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OBJECTIVE: We attempted to investigate whether dehydroepiandrosterone sulfate (DHEA-S) levels are associated with remission of major depressive disorder by assessing scores on the 17-Item Structured Interview Guide for the Hamilton Depression before and after antidepressant treatment. METHODS: Plasma DHEA-S levels in 24 patients diagnosed with major depressive disorder on the basis of Diagnostic and Statistical Manual of Mental Disorders, fourth edition (text revision) before and after antidepressant treatment, and 24 healthy, gender-matched, and age-matched controls were measured using a radioimmunoassay kit. RESULTS: Plasma DHEA-S levels in patients were significantly higher than those in healthy controls. In patients who achieved remission after antidepressant treatment, plasma DHEA-S levels significantly declined compared with the levels before treatment. A significant correlation was observed between changes in DHEA-S levels and Absence of Depressive and Anxious Mood scores, which are calculated from the 2-Item Structured Interview Guide for the Hamilton Depression rating as follows: severity of depressive mood and anxiety in patients before and after antidepressant treatment. CONCLUSIONS: These findings suggest that plasma DHEA-S levels can be used as a putative indicator of the state of remission in patients with major depressive disorder. Copyright © 2014 John Wiley & Sons, Ltd.
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Antidepressivos/uso terapêutico , Sulfato de Desidroepiandrosterona/sangue , Transtorno Depressivo Maior/sangue , Transtorno Depressivo Maior/tratamento farmacológico , Adulto , Análise Química do Sangue/métodos , Transtorno Depressivo Maior/diagnóstico , Manual Diagnóstico e Estatístico de Transtornos Mentais , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Escalas de Graduação Psiquiátrica , Radioimunoensaio , Indução de Remissão , Resultado do Tratamento , Adulto JovemRESUMO
OBJECTIVES: Diallyl disulfide (DADS) is a natural organosulfur compound found in garlic and related plants with various pharmacologic effects. However, whether DADS improves obesity-induced insulin resistance (IR) and its underlying mechanisms remain unclear. The aim of this study was to investigate the effects of DADS on systemic IR in high-fat diet-induced obese mice. METHODS: To induce obesity, 8-wk-old male C57BL/6J mice were fed a high-fat diet (60% fat/kcal). The mice were assigned to three weight-matched groups: control (CON, n = 8), low-dose DADS (DADS-L, n = 8), and high-dose DADS (DADS-H, n = 9). The treated mice were orally administered DADS (25 or 100 mg/kg) 5 d/wk for 8 wk. At 15 wk of age, an intraperitoneal glucose tolerance test (GTT) and insulin tolerance test (ITT) were performed. Twenty-four hours after the final administration of DADS, epididymal fat and the liver were sampled after a 5-h fast. RESULTS: DADS administration significantly attenuated body and fat weight gains during the experimental period. Additionally, systemic IR, as evaluated by ITT, was significantly improved by DADS administration in a dose-dependent manner. High-dose DADS administration significantly decreased liver triacylglycerol levels. Moreover, high-dose DADS administration decreased the c-Jun N-terminal kinase (JNK) phosphorylation and significantly increased heat shock protein 72 expression in the liver. CONCLUSIONS: The results of this study suggested that DADS administration alleviated systemic IR in obese mice. This may be associated with decreased hepatic fat accumulation and a heat shock protein 72-mediated decrease in JNK activity in the liver.
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Compostos Alílicos , Resistência à Insulina , Camundongos , Animais , Dieta Hiperlipídica/efeitos adversos , Proteínas de Choque Térmico HSP72 , Camundongos Obesos , Camundongos Endogâmicos C57BL , Dissulfetos/farmacologia , Compostos Alílicos/farmacologia , Obesidade/tratamento farmacológico , Obesidade/metabolismoRESUMO
Semaphorins and their receptors have diverse functions in axon guidance, organogenesis, vascularization and/or angiogenesis, oncogenesis and regulation of immune responses. The primary receptors for semaphorins are members of the plexin family. In particular, plexin-A1, together with ligand-binding neuropilins, transduces repulsive axon guidance signals for soluble class III semaphorins, whereas plexin-A1 has multiple functions in chick cardiogenesis as a receptor for the transmembrane semaphorin, Sema6D, independent of neuropilins. Additionally, plexin-A1 has been implicated in dendritic cell function in the immune system. However, the role of plexin-A1 in vivo, and the mechanisms underlying its pleiotropic functions, remain unclear. Here, we generated plexin-A1-deficient (plexin-A1(-/-)) mice and identified its important roles, not only in immune responses, but also in bone homeostasis. Furthermore, we show that plexin-A1 associates with the triggering receptor expressed on myeloid cells-2 (Trem-2), linking semaphorin-signalling to the immuno-receptor tyrosine-based activation motif (ITAM)-bearing adaptor protein, DAP12. These findings reveal an unexpected role for plexin-A1 and present a novel signalling mechanism for exerting the pleiotropic functions of semaphorins.
