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1.
Hum Mol Genet ; 29(15): 2508-2522, 2020 08 29.
Artigo em Inglês | MEDLINE | ID: mdl-32620959

RESUMO

Bardet-Biedl syndrome (BBS) is a pleiotropic autosomal recessive ciliopathy affecting multiple organs. The development of potential disease-modifying therapy for BBS will require concurrent targeting of multi-systemic manifestations. Here, we show for the first time that monosialodihexosylganglioside accumulates in Bbs2-/- cilia, indicating impairment of glycosphingolipid (GSL) metabolism in BBS. Consequently, we tested whether BBS pathology in Bbs2-/- mice can be reversed by targeting the underlying ciliary defect via reduction of GSL metabolism. Inhibition of GSL synthesis with the glucosylceramide synthase inhibitor Genz-667161 decreases the obesity, liver disease, retinal degeneration and olfaction defect in Bbs2-/- mice. These effects are secondary to preservation of ciliary structure and signaling, and stimulation of cellular differentiation. In conclusion, reduction of GSL metabolism resolves the multi-organ pathology of Bbs2-/- mice by directly preserving ciliary structure and function towards a normal phenotype. Since this approach does not rely on the correction of the underlying genetic mutation, it might translate successfully as a treatment for other ciliopathies.


Assuntos
Síndrome de Bardet-Biedl/genética , Cílios/genética , Ciliopatias/genética , Proteínas/genética , Animais , Síndrome de Bardet-Biedl/tratamento farmacológico , Síndrome de Bardet-Biedl/patologia , Diferenciação Celular/efeitos dos fármacos , Cílios/patologia , Ciliopatias/tratamento farmacológico , Ciliopatias/patologia , Modelos Animais de Doenças , Inibidores Enzimáticos/farmacologia , Gangliosídeos/biossíntese , Gangliosídeos/genética , Glucosiltransferases/antagonistas & inibidores , Glucosiltransferases/genética , Glicoesfingolipídeos/biossíntese , Glicoesfingolipídeos/genética , Camundongos Knockout
2.
J Neurochem ; 156(5): 692-701, 2021 03.
Artigo em Inglês | MEDLINE | ID: mdl-32743826

RESUMO

Most lysosomal storage diseases (LSDs) have a significant neurological component, including types 2 and 3 Gaucher disease (neuronal forms of Gaucher disease; nGD). No therapies are currently available for nGD since the recombinant enzymes used in the systemic form of Gaucher disease do not cross the blood-brain barrier (BBB). However, a number of promising approaches are currently being tested, including substrate reduction therapy (SRT), in which partial inhibition of the synthesis of the glycosphingolipids (GSLs) that accumulate in nGD lowers their accumulation. We now induce nGD in mice by injection with conduritol B-epoxide (CBE), an irreversible inhibitor of acid beta-glucosidase (GCase), the enzyme defective in nGD, with or without co-injection with Genz-667161, a prototype for SRT which crosses the BBB. Significant neuropathology, and a reduction in lifespan, was observed upon CBE injection, and this was largely reversed by co-injection with Genz-667161, along with a reduction in glucosylceramide and glucosylsphingosine levels. Analysis of gene expression by RNAseq revealed that Genz-667161 largely reversed the changes in genes and pathways that were differentially expressed upon CBE injection, specifically pathways of GSL metabolism, lipoproteins and other lipid metabolic pathways, lipid droplets, astrocyte activation, neuronal function, and to some extent, neuroinflammation. Together, this demonstrates the efficacy of SRT to reverse the effects of substrate accumulation on pathological components and pathways in nGD brain.


Assuntos
Modelos Animais de Doenças , Doença de Gaucher/metabolismo , Doença de Gaucher/patologia , Glucosilceramidase/antagonistas & inibidores , Glicoesfingolipídeos/antagonistas & inibidores , Animais , Barreira Hematoencefálica/efeitos dos fármacos , Barreira Hematoencefálica/metabolismo , Barreira Hematoencefálica/patologia , Inibidores Enzimáticos/farmacologia , Inibidores Enzimáticos/uso terapêutico , Feminino , Doença de Gaucher/tratamento farmacológico , Glucosilceramidase/metabolismo , Glicoesfingolipídeos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Especificidade por Substrato/efeitos dos fármacos , Especificidade por Substrato/fisiologia
3.
Mov Disord ; 36(5): 1216-1228, 2021 05.
Artigo em Inglês | MEDLINE | ID: mdl-33547828

