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1.
Microorganisms ; 10(10)2022 Sep 28.
Artículo en Inglés | MEDLINE | ID: mdl-36296203

RESUMEN

Tuberculosis (TB) management is important for prompt discrimination of latent TB infection (LTBI) from active TB and proper treatment. Whole blood Interferon-gamma (IFN-γ) release assay (IGRA) is used to diagnose LTBI based on the secretion of IFN-γ by T-cells in the whole blood by using a specific antigen of Mycobacterium tuberculosis. However, the ability of IGRA to distinguish active TB from LTBI is considerably limited. Distinguishing active TB from LTBI is necessary to identify indicators that can be used to effectively manage TB and develop diagnostic methods. In the present study, we used a Luminex multiplex bead array (a bead-based antibody−antigen sandwich method). The whole blood level of acute phase proteins (APPs), such as endoglin (ENG), procalcitonin (PCT), C-reactive protein (CRP), and α1-acid glycoprotein (AGP), in active TB, LTBI, and healthy individuals were analyzed and quantified. The APP test results for the serum and whole blood samples showed that the levels of PCT, CRP, and AGP were significantly increased (p < 0.0500; area under curve = 0.955) in active TB. The level of these markers in the whole blood of active TB, LTBI, and healthy individuals could provide data for effective diagnosis and treatment of TB.

2.
J Clin Tuberc Other Mycobact Dis ; 24: 100253, 2021 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-34278005

RESUMEN

Tuberculosis (TB), which is caused by Mycobacterium tuberculosis (MTB), is a serious infectious disease with high infection and mortality rates and is a public health problem around the world. According to the World Health Organization (WHO) report, one-third of the world's population is latently infected with MTB, and 5 to 10% of those with latent TB infection (LTBI) have the potential to develop active TB once in their lifetime. Therefore, TB management for promptly distinguishing LTBI from active TB and for proper treatment is important. LTBI is currently diagnosed using the tuberculin skin test (TST) and interferon gamma (IFN-γ) release assay (IGRA). However, this test is substantially limited by its inability to distinguish active TB from LTBI. It is necessary to discover indicators that can be used for effective TB management and to develop diagnostic methods. In the present study, we used IGRA and complete blood count (CBC) analysis for discrimination of active TB, LTBI, and healthy control groups. The results showed that the number of WBC was significantly increased in the group with active TB (p < 0.0100) and level of hemoglobin (Hb) was significantly decreased (p < 0.0010) in the CBC than those of the healthy control and LTBI groups. In the WBC differential count, the number of neutrophils and monocytes were increased (p < 0.0010) in active TB group, where as those of lymphocytes were significantly decreased (p < 0.0100) in active TB group compared healthy control group. Results verified that the levels of total WBC, Hb, neutrophils, lymphocytes and monocytes were statistically significant (p < 0.0500) and the AUC was approximately 0.8613. In addition, receiver operating characteristic (ROC) curve analysis was performed to confirm the clinical usefulness between active TB and healthy control groups. In conclusion, based on these data demonstrated that the usefulness of these potential indicators for differential diagnosis, according to the result can be provided for effective diagnosis and treatment by comparing the expression patterns of the markers in the whole blood of the active TB, LTBI, and healthy control groups. Furthermore, this study needs to investigate a larger number of clinical specimens later to develop biomarkers according to the state of infection with MTB such as LTBI and active TB, as well as after treatment.

3.
Clin Orthop Surg ; 9(1): 101-108, 2017 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-28261435

RESUMEN

BACKGROUND: The purpose of this study was to analyze the radiographic and functional outcomes of flexible intramedullary (IM) nailing in adolescent patients with forearm fractures at the diaphysis or at the metadiaphyseal junction (MDJ). METHODS: We retrospectively reviewed the results of 40 patients who underwent IM nailing for pediatric forearm fractures. Thirty males and 10 females were followed for an average of 16 months (range, 12 to 20 months). Their average age was 11 years (range, 10 to 16 years). The average duration from the onset of trauma to surgery was 3.8 days (range, 1 to 36 days). Fracture sites were located at the MDJ of the radius in 8 patients (MDJ group) while 32 patients had middle-third fractures (D group). We assessed the magnitude and location of the maximum radial bow and range of movements. Functional outcomes were evaluated using Daruwalla criteria. RESULTS: Open reduction was carried out in 8 cases. Union was achieved at an average of 8.3 weeks postoperatively. The results were classified as good in 38 and excellent in 2 according to Daruwalla criteria with restoration of forearm rotation. The mean angulation at the last follow-up was 1.8° on the anteroposterior radiograph and 3.3° on the lateral radiograph (MDJ group: 1.8° and 2.1°, respectively; D group: 1.9° and 2.8°, respectively). There was no significant difference in the mean angulation between the groups. The mean magnitude of maximal radial bow was 5.7% ± 1.8% (MDJ group, 5.2% ± 0.8%; D group, 5.9% ± 1.9%). The mean location of maximal radial bow was 58.0% ± 8.8% (MDJ group, 56.4% ± 8.9%; D group, 58.6% ± 8.9%). The differences in the mean magnitude and location of maximal radial bow with the normal contralateral arms (7.0% ± 1.2% and 50.9% ± 6.0%, respectively) were not significantly different between the groups. Complications included superficial infection (2), delayed union (1), and refracture (1). CONCLUSIONS: IM nail fixation provided satisfactory results and maintained adequate stability for both forearm bone fractures in adolescents, even though the fracture was located at the MDJ of the radius.