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Proteínas Adaptadoras de Transdução de Sinal/fisiologia , Osso e Ossos/fisiologia , Imunidade , Proteínas do Tecido Nervoso/fisiologia , Receptores de Superfície Celular/fisiologia , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Homeostase , Glicoproteínas de Membrana/metabolismo , Camundongos , Camundongos Knockout , Proteínas do Tecido Nervoso/deficiência , Proteínas do Tecido Nervoso/metabolismo , Receptores de Superfície Celular/deficiência , Receptores de Superfície Celular/metabolismo , Receptores Imunológicos/metabolismo , Transdução de SinaisRESUMO
Methylmercury (MeHg), a global environmental pollutant, could seriously damage the central nervous system (CNS) and cause neurological disorders such as cerebellar symptoms. Although numerous studies have revealed detailed toxicity mechanisms of MeHg in neurons, toxicity in astrocytes is barely known. Here, we tried to shed light on the toxicity mechanisms of MeHg exposure in cultured normal rat cerebellar astrocytes (NRA), focusing on the involvement of reactive oxygen species (ROS) in MeHg toxicity by assessing the effects of major antioxidants Trolox, a free-radical scavenger, N-acetyl-L-cysteine (NAC), a potent thiol-containing antioxidant, and glutathione (GSH), an endogenous thiol-containing antioxidant. Exposure to MeHg at just approximately 2 µM for 96 h increased cell viability, which was accompanied by the increase in intracellular ROS level and at ≥ 5 µM induced significant cell death and lowered ROS level. Trolox and NAC suppressed 2 µM MeHg-induced increases in cell viability and ROS level corresponding to control, although GSH with 2 µM MeHg induced significant cell death and ROS increase. On the contrary, against 4 µM MeHg-induced cell loss and ROS decrease, NAC inhibited both cell loss and ROS decrease, Trolox inhibited cell loss and further enhanced ROS decrease, and GSH moderately inhibited cell loss and increased ROS level above the control level. MeHg-induced oxidative stress was suggested by increases in the protein expression levels of heme oxygenase-1 (HO-1), Hsp70, and Nrf2, except for the decrease in SOD-1 and no change in catalase. Furthermore, MeHg exposure dose-dependently induced increases in the phosphorylation of MAP kinases (ERK1/2, p38MAPK, and SAPK/JNK) and phosphorylation and/or expression levels of transcription factors (CREB, c-Jun, and c-Fos) in NRA. NAC successfully suppressed 2 µM MeHg-induced alterations in all of the above-mentioned MeHg-responsive factors, whereas Trolox suppressed some MeHg-responsive factors but failed to suppress MeHg-induced increases in the protein expression levels of HO-1 and Hsp70 and increase in p38MAPK phosphorylation. Protein expression analyses in NRA exposed to 2 µM MeHg and GSH were excluded because of devastating cell death. These results suggested that MeHg could induce aberrant NRA activation, and ROS must be substantially involved in the toxicity mechanism of MeHg in NRA; however, other factors should be assumed.
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Antioxidantes , Compostos de Metilmercúrio , Ratos , Animais , Espécies Reativas de Oxigênio/metabolismo , Antioxidantes/farmacologia , Antioxidantes/metabolismo , Sistema de Sinalização das MAP Quinases , Astrócitos , Estresse Oxidativo , Glutationa/metabolismo , Acetilcisteína/farmacologia , Acetilcisteína/metabolismo , Células CultivadasRESUMO
Diphenylarsinic acid (DPAA) is a non-natural pentavalent organic arsenic and was detected in well water in Kamisu, Ibaraki, Japan in 2003. Individuals that had consumed this arsenic-contaminated water developed cerebellar symptoms such as myoclonus. We previously revealed that DPAA exposure in rats in vitro and in vivo specifically affected astrocytes rather than neurons among cerebellar cells. Here, we evaluated adverse effects of DPAA in cultured normal human cerebellar astrocytes (NHA), which were compared with those in normal rat cerebellar astrocytes (NRA) exposed to DPAA at 10 µM for 96 h, focusing on aberrant activation of astrocytes; increase in cell viability, activation of MAP kinases (ERK1/2, p38MAPK, and SAPK/JNK) and transcription factors (CREB, c-Jun, and c-Fos), upregulation of oxidative stress-responsive factors (Nrf2, HO-1, and Hsp70), and also hypersecretion of brain cytokines (MCP-1, adrenomedullin, FGF-2, CXCL1, and IL-6) as reported in NRA. While DPAA exposure at 10 µM for 96 h had little effect on NHA, a higher concentration (50 µM for 96 h) and longer exposure (10 µM for 288 h) induced similar aberrant activation. Moreover, exposure to DPAA at 50 µM for 96 h or 10 µM for 288 h in NHA induced hypersecretion of cytokines induced in DPAA-exposed NRA (MCP-1, adrenomedullin, FGF-2, CXCL1, and IL-6), and IL-8 besides into culture medium. These results suggested that aberrantly activated human astrocytes by DPAA exposure might play a pivotal role in the pathogenesis of cerebellar symptoms, affecting adjacent neurons, microglia, brain blood vessels, or astrocyte itself through these brain cytokines in human.