RESUMO

BACKGROUND: With pathway-specific trials in PD associated with variants in the glucocerebrosidase gene (PDGBA ) under way, we need markers that confirm the impact of genetic variants in patient-derived biofluids in order to allow patient stratification merely based on genetics and that might serve as biochemical read-out for target engagement. OBJECTIVE: To explore GBA-pathway-specific biomarker profiles cross-sectionally (TUEPAC-MIGAP, PPMI) and longitudinally (PPMI). METHODS: We measured enzyme activity of the lysosomal glucocerebrosidase, CSF levels of glucosylceramides (upstream substrate of glucocerebrosidase), CSF levels of ceramides (downstream product of glucocerebrosidase), lactosylceramides, sphingosines, sphingomyelin (by-products) and CSF levels of total α-synuclein in PDGBA patients compared to PDGBA_wildtype patients. RESULTS: Cross-sectionally in both cohorts and longitudinally in PPMI: (1) glucocerebrosidase activity was significantly lower in PDGBA compared to PDGBA_wildtype . (2) CSF levels of upstream substrates (glucosylceramides species) were higher in PDGBA compared to PDGBA_wildtype . (3) CSF levels of total α-synuclein were lower in PDGBA compared to PDGBA_wildtype . All of these findings were most pronounced in PDGBA with severe mutations (PDGBA_severe ). Cross-sectionally in TUEPAC-MIGAP and longitudinally in PPMI, CSF levels of downstream-products (ceramides) were higher in PDGBA_severe . Cross-sectionally in TUEPAC-MIGAP by-products sphinganine and sphingosine-1-phosphate and longitudinally in PPMI species of by-products lactosylceramides and sphingomyelin were higher in PDGBA_severe . INTERPRETATION: These findings confirm that GBA mutations have a relevant functional impact on biomarker profiles in patients. Bridging the gap between genetics and biochemical profiles now allows patient stratification for clinical trials merely based on mutation status. Importantly, all findings were most prominent in PDGBA with severe variants. © 2021 International Parkinson and Movement Disorder Society.


Assuntos
Doença de Parkinson , alfa-Sinucleína , Glucosilceramidase/genética , Humanos , Mutação/genética , Doença de Parkinson/genética , Esfingolipídeos , alfa-Sinucleína/genética
4.
Biotechnol Bioeng ; 118(8): 3263-3268, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-33990942

RESUMO

Here, we report a bienzymatic cascade to produce ß-amino acids as an intermediate for the synthesis of the leading oral antidiabetic drug, sitagliptin. A whole-cell biotransformation using recombinant Escherichia coli coexpressing a esterase and transaminase were developed, wherein the desired expression level of each enzyme was achieved by promotor engineering. The small-scale reactions (30 ml) performed under optimized conditions at varying amounts of substrate (100-300 mM) resulted in excellent conversions of 82%-95% for the desired product. Finally, a kilogram-scale enzymatic reaction (250 mM substrate, 220 L) was carried out to produce ß-amino acid (229 mM). Sitagliptin phosphate was chemically synthesized from ß-amino acids with 82% yield and > 99% purity.


Assuntos
Escherichia coli , Esterases , Engenharia Genética , Microrganismos Geneticamente Modificados , Regiões Promotoras Genéticas , Fosfato de Sitagliptina/metabolismo , Transaminases , Escherichia coli/genética , Escherichia coli/metabolismo , Esterases/genética , Esterases/metabolismo , Microrganismos Geneticamente Modificados/genética , Microrganismos Geneticamente Modificados/metabolismo , Transaminases/genética , Transaminases/metabolismo
5.
J Youth Adolesc ; 50(9): 1824-1838, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34263406

RESUMO

Latino college graduation rates continue to fall behind rates of other racial/ethnic groups, highlighting the importance of understanding risk and protective processes across the transition into higher education. The current study examined changes in socio-cultural contexts (i.e., campus and neighborhood co-ethnic composition) and academic achievement across the college transition for Latino adolescents and investigated whether familism values moderated associations. Participants were 188 Latino late-adolescents (Mage = 18.12; SD = 0.40; 62.9% female). Greater campus incongruency (i.e., decrease in co-ethnic composition) was associated with lower achievement for adolescents with low familism values, but not those with average or high levels. Change in neighborhood co-ethnic composition was not associated with achievement. Moving to incongruent campus contexts may be risky for Latino youth who report low levels of familism values, underscoring the importance of sociocultural protective processes in person-context transitions.