Asunto(s)
Fijación Intramedular de Fracturas/métodos , Curación de Fractura , Fracturas del Radio/cirugía , Radio (Anatomía)/diagnóstico por imagen , Fracturas del Cúbito/cirugía , Adolescente , Niño , Diáfisis , Femenino , Antebrazo/fisiopatología , Fijación Intramedular de Fracturas/efectos adversos , Humanos , Masculino , Reducción Abierta , Radiografía , Fracturas del Radio/diagnóstico por imagen , Fracturas del Radio/fisiopatología , Estudios Retrospectivos , Rotación , Fracturas del Cúbito/diagnóstico por imagen , Fracturas del Cúbito/fisiopatología
4.
Microb Pathog ; 96: 10-4, 2016 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-27133265

RESUMEN

To investigate the expression patterns of chitinase on SDS-PAGE gel, Paenibacillus ehimensis MA2012 was incubated in gelatin-chitin medium (GCM) at 30 °C for 7 days. Six major bands (Ch3, Ch4, Ch5, Ch6, Ch7, and Ch8) of chitinase isozymes in GC medium appeared on SDS-PAGE gel during the incubation period. Chitinase activity staining of P. ehimensis MA2012 was detected on 2-DE with different pI values (4-11). After DEAE-Sephadex chromatography, eight bands (Ch1 to Ch8) of chitinase isozymes were stained strongly with Calcofluor white M2R at fraction 45. After Sephadex G-75 gel filtration, six bands (Ch3 to Ch8) of chitinase isozymes were stained with Calcofluor white M2R at fractions of 11-12. The specific activity of the purified chitinase was 3.8 units mg(-1) protein with a purification factor of 0.27. Inhibition rate of the conidial germination of Colletotrichum gloeosporioides was 87% in partial purified chitinase treatment compared with control.


Asunto(s)
Antifúngicos/metabolismo , Quitinasas/metabolismo , Colletotrichum/efectos de los fármacos , Paenibacillus/enzimología , Esporas Fúngicas/efectos de los fármacos , Antifúngicos/aislamiento & purificación , Quitinasas/aislamiento & purificación , Cromatografía en Gel , Cromatografía por Intercambio Iónico , Medios de Cultivo/química , Electroforesis en Gel Bidimensional , Electroforesis en Gel de Poliacrilamida , Perfilación de la Expresión Génica , Isoenzimas/aislamiento & purificación , Isoenzimas/metabolismo , Paenibacillus/crecimiento & desarrollo , Temperatura
5.
Clin Psychopharmacol Neurosci ; 14(1): 26-32, 2016 Feb 29.
Artículo en Inglés | MEDLINE | ID: mdl-26792037

RESUMEN

OBJECTIVE: Cognitive symptoms are an important component of depression and the Perceived Deficits Questionnaire-Depression is one of only a few instruments available for the subjective assessment of cognitive dysfunction in depression. Thus, the present study aimed to validate a Korean version of the PDQ-D (K-PDQ-D) using patients with major depressive disorder (MDD). METHODS: This study included 128 MDD patients who were assessed at study entry and 86 of these patients were then completed 12 weeks of antidepressant monotherapy. All subjects were assessed with the K-PDQ-D, the Montgomery-Asberg Depression Rating Scale (MADRS), the Sheehan Disability Scale (SDS), the EuroQol-5 dimensions questionnaire (EQ-5D), and the number of sick leave days taken in the previous week. The internal consistency, Guttman's split-half and test-retest reliabilities, factorial analyses, and concurrent and predictive validities of the K-PDQ-D were investigated. RESULTS: The K-PDQ-D exhibited excellent internal consistency and reliabilities, and was composed of four factors with high coefficients of determination. The concurrent validity analyses revealed that the K-PDQ-D scores were significantly correlated with the MADRS, SDS, and EQ-5D scores and the number of sick leave days taken. The K-PDQ-D scores at study entry significantly predicted changes in sick leave days and EQ-5D score from study entry to the 12-week endpoint. CONCLUSION: The newly developed K-PDQ-D is a reliable and valid instrument for the evaluation of subjective cognitive symptoms in MDD patients. The K-PDQ-D may assist in the gathering of unique information regarding subjective cognitive complaints, which is important for the comprehensive evaluation of patients with MDD.