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Arsenicais/efeitos adversos , Astrócitos/efeitos dos fármacos , Cerebelo/efeitos dos fármacos , Citocinas/metabolismo , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Fatores de Transcrição/metabolismo , Animais , Arsenicais/administração & dosagem , Astrócitos/metabolismo , Western Blotting , Cerebelo/citologia , Cerebelo/metabolismo , Relação Dose-Resposta a Droga , Ensaio de Imunoadsorção Enzimática , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Humanos , Ratos , Ratos WistarRESUMO
Chronic inflammation is considered as an etiology of obesity and type 2 diabetes. Brown adipose tissue (BAT) of obese animals shows increased inflammation. Regular exercise has anti-inflammatory effects; however, the effects of exercise training on BAT inflammation in obese animals remain unclear. Thus, this study aimed to investigate the effects of exercise training on inflammation-related signaling in the BAT of obese and diabetic rats. Male Otsuka Long-Evans Tokushima Fatty (OLETF) rats, an obese/diabetic rodent model, were randomly divided into either sedentary (n = 11) or exercise training (n = 8) groups. Long-Evans Tokushima Otsuka (LETO; n = 9) rats were used as the nondiabetic sedentary controls. Exercise training using a treadmill was conducted 4 days per week for 20 weeks, starting at 5 weeks old. As a result, exercise training attenuated the phosphorylation levels of p65 and mitogen-activated protein kinases in the BAT of OLETF rats, concurrently with the improvement of obesity and systemic glucose tolerance. Moreover, exercise training decreased oxidative stress and increased the antioxidant and anti-inflammatory protein levels in the BAT. Conversely, exercise training did not alter the expression levels of uncoupling protein-1 and oxidative phosphorylation-related proteins in the BAT, which were lower in the OLETF rats than the LETO rats. In conclusion, our data suggest that exercise training prevents the activation of inflammatory signaling in the BAT of obese/diabetic rats.
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PlexinA1 (PlxnA1) is a transmembrane receptor for semaphorins (Semas), a large family of axonal guidance cues vital during neural development. PlxnA1 is expressed in embryonic interneurons, and PlxnA1 deletion in mice leads to less interneurons in the developing cortex. In addition, PlxnA1 has been identified as a schizophrenia susceptibility gene. In our previous study, PlxnA1 knockout (KO) mice under a BALB/cAJ genetic background exhibited significantly increased self-grooming and reduced prepulse inhibition, a reliable phenotype for investigating the neurobiology of schizophrenia. However, the mechanism underlying the abnormal behavior of PlxnA1 KO mice remains unclear. We first confirmed PlxnA1 mRNA expression in parvalbumin-expressing interneurons (PV cells) in the medial prefrontal cortex (mPFC) of adult mice. Immunohistochemical analysis (IHC) showed significantly decreased densities of both GABAergic neurons and PV cells in the mPFC of PlxnA1 KO mice compared with wild type mice (WT). PV cells were found to express molecule interacting with CasL 1 (MICAL1), an effector involved in Sema-Plxn signaling for axon guidance, suggesting MICAL1 and PlxnA1 co-expression in PV cells. Furthermore, IHC analysis of 8-oxo-dG, an oxidative stress marker, revealed significantly increased oxidative stress in PlxnA1-deficient PV cells compared with WT. Thus, increased oxidative stress and decreased PV cell density in the mPFC may determine the onset of PlxnA1 KO mice's abnormal behavior. Accordingly, deficient PlxnA1-mediated signaling may increase oxidative stress in PV cells, thereby disrupting PV-cell networks in the mPFC and causing abnormal behavior related to neuropsychiatric diseases.
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This study investigated the combined effects of exercise training and D-allulose intake on endurance capacity in mice. Male C57BL/6J mice were fed either a control diet (Con) or a 3% D-allulose diet (Allu) and further divided into the sedentary (Sed) or exercise training (Ex) groups (Con-Sed, Con-Ex, Allu-Sed, Allu-Ex, respectively; n = 6-7/group). The mice in the Ex groups were trained on a motor-driven treadmill 5 days/week for 4 weeks (15-18 m/min, 60 min). After the exercise training period, all mice underwent an exhaustive running test to assess their endurance capacity. At 48 h after the running test, the mice in the Ex groups were subjected to run at 18 m/min for 60 min again. Then the gastrocnemius muscle and liver were sampled immediately after the exercise bout. The running time until exhaustion tended to be higher in the Allu-Ex than in the Con-Ex group (p = 0.08). The muscle glycogen content was significantly lower in the Con-Ex than in the Con-Sed group and was significantly higher in the Allu-Ex than in the Con-Ex group (p < 0.05). Moreover, exercise training increased the phosphorylation levels of adenosine monophosphate-activated protein kinase (AMPK) in the muscle and liver. The phosphorylation levels of acetyl coenzyme A carboxylase (ACC), a downstream of AMPK, in the muscle and liver were significantly higher in the Allu-Ex than in the Con-Sed group (p < 0.05), suggesting that the combination of exercise training and D-allulose might have activated the AMPK-ACC signaling pathway, which is associated with fatty acid oxidation in the muscle and liver. Taken together, our data suggested the combination of exercise training and D-allulose intake as an effective strategy to upregulate endurance capacity in mice. This may be associated with sparing glycogen content and enhancing activation of AMPK-ACC signaling in the skeletal muscle.