Assuntos
Sucesso Acadêmico , Hispânico ou Latino , Adolescente , Escolaridade , Etnicidade , Feminino , Humanos , Masculino , Instituições Acadêmicas
6.
J Youth Adolesc ; 50(3): 506-520, 2021 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-33025287

RESUMO

Spending time with family ("family connection") is a salient aspect of adolescents' daily lives linked with healthy sleep. Less is known regarding the unique effects of parent and sibling connection on sleep. This study examined daily and average associations between parent/sibling connection and objective sleep (duration, efficiency) in a sample of Latinx adolescents (N = 195; Mage = 18.11, SD = 0.41; 65.6% female) and explored familism values and family communication as moderators. Adolescents slept longer on days that they spent more time with siblings, and youth who typically spent more time with parents had longer sleep durations. Family communication and familism-obligation moderated associations between family connection and sleep. These results provide support for the role of family interactions in promoting healthy sleep among Latinx adolescents.


Assuntos
Pais , Sono , Adolescente , Comunicação , Feminino , Objetivos , Humanos , Masculino
7.
Mol Ther ; 27(8): 1495-1506, 2019 08 07.
Artigo em Inglês | MEDLINE | ID: mdl-31208914

RESUMO

Neuronopathic glycosphingolipidoses are a sub-group of lysosomal storage disorders for which there are presently no effective therapies. Here, we evaluated the potential of substrate reduction therapy (SRT) using an inhibitor of glucosylceramide synthase (GCS) to decrease the synthesis of glucosylceramide (GL1) and related glycosphingolipids. The substrates that accumulate in Sandhoff disease (e.g., ganglioside GM2 and its nonacylated derivative, lyso-GM2) are distal to the drug target, GCS. Treatment of Sandhoff mice with a GCS inhibitor that has demonstrated CNS access (Genz-682452) reduced the accumulation of GL1 and GM2, as well as a variety of disease-associated substrates in the liver and brain. Concomitant with these effects was a significant decrease in the expression of CD68 and glycoprotein non-metastatic melanoma B protein (Gpnmb) in the brain, indicating a reduction in microgliosis in the treated mice. Moreover, using in vivo imaging, we showed that the monocytic biomarker translocator protein (TSPO), which was elevated in Sandhoff mice, was normalized following Genz-682452 treatment. These positive effects translated in turn into a delay (∼28 days) in loss of motor function and coordination, as measured by rotarod latency, and a significant increase in longevity (∼17.5%). Together, these results support the development of SRT for the treatment of gangliosidoses, particularly in patients with residual enzyme activity.


Assuntos
Carbamatos/farmacologia , Inibidores Enzimáticos/farmacologia , Glucosiltransferases/antagonistas & inibidores , Quinuclidinas/farmacologia , Doença de Sandhoff/enzimologia , Animais , Encéfalo/efeitos dos fármacos , Encéfalo/metabolismo , Encéfalo/patologia , Modelos Animais de Doenças , Feminino , Glucosiltransferases/genética , Glucosiltransferases/metabolismo , Ligantes , Fígado/efeitos dos fármacos , Fígado/metabolismo , Fígado/patologia , Masculino , Espectrometria de Massas , Camundongos , Camundongos Knockout , Imagem Molecular , Receptores de GABA/metabolismo , Doença de Sandhoff/diagnóstico , Doença de Sandhoff/genética , Doença de Sandhoff/terapia , Esfingolipídeos/metabolismo , Cadeia beta da beta-Hexosaminidase/genética , Cadeia beta da beta-Hexosaminidase/metabolismo
8.
Proc Natl Acad Sci U S A ; 114(10): 2699-2704, 2017 03 07.
Artigo em Inglês | MEDLINE | ID: mdl-28223512

RESUMO

Mutations in the glucocerebrosidase gene (GBA) confer a heightened risk of developing Parkinson's disease (PD) and other synucleinopathies, resulting in a lower age of onset and exacerbating disease progression. However, the precise mechanisms by which mutations in GBA increase PD risk and accelerate its progression remain unclear. Here, we investigated the merits of glucosylceramide synthase (GCS) inhibition as a potential treatment for synucleinopathies. Two murine models of synucleinopathy (a Gaucher-related synucleinopathy model, GbaD409V/D409V and a A53T-α-synuclein overexpressing model harboring wild-type alleles of GBA, A53T-SNCA mouse model) were exposed to a brain-penetrant GCS inhibitor, GZ667161. Treatment of GbaD409V/D409V mice with the GCS inhibitor reduced levels of glucosylceramide and glucosylsphingosine in the central nervous system (CNS), demonstrating target engagement. Remarkably, treatment with GZ667161 slowed the accumulation of hippocampal aggregates of α-synuclein, ubiquitin, and tau, and improved the associated memory deficits. Similarly, prolonged treatment of A53T-SNCA mice with GZ667161 reduced membrane-associated α-synuclein in the CNS and ameliorated cognitive deficits. The data support the contention that prolonged antagonism of GCS in the CNS can affect α-synuclein processing and improve behavioral outcomes. Hence, inhibition of GCS represents a disease-modifying therapeutic strategy for GBA-related synucleinopathies and conceivably for certain forms of sporadic disease.