6.
J Biol Chem ; 290(28): 17401-14, 2015 Jul 10.
Artículo en Inglés | MEDLINE | ID: mdl-26023233

RESUMEN

Recent groundbreaking work has demonstrated that combined expression of the transcription factors Brn2, Ascl1, and Myt1L (BAM; also known as Wernig factors) convert mouse fibroblasts into postmitotic neuronal cells. However, questions remain regarding whether trans-conversion is achieved directly or involves an intermediary precursor stage. Trans-conversion toward expandable neural precursor cells (NPCs) is more useful than direct one-step neuron formation with respect to yielding a sufficient number of cells and the feasibility of manipulating NPC differentiation toward certain neuron subtypes. Here, we show that co-expression of Wernig factors and Bcl-xL induces fibroblast conversion into NPCs (induced NPCs (iNPCs)) that are highly expandable for >100 passages. Gene expression analyses showed that the iNPCs exhibited high expression of common NPC genes but not genes specific to defined embryonic brain regions. This finding indicated that a regional identity of iNPCs was not established. Upon induction, iNPCs predominantly differentiated into astrocytes. However, the differentiation potential was not fixed and could be efficiently manipulated into general or specific subtypes of neurons by expression of additional genes. Specifically, overexpression of Nurr1 and Foxa2, transcription factors specific for midbrain dopamine neuron development, drove iNPCs to yield mature midbrain dopamine neurons equipped with presynaptic DA neuronal functions. We further assessed the therapeutic potential of iNPCs in Parkinson disease model rats.


Asunto(s)
Transdiferenciación Celular , Dopamina/metabolismo , Neuronas Dopaminérgicas/citología , Neuronas Dopaminérgicas/metabolismo , Fibroblastos/citología , Fibroblastos/metabolismo , Células-Madre Neurales/citología , Células-Madre Neurales/metabolismo , Animales , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Reprogramación Celular , Expresión Génica , Factor Nuclear 3-beta del Hepatocito/genética , Mesencéfalo/citología , Mesencéfalo/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos ICR , Ratones Transgénicos , Proteínas del Tejido Nervioso/genética , Miembro 2 del Grupo A de la Subfamilia 4 de Receptores Nucleares/genética , Factores del Dominio POU/genética , Trastornos Parkinsonianos/metabolismo , Trastornos Parkinsonianos/patología , Trastornos Parkinsonianos/terapia , Ratas , Ratas Endogámicas Lew , Ratas Wistar , Factores de Transcripción/genética
7.
Artículo en Inglés | MEDLINE | ID: mdl-24929137

RESUMEN

Doublesex and Mab-3-related transcription factor (Dmrt) gene family members have rarely been identified or characterized in aquatic invertebrates. In this study, we identified and characterized three DMdomain-containing genes - Dmrt11E, Dmrt93B, and Dmrt99B - in the monogonont rotifer, Brachionus koreanus. DMdomains of the proteins encoded by the B.koreanus Dmrt (Bk-Dmrt) genes had high similarities to DM domains of other invertebrate species. To understand the potential effects of environmental stressors on the transcriptional expression of Dmrt genes in rotifers, we exposed B.koreanus to a wide range of UV-B radiation and different concentrations of benzo[a]pyrene (B[a]P) over different time courses. Transcript levels of all Bk-Dmrt genes decreased significantly in response to relatively high doses of UV-B irradiation, and were also downregulated in response to exposure to UV-B radiation over time. Transcript levels of all Bk-Dmrt genes were downregulated in response to B[a]P exposure for 24h. This decrease in expression of all Bk-Dmrt genes was concomitant with the growth retardation induced by UV-B and B[a]P exposure. We concluded that both environmental stressors have detrimental effects on transcriptional regulation of all Bk-Dmrt genes, especially relatively high doses of these stressors, leading to growth retardation. However, further studies are required to better understand the potential role of Dmrt genes in environmental stressor-triggered growth retardation in the rotifer B.koreanus.