Assuntos
Proteínas Quinases Ativadas por AMP , Condicionamento Físico Animal , Proteínas Quinases Ativadas por AMP/metabolismo , Animais , Frutose/metabolismo , Glicogênio/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Músculo Esquelético/metabolismo , Condicionamento Físico Animal/fisiologiaRESUMO
PlexinA1 (PlxnA1) is a transmembrane receptor for semaphorins, a large family of proteins that act as axonal guidance cues during nervous system development. However, there are limited studies on PlxnA1 function in neurobehavior. The present study examined if PlxnA1 deficiency leads to behavioral abnormalities in BALB/cAJ mice. PlxnA1 knockout (KO) mice were generated by homologous recombination and compared to wild type (WT) littermates on a comprehensive battery of behavioral tests, including open field assessment of spontaneous ambulation, state anxiety, and grooming, home cage grooming, the wire hang test of muscle strength, motor coordination on the rotarod task, working memory on the Y maze alternation task, cued and contextual fear conditioning, anxiety on the elevated plus maze, sociability to intruders, and sensory processing as measured by prepulse inhibition (PPI). Measures of motor performance, working memory, fear memory, and sociability did not differ significantly between genotypes, while PlxnA1 KO mice displayed excessive self-grooming, impaired PPI, and slightly lower anxiety. These results suggest a crucial role for PlxnA1 in the development and function of brain regions controlling self-grooming and sensory gating. PlxnA1 KO mice may be a valuable model to investigate the repetitive behaviors and information processing deficits characteristic of many neurodevelopmental and psychiatric disorders.
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Although death-associated protein kinase (DAPK) is a Ca2+/calmodulin-regulated serine/threonine kinase that plays important roles in various types of apoptotic cell death, there have been no reports of its tissue distributions and functions in female reproductive organs. By comparing C57BL/6 wild-type mice with DAPK-mutant mice lacking the 74-amino acid catalytic kinase domain of DAPK, the cellular distributions and biological functions of DAPK in murine ovaries were investigated. In situ hybridization analyses with sense and antisense riboprobes revealed that DAPK mRNA was selectively and highly expressed in granulosa cells in the ovaries of both types of mice. There were no significant differences in the body weights, ovarian weights and unstimulated ovarian follicular numbers between the wild-type and DAPK-mutant mice. Intraperitoneal injection of CPT-11, an anticancer topoisomerase I inhibitor that causes granulosa cell-specific apoptosis partly through Fas-Fas ligand (FasL) interactions in MCH mice, induced follicular apoptosis in both the wild-type and DAPK-mutant mice. However, the numbers of apoptotic follicles were significantly reduced in the DAPK-mutant mice. The Fas and FasL expression levels in the CPT-11-injected mice did not differ significantly between the wild-type and DAPK-mutant mice. These results indicate that DAPK positively regulates intracellular signaling pathways for CPT-11-induced granulosa cell apoptosis.
Assuntos
Proteínas Reguladoras de Apoptose/química , Proteínas Reguladoras de Apoptose/genética , Apoptose , Proteínas Quinases Dependentes de Cálcio-Calmodulina/química , Proteínas Quinases Dependentes de Cálcio-Calmodulina/genética , Camptotecina/análogos & derivados , Regulação Neoplásica da Expressão Gênica , Folículo Ovariano/metabolismo , Neoplasias Ovarianas/tratamento farmacológico , Animais , Camptotecina/farmacologia , Proteínas Quinases Associadas com Morte Celular , Proteína Ligante Fas , Feminino , Deleção de Genes , Células da Granulosa/citologia , Humanos , Irinotecano , Camundongos , Camundongos Endogâmicos C57BL , Modelos Biológicos , Ovário/metabolismoRESUMO
BACKGROUND: Sema4D/CD100 is a type of class 4 semaphorin, exhibiting crucial roles in growth cone guidance in developing neurons. Sema4D is widely expressed throughout the central nervous system in embryonic mouse brain, and is selectively localized to oligodendrocytes and myelin in the postnatal brain. However, direct evidence of the actual involvement of Sema4D in the neuronal network development crucial for neurobehavioral performance is still lacking. The present study therefore examined whether Sema4D deficiency leads to abnormal behavioral development. METHODS: Both wild-type and Sema4D-deficient mice were subjected to behavioral analyses including open-field, adhesive tape removal, rotarod tests and a water maze task. RESULTS: Open-field tests revealed increased locomotor activity in Sema4D-deficient mice with less percentage of time spent in the center of the field. In both the adhesive tape removal and rotarod tests, which examine motor coordination and balance, Sema4D-deficient mice showed significantly superior performance, suggesting facilitated motor behavior. Both Sema4D-deficient and wild-type mice successfully learnt the water maze task, locating a hidden escape platform, and also showed precise memory for the platform position in probe tests. However, the swimming speed of Sema4D-deficient mice was significantly faster than that of wild-type mice, providing further evidence of their accelerated motor behavior. CONCLUSION: Our mouse behavioral analyses revealed enhanced motor activity in Sema4D-deficient mice, suggesting the crucial involvement of Sema4D in the neurodevelopmental processes of the central structures mediating motor behavior in mice.