Assuntos
Carbamatos/farmacologia , Inibidores Enzimáticos/administração & dosagem , Glucosiltransferases/antagonistas & inibidores , Doença de Parkinson/tratamento farmacológico , Quinuclidinas/farmacologia , alfa-Sinucleína/genética , Animais , Modelos Animais de Doenças , Regulação da Expressão Gênica , Glucosiltransferases/genética , Humanos , Camundongos , Mutação , Doença de Parkinson/enzimologia , Doença de Parkinson/patologia , Agregação Patológica de Proteínas/tratamento farmacológico , Agregação Patológica de Proteínas/genética , Agregação Patológica de Proteínas/patologia , Ubiquitina/metabolismo , Proteínas tau/metabolismo
9.
J Korean Med Sci ; 35(15): e101, 2020 Apr 20.
Artigo em Inglês | MEDLINE | ID: mdl-32301293

RESUMO

BACKGROUND: Despite the increasing importance of rehabilitation for critically ill patients, there is little information regarding how rehabilitation therapy is utilized in clinical practice. Our objectives were to evaluate the implementation rate of rehabilitation therapy in the intensive care unit (ICU) survivors and to investigate the effects of rehabilitation therapy on outcomes. METHODS: A retrospective nationwide cohort study with including > 18 years of ages admitted to ICU between January 2008 and May 2015 (n = 1,465,776). The analyzed outcomes were readmission to ICU readmission and emergency room (ER) visit. RESULTS: During the study period, 249,918 (17.1%) patients received rehabilitation therapy. The percentage of patients receiving any rehabilitation therapy increased annually from 14% in 2008 to 20% in 2014, and the percentages for each type of therapy also increased over time. The most common type of rehabilitation was physical therapy (91.9%), followed by neuromuscular electrical stimulation (29.6%), occupational (28.6%), respiratory, (11.6%) and swallowing (10.3%) therapies. After adjusting for confounding variables, the risk of 30-day ICU readmission was lower in patients who received rehabilitation therapy than in those who did not (P < 0.001; hazard ratio [HR], 0.70; 95% confidence interval [CI], 0.65-0.75). And, the risk of 30-day ER visit was also lower in patients who received rehabilitation therapy (P < 0.001; HR, 0.83; 95% CI, 0.77-0.88). CONCLUSION: In this nationwide cohort study in Korea, only 17% of all ICU patients received rehabilitation therapy. However, rehabilitation is associated with a significant reduction in the risk of 30-day ICU readmission and ER visit.


Assuntos
Doenças do Sistema Nervoso Central/reabilitação , Serviços Médicos de Emergência/estatística & dados numéricos , Readmissão do Paciente/estatística & dados numéricos , Sobreviventes/estatística & dados numéricos , Adulto , Idoso , Doenças do Sistema Nervoso Central/mortalidade , Doenças do Sistema Nervoso Central/patologia , Comorbidade , Bases de Dados Factuais , Feminino , Humanos , Unidades de Terapia Intensiva , Masculino , Pessoa de Meia-Idade , Alta do Paciente , Modelos de Riscos Proporcionais , República da Coreia , Estudos Retrospectivos
10.
Mil Psychol ; 32(6): 408-416, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-38536242

RESUMO

Given over 2.77 million US service members have been deployed in the past 20 years and the intense process of reintegration to civilian life, understanding factors that contribute to Veterans' mental health and substance use is critical. This study sought to understand the effects of US identity exploration, US identity commitment, US identity affirmation, and US identity centrality on substance use and symptoms of depression and anxiety. The sample consisted of 195 US military Veterans (n = 184, 53.3% women; 73.3% White; Mage = 35.12 years, SD = 9.60 years). Bivariate correlations indicated US identity affirmation was negatively associated with substance use and symptoms of depression and anxiety whereas US identity centrality was positively correlated with alcohol use. Utilizing structural equation model, US identity affirmation and US identity centrality were, respectively, negatively and positively associated with alcohol use, substance use, and symptoms of depression and anxiety. Partially consistent with our hypothesis, US identity exploration was positively associated with symptoms of anxiety. In contrast to our hypothesis, US identity commitment was not significantly associated with any outcome. Results are discussed in terms of important directions for identity research in the transition to civilian life.

