veratrum/farmacologiaRESUMO
Lymphoma is a heterogeneous disease with a highly variable clinical course and prognosis. Improving the prognosis for patients with relapsed and treatment-resistant lymphoma remains challenging. Current in vitro drug testing models based on 2D cell culture lack natural tissue-like structural organization and result in disappointing clinical outcomes. The development of efficient drug testing models using 3D cell culture that more accurately reflects in vivo behaviors is vital. Our aim was to establish an in vitro 3D lymphoma model that can imitate the in vivo 3D lymphoma microenvironment. Using this model, we explored strategies to enhance chemosensitivity to doxorubicin, an important chemotherapeutic drug widely used for the treatment of hematological malignancies. Lymphoma cells grown in this model exhibited excellent biomimetic properties compared to conventional 2D culture including (1) enhanced chemotherapy resistance, (2) suppressed rate of apoptosis, (3) upregulated expression of drug resistance genes (MDR1, MRP1, BCRP and HIF-1α), (4) elevated levels of tumor aggressiveness factors including Notch (Notch-1, -2, -3, and -4) and its downstream molecules (Hes-1 and Hey-1), VEGF and MMPs (MMP-2 and MMP-9), and (5) enrichment of a lymphoma stem cell population. Tiam1, a potential biomarker of tumor progression, metastasis, and chemoresistance, was activated in our 3D lymphoma model. Remarkably, we identified two synergistic therapeutic oncotargets, Tiam1 and Notch, as a strategy to combat resistance against doxorubicin in EL4 T and A20 B lymphoma. Therefore, our data suggest that our 3D lymphoma model is a promising in vitro research platform for studying lymphoma biology and therapeutic approaches.
RESUMO
The marked induction of glial fibrillary acidic protein (GFAP) has been observed in astrocytes during neuropathological processes accompanying reactive gliosis; however, the precise molecular mechanism(s) underlying this GFAP induction remains poorly resolved. Therefore, in this study, we examined whether the change of nuclear factor-kappa B (NF-kappaB) activity can influence GFAP expression levels. Aspirin, widely used to prevent NF-kappaB activity, reduced the levels of GFAP mRNA and protein in human astroglial cells including human glioblastoma A172 cells and primary human brain astrocyte cells (HBAs). Furthermore, aspirin inhibited the effects of hypoxic injury on the up-regulation of GFAP expression in HBAs. We confirmed the repressive effect of aspirin on GFAP transcription by GFAP promoter-driven reporter assay and found that one NF-kappaB binding site conserved in the mouse and human GFAP gene promoters is critical for this effect. To further delineate whether NF-kappaB is directly involved in the regulation of GFAP gene expression, we transfected A172 cells with an expression vector encoding a super-repressor IkappaBalpha protein (IkappaBalpha-SR) to specifically inhibit NF-kappaB activity and found the marked reduction of GFAP protein levels in IkappaBalpha-SR-transfectant cells. Taken together, our results suggest that NF-kappaB may play pivotal roles in GFAP gene expression.
Assuntos
Aspirina/farmacologia , Astrócitos/efeitos dos fármacos , Astrócitos/metabolismo , Proteína Glial Fibrilar Ácida/genética , Proteína Glial Fibrilar Ácida/metabolismo , NF-kappa B/antagonistas & inibidores , Regulação para Cima/efeitos dos fármacos , Astrócitos/citologia , Hipóxia Celular/efeitos dos fármacos , Regulação para Baixo/genética , Humanos , Quinase I-kappa B/metabolismo , NF-kappa B/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Transcrição Gênica/efeitos dos fármacos , Células Tumorais CultivadasRESUMO
Neurogenesis can be induced by pathological conditions such as cerebral ischemia. However the molecular mechanisms or modulating reagents of the reactive neurogenesis after the cerebral ischemia are poorly characterized. Retinoic acid (RA) has been shown to increase neurogenesis by enhancing the proliferation and neuronal differentiation of forebrain neuroblasts. Here, we examined whether RA can modulate the reactive neurogenesis after the cerebral ischemia. In contrast to our expectation, RA treatment decreased the reactive neurogenesis in subventricular zone (SVZ), subgranular zone (SGZ) and penumbral region. Furthermore, RA treatment also decreased the angiogenesis and gliosis in penumbral region.