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Comportamento Exploratório/fisiologia , Locomoção/genética , Atividade Motora/genética , Desempenho Psicomotor/fisiologia , Semaforinas/deficiência , Animais , Aprendizagem da Esquiva/fisiologia , Comportamento Animal/fisiologia , Calbindinas , Cerebelo/metabolismo , Aprendizagem em Labirinto/fisiologia , Camundongos , Camundongos Knockout , Tempo de Reação/genética , Teste de Desempenho do Rota-Rod/métodos , Proteína G de Ligação ao Cálcio S100/metabolismo , Comportamento Espacial/fisiologia , Fatores de TempoRESUMO
The corpus callosum (CC) is the biggest commissure that links cerebral hemispheres. Guidepost structures develop in the cortical midline during CC development and express axon guidance molecules that instruct neurons regarding the proper direction of axonal elongation toward and across the cortical midline. Neuropilin-1 (Npn1), a high affinity receptor for class 3 semaphorins (Sema3s) localized on cingulate pioneering axons, plays a crucial role in axon guidance to the midline through interactions with Sema3s. However, it remains unclear which type of Plexin is a component of Sema3 holoreceptors with Npn1 during the guidance of cingulate pioneering axons. To address the role of PlexinA1 in CC development, we examined with immunohistochemistry the localization of PlexinA1, Npn1, and Sema3s using embryonic brains from wild-type (WT) and PlexinA1-deficient (PlexinA1 knock-out (KO)) mice with a BALB/cAJ background. The immunohistochemistry confirmed the expression of PlexinA1 in callosal axons derived from the cingulate and neocortex of the WT mice on embryonic day 17.5 (E17.5) but not in the PlexinA1 KO mice. To examine the role of PlexinA1 in the navigation of callosal axons, the extension of callosal axons toward and across the midline was traced in brains of WT and PlexinA1 KO mice at E17.5. As a result, callosal axons in the PlexinA1 KO brains had a significantly lower incidence of midline crossing at E17.5 compared with the WT brains. To further examine the role of PlexinA1 in CC development, the CC phenotype was examined in PlexinA1 KO mice at postnatal day 0.5 (P0.5). Most of the PlexinA1 KO mice at P0.5 showed agenesis of the CC. These results indicate the crucial involvement of PlexinA1 in the midline crossing of callosal axons during CC development in BALB/cAJ mice.
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Axônios/metabolismo , Corpo Caloso/embriologia , Corpo Caloso/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Receptores de Superfície Celular/metabolismo , Agenesia do Corpo Caloso/embriologia , Agenesia do Corpo Caloso/patologia , Animais , Receptor DCC/metabolismo , Embrião de Mamíferos/metabolismo , Ligantes , Camundongos Endogâmicos BALB C , Camundongos Knockout , Neocórtex/metabolismo , Neuropilina-1/metabolismo , Fenótipo , Semaforina-3A/metabolismoRESUMO
Minimal optimal surgery without chemotherapy is often performed for patients with ovarian immature teratoma, which frequently occurs in young women who hope for future pregnancies. If tumors recur after the operation, anticancer drug chemotherapy is often administered, although few studies have highlighted differences between the recurrent and the primary tumor cells. Therefore, we have established experimental animal models of recurrent ovarian immature teratoma cells after optimal surgery and characterized the anticancer drug sensitivity and antigenicity of the recurrent tumors. Surgically-excised tumor cells of a grade II ovarian immature teratoma were cultured in vitro and transplanted into nude mice to establish stable cell lines. Differential drug sensitivity and antigenicity of the tumor cells were compared between the primary and the nude mouse tumors. Nude mouse tumor cells showed a normal 46XX karyotype. Cultured primary cells showed a remarkably high sensitivity to paclitaxel, docetaxel, adriamycin and pirarubicin, compared to peritoneal cancer cells obtained from a patient with ovarian adenocarcinomatous peritonitis. The drug sensitivity of teratoma cells to 5-fluorouracil, bleomycin or peplomycin was also significantly higher. However, there was no significant difference in sensitivity to platinum drugs between the primary teratoma and the peritoneal adenocarcinoma cells. As for nude mouse tumor cells, sensitivity to 12 anticancer drugs was significantly lower than that of the primary tumor cells, while there was little difference in sensitivity to carboplatin or peplomycin between the primary and nude mouse tumor cells. Flow cytometry showed that the expression of smooth muscle actin (SMA) significantly decreased in nude mouse tumor cells when compared to cultured primary cells. In conclusion, ovarian immature teratomas with normal karyotypes have a malignant potential to recur after minimal surgery. During nude mouse transplantation, SMA-overexpressing cells appeared to be selectively excluded and nude mouse tumor cells were less sensitive to the majority of anticancer drugs than the primary tumor cells. These results indicate that after optimal surgery for ovarian immature teratoma, recurrent cells can be more resistant to anticancer drugs than the primary tumors. Therefore, it is likely that adjuvant chemotherapy lowers the risk of ovarian immature teratomas recurring after optimal surgery. BEP and PBV regimens are frequently given to teratoma patients. However, paclitaxel/carboplatin or docetaxel/carboplatin, which are the most effective chemotherapy treatments for epithelial ovarian cancer patients, are considered to be an alternative regimen, especially in the prevention of reproductive toxicity.