11.
Hum Mol Genet ; 25(13): 2645-2660, 2016 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-27126635

RESUMO

Mutations in GBA1, the gene encoding glucocerebrosidase, are associated with an enhanced risk of developing synucleinopathies such as Parkinson's disease (PD) and dementia with Lewy bodies. A higher prevalence and increased severity of motor and non-motor symptoms is observed in PD patients harboring mutant GBA1 alleles, suggesting a link between the gene or gene product and disease development. Interestingly, PD patients without mutations in GBA1 also exhibit lower levels of glucocerebrosidase activity in the central nervous system (CNS), implicating this lysosomal enzyme in disease pathogenesis. Here, we investigated whether modulation of glucocerebrosidase activity in murine models of synucleinopathy (expressing wild type Gba1) affected α-synuclein accumulation and behavioral phenotypes. Partial inhibition of glucocerebrosidase activity in PrP-A53T-SNCA mice using the covalent inhibitor conduritol-B-epoxide induced a profound increase in soluble α-synuclein in the CNS and exacerbated cognitive and motor deficits. Conversely, augmenting glucocerebrosidase activity in the Thy1-SNCA mouse model of PD delayed the progression of synucleinopathy. Adeno-associated virus-mediated expression of glucocerebrosidase in the Thy1-SNCA mouse striatum led to decrease in the levels of the proteinase K-resistant fraction of α-synuclein, amelioration of behavioral aberrations and protection from loss of striatal dopaminergic markers. These data indicate that increasing glucocerebrosidase activity can influence α-synuclein homeostasis, thereby reducing the progression of synucleinopathies. This study provides robust in vivo evidence that augmentation of CNS glucocerebrosidase activity is a potential therapeutic strategy for PD, regardless of the mutation status of GBA1.


Assuntos
Glucosilceramidase/metabolismo , Glucosilceramidase/fisiologia , Animais , Cognição/efeitos dos fármacos , Modelos Animais de Doenças , Dopamina , Doença de Gaucher/genética , Expressão Gênica , Glucosilceramidase/genética , Glucosilceramidase/uso terapêutico , Humanos , Camundongos , Atividade Motora/efeitos dos fármacos , Mutação , Doença de Parkinson/tratamento farmacológico , Doença de Parkinson/genética , alfa-Sinucleína/líquido cefalorraquidiano , alfa-Sinucleína/metabolismo
12.
Dev Psychopathol ; 30(5): 1779-1796, 2018 12.
Artigo em Inglês | MEDLINE | ID: mdl-30259820

RESUMO

Culturally linked family influences during adolescence are important predictors of health and well-being for Latino youth, yet few studies have examined whether these familial influences are associated with indicators of typical physiological stress processes. Following a cultural neurobiology framework, we examined the role of family in the everyday lives of Latino adolescents (N = 209; Mage = 18.10; 85.1% Mexican descent; 64.4% female) by investigating familism values and perceptions of parent support as well as daily family assistance behaviors in relation to hypothalamic-pituitary-adrenal axis diurnal patterns, indexed by salivary cortisol five times a day for 3 weekdays. Three-level growth curve analyses revealed that perceptions of parental support were associated with greater cortisol awakening responses, whereas familism values were not associated with diurnal cortisol patterns. In day-to-day analyses, assisting family during the day (compared to not assisting family) was associated with lower waking cortisol levels and flatter diurnal slopes the next day. Our findings highlight the dynamic associations and multiple time courses between cultural values and behaviors, daily experiences, and physiological stress processes for Latino adolescents. Further, we identified important cultural risk and promotive factors associated with physiological regulation in daily life and potential pathways toward health outcomes in adulthood.


Assuntos
Ritmo Circadiano/fisiologia , Relações Familiares , Hispânico ou Latino , Hidrocortisona/metabolismo , Sistema Hipotálamo-Hipofisário/metabolismo , Sistema Hipófise-Suprarrenal/metabolismo , Apoio Social , Adolescente , Relações Familiares/etnologia , Feminino , Humanos , Masculino , Relações Pais-Filho/etnologia , Saliva
13.
Mol Ther ; 24(6): 1019-1029, 2016 06.
Artigo em Inglês | MEDLINE | ID: mdl-26948439