Assuntos
Ataque Isquêmico Transitório/patologia , Neurônios/fisiologia , Tretinoína/farmacologia , Animais , Encéfalo/irrigação sanguínea , Diferenciação Celular , Proliferação de Células , Ataque Isquêmico Transitório/metabolismo , Masculino , Neovascularização Patológica , Neuroglia/patologia , Neuroglia/fisiologia , Neurônios/patologia , Ratos , Ratos Sprague-Dawley , Tretinoína/fisiologiaRESUMO
Skin diseases associated with inflammation or oxidative stress represent the most common problem in dermatology. The present study demonstrates that fish scale collagen peptides (FSCP) protect against CoCl2-induced cytotoxicity and TNF-α-induced inflammatory responses in human HaCaT keratinocyte cells. Our study is the first to report that FSCP increase cell viability and ameliorate oxidative injury in HaCaT cells through mechanisms mediated by the downregulation of key proinflammatory cytokines, namely, TNF-α, IL-1ß, IL-8, and iNOS. FSCP also prevent cell apoptosis by repressing Bax expression, caspase-3 activity, and cytochrome c release and by upregulating Bcl-2 protein levels in CoCl2- or TNF-α-stimulated HaCaT cells. In addition, the inhibitory effects of FSCP on cytotoxicity and the induction of proinflammatory cytokine expression were found to be associated with suppression of the ROS, MAPK (p38/MAPK, ERK, and JNK), and NF-κB signaling pathways. Taken together, our data suggest that FSCP are useful as immunomodulatory agents in inflammatory or immune-mediated skin diseases. Furthermore, our results provide new insights into the potential therapeutic use of FSCP in the prevention and treatment of various oxidative- or inflammatory stress-related inflammation and injuries.
Assuntos
Colágeno/metabolismo , Inflamação/metabolismo , NF-kappa B/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Peptídeos/metabolismo , Pele/patologia , Fator de Necrose Tumoral alfa/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Humanos , Espécies Reativas de Oxigênio/metabolismoRESUMO
It has been proposed that susceptibility-weighted imaging is a sensitive magnetic resonance imaging (MRI) technique for identifying white matter (WM) pathologic changes involving demyelination and iron accumulation. We identified the tree silhouette-like configuration with a paramagnetic phase shift in the frontal subcortical WM lesions of 4 patients with adult-onset leukoencephalopathy with axonal spheroids and pigmented glia who underwent 3T MRI. According to our postmortem 7T MRI and histologic correlation study to investigate the origin of the susceptibility-related phase contrast, changes in the subcortical WM architecture and central WM loss with the relative preservation of iron-rich U-fibers may contribute to the paramagnetic susceptibility.
Assuntos
Axônios/patologia , Leucoencefalopatias/patologia , Neuroglia/patologia , Pigmentação , Esferoides Celulares/patologia , Substância Branca/patologia , Espectroscopia de Ressonância de Spin Eletrônica/métodos , Feminino , Humanos , Leucoencefalopatias/diagnóstico por imagem , Imageamento por Ressonância Magnética/métodos , Masculino , Pessoa de Meia-Idade , Substância Branca/diagnóstico por imagemRESUMO
Trichostatin A (TSA), histone deacetylase inhibitor, shows a promising therapeutic effect on cancer cells in combination with radiotherapy or chemotherapy. However, little has been reported on the combined treatment of TSA with hyperthermia. Here, we have assessed the effect of TSA/hyperthermia on human glioblastoma A172 cells and found that TSA increases the thermosensitivity of A172 cells, resulting in cellular apoptosis. The underlying mechanism of this effect consists of reduction in the level of phosphorylated STAT3 (Tyr705), a transcription factor required for survival of A172 cells, which leads to down-regulation of STAT3 target genes, cyclin D1 and Bcl-xL. Furthermore, the level of VEGF mRNA was also decreased by TSA/hyperthermia, suggesting the antiangiogenic effect of TSA/hyperthermia on human glioblastoma. Collectively, our results show the role of TSA as a chemical thermosensitizer, suggesting the possible therapeutic application of combined treatment of TSA/hyperthermia on STAT3-dependent tumors.