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Recidiva Local de Neoplasia/patologia , Neoplasias Ovarianas/patologia , Teratoma/patologia , Adolescente , Animais , Linhagem Celular Tumoral , Ensaios de Seleção de Medicamentos Antitumorais , Feminino , Citometria de Fluxo , Humanos , Cariotipagem , Camundongos , Neoplasias Ovarianas/tratamento farmacológico , Neoplasias Ovarianas/cirurgia , Teratoma/tratamento farmacológico , Teratoma/cirurgiaRESUMO
Diphenylarsinic acid (DPAA) was a major compound found in the arsenic poisoning incident that occurred in Kamisu, Ibaraki, Japan in 2003. People exposed to DPAA via contaminated well water suffered from several neurological disorders, including cerebellar symptoms. We previously reported that DPAA induces cellular activation in cultured rat cerebellar astrocytes, dose-dependent promotion of cell growth (low DPAA), cell death (high DPAA), and increased phosphorylation of mitogen-activated protein (MAP) kinases (p38MAPK, SAPK/JNK, and ERK1/2). Moreover, DPAA induces up-regulation of oxidative stress-counteracting proteins, activation of CREB phosphorylation, increased protein expression of c-Jun and c-Fos, and aberrant secretion of brain-active cytokines (MCP-1, adrenomedullin, FGF2, CXCL1, and IL-6). Here, we explored the role of MAP kinases in DPAA-induced activation of astrocytes using specific MAP kinase signaling inhibitors [SB203580 (p38MAPK), SP600125 (SAPK/JNK), SCH772984 (ERK1/2), and U0126 (MEK1/2, a kinase for ERK1/2)]. DPAA-induced activation of MAP kinases had little contribution to DPAA-induced cell growth and death. On the other hand, a power relationship among MAP kinases was also observed, in which p38MAPK suppressed DPAA-induced SAPK/JNK and ERK1/2 activation, whereas ERK1/2 and MEK1/2 facilitated p38MAPK and SAPK/JNK activation. In addition, SAPK/JNK had minimal effects on the activation of other MAP kinases. DPAA-induced activation of transcription factors and secretion of brain-active cytokines were submissively but intricately dominated by MAP kinases. Collectively, our results indicate that DPAA-induced activation of MAP kinases is neither a cell growth-promoting response nor a cytoprotective one but leads to transcriptional disruption and aberrant secretion of brain-active cytokines in cerebellar astrocytes.
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Arsenicais/farmacologia , Astrócitos/efeitos dos fármacos , Cerebelo/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Animais , Astrócitos/enzimologia , Western Blotting , Células Cultivadas , Cerebelo/citologia , Cerebelo/enzimologia , Técnicas Imunoenzimáticas , RatosRESUMO
The molecular mechanism for cisplatin (CDDP)-resistance of cancer cells has not yet been clarified, despite extensive studies. Here, we investigated whether death-associated protein (DAP) kinase, an apoptosis modulator, was involved in CDDP-resistance by examining the ME180 human cervical squamous cancer cell line and 6 monoclonal ME180-derived CDDP-resistant subclones. Co-treatment with CDDP and 5-aza-2'-deoxycytidine (5-aza-CdR), a demethylating agent, significantly enhanced the CDDP-sensitivities of the parent cells and CDDP-resistant subclones. Subsequent removal of 5-aza-CdR rapidly reversed the CDDP-sensitivity of the CDDP-resistant subclones to their original levels, whereas the parent cells retained the enhanced CDDP-sensitivity for at least 24 h. Quantitative RT-PCR revealed that the CDDP-resistant subclones expressed higher DNA methyltransferase (DNMT) mRNA levels than the parent cells, suggesting that increased DNMT expressions easily restored the CDDP-resistance of the CDDP-resistant subclones following 5-aza-CdR removal. Although the parent cells showed hypermethylation in the DAP kinase promoter region, corresponding methylated bands were not detected in 2 of the 6 CDDP-resistant subclones by methylation-specific PCR. All 6 CDDP-resistant subclones expressed higher DAP kinase mRNA levels than the parent cells, as evaluated by quantitative RT-PCR. Although DAP kinase protein expression was strongly suppressed in the parent cells and CDDP-resistant subclones, 5-aza-CdR treatment of the parent cells dose-dependently stimulated the DAP kinase protein expression, and this was synergistically enhanced by inhibiting histone deacetylation via trichostatin treatment in addition to 5-aza-CdR. However, DAP kinase protein expression in the CDDP-resistant subclones was not stimulated by treatment with 5-aza-CdR and/or trichostatin. These results indicate that post-transcriptional translation of DAP kinase mRNA is strongly suppressed and insensitive to treatment with 5-aza-CdR and trichostatin in the CDDP-resistant subclones established from ME180 human cervical squamous cancer cells. This CDDP-resistance is accompanied by molecular changes that disturb the post-transcriptional translation of the DAP kinase mRNA, and these molecular changes are transiently restored by demethylation.