RESUMO

Gaucher disease (GD) is caused by a deficiency of glucocerebrosidase and the consequent lysosomal accumulation of unmetabolized glycolipid substrates. Enzyme-replacement therapy adequately manages the visceral manifestations of nonneuronopathic type-1 Gaucher patients, but not the brain disease in neuronopathic types 2 and 3 GD. Substrate reduction therapy through inhibition of glucosylceramide synthase (GCS) has also been shown to effectively treat the visceral disease. Here, we evaluated the efficacy of a novel small molecule inhibitor of GCS with central nervous system (CNS) access (Genz-682452) to treat the brain disease. Treatment of the conduritol ß epoxide-induced mouse model of neuronopathic GD with Genz-682452 reduced the accumulation of liver and brain glycolipids (>70% and >20% respectively), extent of gliosis, and severity of ataxia. In the genetic 4L;C* mouse model, Genz-682452 reduced the levels of substrate in the brain by >40%, the extent of gliosis, and paresis. Importantly, Genz-682452-treated 4L;C* mice also exhibited an ~30% increase in lifespan. Together, these data indicate that an orally available antagonist of GCS that has CNS access is effective at attenuating several of the neuropathologic and behavioral manifestations associated with mouse models of neuronopathic GD. Therefore, Genz-682452 holds promise as a potential therapeutic approach for patients with type-3 GD.


Assuntos
Carbamatos/administração & dosagem , Sistema Nervoso Central/metabolismo , Inibidores Enzimáticos/administração & dosagem , Doença de Gaucher/tratamento farmacológico , Glucosiltransferases/antagonistas & inibidores , Glicolipídeos/metabolismo , Quinuclidinas/administração & dosagem , Administração Oral , Animais , Carbamatos/farmacologia , Modelos Animais de Doenças , Inibidores Enzimáticos/farmacologia , Doença de Gaucher/induzido quimicamente , Doença de Gaucher/metabolismo , Humanos , Inositol/análogos & derivados , Fígado/metabolismo , Lisossomos/efeitos dos fármacos , Lisossomos/enzimologia , Camundongos , Quinuclidinas/farmacologia , Distribuição Tecidual , Resultado do Tratamento
14.
Drug Chem Toxicol ; 39(1): 111-8, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-26045230

RESUMO

Three chemotoxins including dimethylnitrosamine (DMN), carbon tetrachloride (CCl4), and thioacetamide (TAA) are commonly used in hepatofibrotic models. We aimed to draw characteristics of histopathology and pro-fibrogenic cytokines including TGF-ß, PDGF and CTGF among three models. Rats were divided into six groups and intra-peritoneally injected with DMN (10 mg/kg, for three weeks, three consecutive days weekly), CCl4 (1.6 g/kg, for 10 weeks, twice weekly), TAA (200 mg/kg, for 12 weeks, twice weekly) or their corresponded treatment for each control group. The liver weights were decreased in DMN model, but not other models. Ascites were occurred as 3-, 2-, and 7-rats in DMN, CCl4, and TAA model, respectively. The lipid peroxidation was highest in CCl4 model, serum levels of liver enzymes were increased as similar severity. The hepatofibrotic alterations were remarkable in DMN and TAA model, but not CCl4 as evidenced by the Masson trichrome staining and hydroxyproline. The immunohistochemistry for α-SAM showed that the DMN model was most severely enhanced than other models. On the other hand, hepatic tissue levels of pro-fibrogenic cytokines including TGF-ß, PDGF, and CTGF were generally increased in three models, but totally different among models or measurement resources. Especially, serum levels of three cytokines were remarkably increased by CCl4 injection and CTGF levels in both hepatic tissue and serum were highest in CCl4 group. Our results firstly demonstrated comparative study for features of morphological finding and pro-fibrogenic cytokines in serum and hepatic protein levels among three models. Above results would be a helpful reference for hepatofibrotic studies.


Assuntos
Tetracloreto de Carbono/toxicidade , Dimetilnitrosamina/toxicidade , Cirrose Hepática Experimental/fisiopatologia , Tioacetamida/toxicidade , Animais , Fator de Crescimento do Tecido Conjuntivo/metabolismo , Modelos Animais de Doenças , Imuno-Histoquímica , Peroxidação de Lipídeos/efeitos dos fármacos , Cirrose Hepática Experimental/induzido quimicamente , Masculino , Fator de Crescimento Derivado de Plaquetas/metabolismo , Ratos , Ratos Sprague-Dawley , Fator de Crescimento Transformador beta/metabolismo
15.
J Lipid Res ; 55(4): 773-81, 2014 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-24482490

RESUMO

Gangliosides and sulfatides (STs) are acidic glycosphingolipids (GSLs) that have one or more sialic acids or sulfate substituents, in addition to neutral sugars, attached to the C-1 hydroxyl group of the ceramide long chain base. TLC is a widely employed and convenient technique for separation and characterization of GSLs. When TLC is directly coupled to MS, it provides both the molecular mass and structural information without further purification. Here, after development of the TLC plates, the structural analyses of acidic GSLs, including gangliosides and STs, were investigated using the liquid extraction surface analysis (LESA™) and CAMAG TLC-MS interfaces coupled to an ESI QSTAR Pulsar i quadrupole orthogonal TOF mass spectrometer. Coupling TLC with ESI-MS allowed the acquisition of high resolution mass spectra of the acidic GSLs with high sensitivity and mass accuracy, without the loss of sialic acid residues that frequently occurs during low-pressure MALDI MS. These systems were then applied to the analysis of total lipid extracts from bovine brain. This allowed profiling of many different lipid classes, not only gangliosides and STs, but also SMs, neutral GSLs, and phospholipids.


Assuntos
Glicolipídeos/química , Glicoesfingolipídeos/química , Ácidos Siálicos/química , Animais , Química Encefálica , Bovinos , Cromatografia em Camada Fina , Sensibilidade e Especificidade , Espectrometria de Massas por Ionização por Electrospray , Espectrometria de Massas em Tandem
16.
Anal Chem ; 86(12): 5697-705, 2014 Jun 17.
Artigo em Inglês | MEDLINE | ID: mdl-24832382

RESUMO

A simple, fast approach elucidated carbon-carbon double bond positions in unsaturated lipids. Lipids were deposited onto various surfaces and the products from their oxidation in ambient air were observed by electrospray ionization (ESI) mass spectrometry (MS). The most common oxidative products, aldehydes, were detected as transformations at the cleaved double bond positions. Ozonides and carboxylic acids were generated in certain lipids. Investigations of the conditions controlling the appearance of these products indicated that the surface oxidation depends on light and ambient air. Since the lipid oxidation was slower in a high concentration of ozone, singlet oxygen appeared to be a parallel oxidant for unsaturated lipids. The 3-hydroxyl group in the sphingoid base of sulfatides offered some protection from oxidation for the Δ4,5-double bond, slowing its oxidation rate relative to that of the isolated double bond in the N-linked fatty acyl chain. Direct sampling by thin-layer chromatography (TLC)-ESI-MS provides a powerful approach to elucidate detailed structural information on biological samples. Co-localization of the starting lipids and their oxidation products after TLC separation allowed assignment of the native unsaturation sites. Phosphatidylserine and N,N-dimethyl phosphatidylethanolamine isomers in a bovine brain total lipid extract were distinguished on the basis of their oxidation products. Meanwhile, the findings reported herein reveal a potential pitfall in the assignment of structures to lipids extracted from TLC plates because of artifactual oxidation after the plate development.


Assuntos
Ar , Lipídeos/química , Cromatografia em Camada Fina , Oxirredução , Espectrometria de Massas por Ionização por Electrospray
17.
J Biol Chem ; 287(12): 9128-36, 2012 Mar 16.
Artigo em Inglês | MEDLINE | ID: mdl-22291021

RESUMO

Sphingosine 1-phosphate lyase (S1P lyase) irreversibly cleaves sphingosine 1-phosphate (S1P) in the final step of sphingolipid catabolism. As sphingoid bases and their 1-phosphate are not only metabolic intermediates but also highly bioactive lipids that modulate a wide range of physiological processes, it would be predicted that their elevation might induce adjustments in other facets of sphingolipid metabolism and/or alter cell behavior. Indeed, we have previously reported that S1P lyase deficiency causes neurodegeneration and other adverse symptoms. We next asked the question whether and how S1P lyase deficiency affects the metabolism of (glyco)sphingolipids and cholesterol, two lipid classes that might be involved in the neurodegenerative processes observed in S1P lyase-deficient mice. As predicted, there was a considerable increase in free and phosphorylated sphingoid bases upon elimination of S1P lyase, but to our surprise, rather than increasing, the mass of (glyco)sphingolipids persisted at wild type levels. This was discovered to be due to reduced de novo sphingoid base biosynthesis and a corresponding increase in the recycling of the backbones via the salvage pathway. There was also a considerable increase in cholesterol esters, although free cholesterol persisted at wild type levels, which might be secondary to the shifts in sphingolipid metabolism. All in all, these findings show that accumulation of free and phosphorylated sphingoid bases by loss of S1P lyase causes an interesting readjustment of the balance between de novo biosynthesis and recycling to maintain (glyco)sphingolipid homeostasis. These changes, and their impact on the metabolism of other cellular lipids, should be explored as possible contributors to the neurodegeneration in S1P lyase deficiency.