Assuntos
Antineoplásicos/farmacologia , Proteínas Reguladoras de Apoptose/metabolismo , Proteínas Quinases Dependentes de Cálcio-Calmodulina/metabolismo , Cisplatino/farmacologia , Resistencia a Medicamentos Antineoplásicos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Proteínas Reguladoras de Apoptose/genética , Azacitidina/análogos & derivados , Azacitidina/farmacologia , Proteínas Quinases Dependentes de Cálcio-Calmodulina/genética , Carcinoma de Células Escamosas , Linhagem Celular Tumoral , Metilação de DNA/efeitos dos fármacos , Metilases de Modificação do DNA/antagonistas & inibidores , Metilases de Modificação do DNA/genética , Metilases de Modificação do DNA/metabolismo , Proteínas Quinases Associadas com Morte Celular , Decitabina , Relação Dose-Resposta a Droga , Feminino , Humanos , Ácidos Hidroxâmicos/farmacologia , Regiões Promotoras Genéticas , Biossíntese de Proteínas/efeitos dos fármacos , Inibidores da Síntese de Proteínas/farmacologia , RNA Mensageiro/metabolismo , Neoplasias do Colo do ÚteroRESUMO
Patients with unresectable advanced carcinoma of the uterine cervix are usually treated with chemotherapy or chemoradiotherapy. In the present study, the optimal administration protocol for etoposide in chemotherapy and chemoradiotherapy for advanced cervical cancer patients was investigated in vitro using the radio-sensitive and anticancer drug-sensitive human cervical squamous cell carcinoma cell line ME180. Therapeutic doses of concurrent irradiation reduced the cellular etoposide sensitivity in a dose-dependent manner, while postirradiation-surviving subclones established from repeatedly irradiated ME180 cells showed significantly higher etoposide sensitivities than the non-irradiated parent cells. Of the 6 monoclonal etoposide-resistant subclones established from ME180 cells, 5 were significantly radioresistant. Although the etoposide-resistant subclones were also significantly resistant to other anticancer drugs, such as cisplatin, carboplatin, nedaplatin, pirarubicin, paclitaxel and docetaxel, they were more sensitive to 5-fluorouracil, mitomycin C and SN38 than the parent cells. Flow cytometric analyses revealed that the etoposide-resistant subclones showed significantly increased cell surface expression of CD40 compared to the parent cells, which expressed undetectable levels of CD40. However, the expression of some integrin receptor subunits, such as CD29, CD49a and CD49f, was apparently reduced in the etoposide-resistant subclones. These results indicate that etoposide should be administered to advanced cervical squamous cancer patients after the completion of radiotherapy, rather than as a concurrent chemoradiotherapy. In order to kill surviving etoposide-resistant cancer cells more effectively, 5-fluorouracil, mitomycin C and irinotecan may be candidate combination drugs for use with etoposide. Differential expression of integrin receptors and CD40 may be involved in the acquisition of etoposide resistance by cervical squamous cancer cells.
Assuntos
Antineoplásicos Fitogênicos/farmacologia , Etoposídeo/farmacologia , Raios X , Antígenos CD20/análise , Antígenos CD40/análise , Camptotecina/análogos & derivados , Camptotecina/farmacologia , Carcinoma de Células Escamosas/tratamento farmacológico , Carcinoma de Células Escamosas/patologia , Carcinoma de Células Escamosas/radioterapia , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos da radiação , Células Clonais/efeitos dos fármacos , Células Clonais/metabolismo , Células Clonais/efeitos da radiação , Terapia Combinada , Relação Dose-Resposta a Droga , Relação Dose-Resposta à Radiação , Resistencia a Medicamentos Antineoplásicos , Feminino , Citometria de Fluxo , Fluoruracila/farmacologia , Humanos , Integrina alfa1/análise , Integrina alfa6/análise , Irinotecano , Mitomicina/farmacologia , Neoplasias do Colo do Útero/tratamento farmacológico , Neoplasias do Colo do Útero/patologia , Neoplasias do Colo do Útero/radioterapiaRESUMO
Death-associated protein kinase (DAPK) is a Ca2+/calmodulin-dependent serine/threonine kinase that is thought to mediate apoptosis. DAPK is highly expressed in hippocampal neurons which are essential elements for memory formation. To examine if DAPK is implicated in spatial learning and memory, both wild-type and DAPK-mutant mice were subjected to Morris water maze tests. DAPK-mutant mice were generated by deleting 74 amino acids from the catalytic kinase domain of DAPK, and were used to investigate roles of the DAPK kinase domain in regulating spatial memory. Both mutant and wild-type mice were able to learn the water maze tasks to locate a hidden escape platform. In the first probe test, mutant mice showed a more precise memory for platform position compared to wild-type mice. In the reversal training in which the platform was located opposite from the original position, DAPK-mutant mice exhibited superior spatial learning compared to wild-type mice. DAPK-mutant mice also showed a more precise memory than their wild-type littermates in the probe trial of reversal test. Thus, the present results revealed crucial implications of DAPK in regulating spatial memory in mice.