Assuntos
Aldeído Liases/deficiência , Neurônios/enzimologia , Esfingolipídeos/biossíntese , Aldeído Liases/genética , Animais , Células Cultivadas , Camundongos , Camundongos Knockout , Neurônios/metabolismo
18.
Molecules ; 18(5): 5317-25, 2013 May 08.
Artigo em Inglês | MEDLINE | ID: mdl-23698045

RESUMO

Changes in the volatile components of unpasteurized and pasteurized makgeolli during 30 days of storage were investigated by gas chromatography-mass spectrometry (GC-MS) and GC-olfactometry (GC-O). A total of 11 odor-active compounds such as 3-methyl-1-butanol (isoamyl alcohol), 2-methyl-1-butanol, 2,3-butanediol, butanoic acid, 3-methylbutanoic acid (isovaleric acid), 2-methylbutanoic acid, 3-(methylthio)-1-propanol (methionol), 2-phenylethanol, ethyl decanoate, ethyl dodecanoate, and ethyl tetradecanoate were determined in both the pasteurized and unpasteurized makgeolli during 30 days of storage. Although there were no significant differences in the concentrations of odor-active compounds at the initial storage time, most of odor-active compounds were more significantly increased in unpasteurized makgeolli compared to the pasteurized one during the storage period.


Assuntos
Análise de Alimentos , Compostos Orgânicos Voláteis/análise , Vinho/análise , Oryza/química , Pasteurização , Fatores de Tempo , Compostos Orgânicos Voláteis/química
19.
Arch Pharm Res ; 46(2): 111-116, 2023 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-36564599

RESUMO

Tolperisone hydrochloride is a centrally-acting muscle relaxant used for relieving spasticities of neurological origin and muscle spasms associated with painful locomotor diseases. It is metabolized to the inactive metabolite mainly by CYP2D6 and, to a lesser extent, by CYP2C19 and CYP1A2. In our previous study, the pharmacokinetics of tolperisone was significantly affected by the genetic polymorphism of CYP2D6, but the wide interindividual variation of tolperisone pharmacokinetics was not explained by genetic polymorphism of CYP2D6 alone. Thus, we studied the effects of CYP2C19 genetic polymorphism on tolperisone pharmacokinetics. Eighty-one subjects with different CYP2C19 genotypes received a single oral dose of 150 mg tolperisone with 240 mL of water, and blood samples were collected up to 12 h after dosing. The plasma concentration of tolperisone was measured by a liquid chromatography-tandem mass spectrometry system. The CYP2C19PM group had significantly higher Cmax and lower CL/F values than the CYP2C19EM and CYP2C19IM groups. The AUCinf of the CYP2C19PM group was 2.86-fold and 3.00-fold higher than the CYP2C19EM and CYP2C19IM groups, respectively. In conclusion, the genetic polymorphism of CYP2C19 significantly affected tolperisone pharmacokinetics.


Assuntos
Tolperisona , Humanos , Tolperisona/farmacocinética , Citocromo P-450 CYP2D6/genética , Voluntários Saudáveis , Citocromo P-450 CYP2C19/genética , Genótipo , Polimorfismo Genético
20.
Blood ; 116(11): 1857-66, 2010 Sep 16.
Artigo em Inglês | MEDLINE | ID: mdl-20511539

RESUMO

The balance between survival and death in many cell types is regulated by small changes in the intracellular content of bioactive sphingolipids. Enzymes that either produce or degrade these sphingolipids control this equilibrium. The findings here described indicate that the lysosomal galactocerebrosidase (GALC) enzyme, defective in globoid cell leukodystrophy, is involved in the maintenance of a functional hematopoietic stem/progenitor cell (HSPC) niche by contributing to the control of the intracellular content of key sphingolipids. Indeed, we show that both insufficient and supraphysiologic GALC activity-by inherited genetic deficiency or forced gene expression in patients' cells and in the disease model-induce alterations of the intracellular content of the bioactive GALC downstream products ceramide and sphingosine, and thus affect HSPC survival and function and the functionality of the stem cell niche. Therefore, GALC and, possibly, other enzymes for the maintenance of niche functionality and health tightly control the concentration of these sphingolipids within HSPCs.


Assuntos
Medula Óssea/enzimologia , Galactosilceramidase/metabolismo , Células-Tronco Hematopoéticas/enzimologia , Nicho de Células-Tronco/enzimologia , Animais , Apoptose/efeitos dos fármacos , Medula Óssea/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Citometria de Fluxo , Galactosilceramidase/deficiência , Galactosilceramidase/genética , Genótipo , Transplante de Células-Tronco Hematopoéticas , Células-Tronco Hematopoéticas/metabolismo , Humanos , Imunofenotipagem , Marcação In Situ das Extremidades Cortadas , Fator de Crescimento Insulin-Like I/farmacologia , Leucodistrofia de Células Globoides/enzimologia , Leucodistrofia de Células Globoides/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Mutantes , Esfingolipídeos/metabolismo , Nicho de Células-Tronco/metabolismo , Transfecção , Células U937
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