Assuntos
Proteínas Reguladoras de Apoptose/genética , Proteínas Quinases Dependentes de Cálcio-Calmodulina/genética , Deleção de Genes , Memória/fisiologia , Comportamento Espacial/fisiologia , Animais , Proteínas Reguladoras de Apoptose/fisiologia , Comportamento Animal/fisiologia , Sítios de Ligação/genética , Proteínas Quinases Dependentes de Cálcio-Calmodulina/fisiologia , Proteínas Quinases Associadas com Morte Celular , Genótipo , Aprendizagem em Labirinto/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos KnockoutRESUMO
Diphenylarsinic acid (DPAA) was detected as the primary compound responsible for the arsenic poisoning that occurred in Kamisu, Ibaraki, Japan, where people using water from a well that was contaminated with a high level of arsenic developed neurological (mostly cerebellar) symptoms and dysregulation of regional cerebral blood flow. To understand the underlying molecular mechanism of DPAA-induced cerebellar symptoms, we focused on astrocytes, which have a brain-protective function. Incubation with 10 µM DPAA for 96 h promoted cell proliferation, increased the expression of antioxidative stress proteins (heme oxygenase-1 and heat shock protein 70), and induced the release of cytokines (MCP-1, adrenomedullin, FGF2, CXCL1, and IL-6). Furthermore, DPAA overpoweringly increased the phosphorylation of three major mitogen-activated protein kinases (MAPKs) (ERK1/2, p38MAPK, and SAPK/JNK), which indicated MAPK activation, and subsequently induced expression and/or phosphorylation of transcription factors (Nrf2, CREB, c-Jun, and c-Fos) in cultured rat cerebellar astrocytes. Structure-activity relationship analyses of DPAA and other related pentavalent organic arsenicals revealed that DPAA at 10 µM activated astrocytes most effective among organic arsenicals tested at the same dose. These results suggest that in a cerebellum exposed to DPAA, abnormal activation of the MAPK-transcription factor pathway and irregular secretion of these neuroactive, glioactive, and/or vasoactive cytokines in astrocytes can be the direct/indirect cause of functional abnormalities in surrounding neurons, glial cells, and vascular cells: This in turn might lead to the onset of cerebellar symptoms and disruption of cerebral blood flow.
Assuntos
Arsenicais/efeitos adversos , Astrócitos/efeitos dos fármacos , Cerebelo/efeitos dos fármacos , Citocinas/metabolismo , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Fatores de Transcrição/genética , Poluentes Químicos da Água/toxicidade , Animais , Animais Recém-Nascidos , Arsenicais/química , Astrócitos/enzimologia , Técnicas de Cultura de Células , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Cerebelo/citologia , Cerebelo/enzimologia , Relação Dose-Resposta a Droga , Fosforilação , Ratos Wistar , Relação Estrutura-Atividade , Fatores de Tempo , Regulação para Cima , Poluentes Químicos da Água/químicaRESUMO
Ghrelin is a gut-brain peptide and its endocrine activities are mediated by GH secretagogue receptor (GHSR)-1a. Des-acyl ghrelin does not activate GHSR-1a and is devoid of endocrine activities. While the microinjection of ghrelin into rat nucleus tractus solitarii (NTS) elicited hypotensive effects, this was not the case upon injection into GHSR-expressing rostral ventrolateral medulla or caudal ventrolateral medulla. To make clear the reason of the discrepancy between receptor distribution and neuronal responses, we examined the cardiovascular response of rats microinjected with des-acyl ghrelin into NTS. Intra-NTS injection of des-acyl ghrelin significantly reduced mean arterial pressure and heart rate. The hypotensive and bradycardic activity evoked by des-acyl ghrelin was not significantly different from that of native ghrelin. These results suggest that des-acyl ghrelin contribute to the regulation of cardiovascular control and that a receptor other than GHSR-1a exists in NTS.
Assuntos
Anti-Hipertensivos/administração & dosagem , Hipotensão/induzido quimicamente , Hormônios Peptídicos/administração & dosagem , Núcleo Solitário/efeitos dos fármacos , Animais , Pressão Sanguínea/efeitos dos fármacos , Pressão Sanguínea/fisiologia , Grelina , Hipotensão/fisiopatologia , Masculino , Microinjeções , Ratos , Ratos Wistar , Núcleo Solitário/fisiologiaRESUMO
Attempts at protein transduction into specific restricted brain areas have remained unsuccessful. We attempted targeted, direct in vivo protein transduction by microinjecting beta-galactosidase (beta-gal) with hemagglutinating virus of Japan envelope (HVJ-E) vector into the rat nucleus tractus solitarius (NTS). The medulla oblongata including the NTS was removed 6h post-injection and cryostat sections were histochemically stained to detect beta-gal enzymatic activity. beta-gal-positive cells were present in these sections as was beta-gal activity determined by colorimetric analysis. beta-gal-positive cells were not present in the rats microinjected only beta-gal protein without HVJ-E vector. Our findings suggest that direct in vivo protein transduction into specific restricted brain areas is possible. The type of targeted delivery system we present may have wide applications in the administration of therapeutic proteins to the central nervous system.