Asunto(s)
Benzo(a)pireno/toxicidad , Rotíferos/metabolismo , Factores de Transcripción/metabolismo , Rayos Ultravioleta , Contaminantes Químicos del Agua/toxicidad , Secuencia de Aminoácidos , Animales , Datos de Secuencia Molecular , Crecimiento Demográfico , Rotíferos/efectos de los fármacos , Rotíferos/genética , Rotíferos/efectos de la radiación , Estrés Fisiológico , Factores de Transcripción/genética , Transcripción Genética , Transcriptoma
8.
Aquat Toxicol ; 152: 232-43, 2014 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-24794342

RESUMEN

Despite being a strong toxicant for aquatic ecosystems, the effect of benzo[a]pyrene (B[a]P) on whole cytochrome P450 (CYP) biotransformation mechanisms has not been deeply investigated in aquatic organisms. To understand the mode of action of B[a]P on CYP molecular responses in fish, we analyzed the full spectrum of cyp genes and the activities of enzymes that are involved in detoxification and antioxidant defense systems after exposure to different concentrations of B[a]P over different time courses in the marine medaka, Oryzias melastigma. Upon B[a]P exposure, we found significant downregulation of cyp genes associated with steroidogenesis with decreased concentrations of actual hormones including estradiol (E2) and testosterone (11-KT), indicating that B[a]P-treated groups were closely associated with the dysfunction of hormone synthesis in a dose-dependent manner. In addition, B[a]P exposure strongly influenced transcriptional levels of antioxidant-related genes and their enzyme activities. Based on these results, we suggest that B[a]P induced the CYPs-involved systematic biotransformation mechanism with oxidative stress in the juvenile marine medaka, resulting in changes of endogenous hormonal levels and transcriptional levels of several steroidogenic metabolism-related CYPs.


Asunto(s)
Benzo(a)pireno/toxicidad , Sistema Enzimático del Citocromo P-450/metabolismo , Regulación de la Expresión Génica/efectos de los fármacos , Oryzias/fisiología , Contaminantes Químicos del Agua/toxicidad , Animales , Biotransformación/efectos de los fármacos , Hormonas/sangre , Hormonas/genética , Oryzias/genética , Estrés Oxidativo/efectos de los fármacos
9.
Aquat Toxicol ; 152: 264-72, 2014 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-24800869

RESUMEN

Nuclear radioisotope accidents are potentially ecologically devastating due to their impact on marine organisms. To examine the effects of exposure of a marine organism to radioisotopes, we irradiated the intertidal copepod Tigriopus japonicus with several doses of gamma radiation and analyzed the effects on mortality, fecundity, and molting by assessing antioxidant enzyme activities and gene expression patterns. No mortality was observed at 96h, even in response to exposure to a high dose (800Gy) of radiation, but mortality rate was significantly increased 120h (5 days) after exposure to 600 or 800Gy gamma ray radiation. We observed a dose-dependent reduction in fecundity of ovigerous females; even the group irradiated with 50Gy showed a significant reduction in fecundity, suggesting that gamma rays are likely to have a population level effect. In addition, we observed growth retardation, particularly at the nauplius stage, in individuals after gamma irradiation. In fact, nauplii irradiated with more than 200Gy, though able to molt to copepodite stage 1, did not develop into adults. Upon gamma radiation, T. japonicus showed a dose-dependent increase in reactive oxygen species (ROS) levels, the activities of several antioxidant enzymes, and expression of double-stranded DNA break damage genes (e.g. DNA-PK, Ku70, Ku80). At a low level (sub-lethal dose) of gamma irradiation, we found dose-dependent upregulation of p53, implying cellular damage in T. japonicus in response to sub-lethal doses of gamma irradiation, suggesting that T. japonicus is not susceptible to sub-lethal doses of gamma irradiation. Additionally, antioxidant genes, phase II enzyme (e.g. GSTs), and cellular chaperone genes (e.g. Hsps) that are involved in cellular defense mechanisms also showed the same expression patterns for sublethal doses of gamma irradiation (50-200Gy). These findings indicate that sublethal doses of gamma radiation can induce oxidative stress-mediated DNA damage and increase the expression of antioxidant enzymes and proteins with chaperone-related functions, thereby significantly affecting life history parameters such as fecundity and molting in the copepod T. japonicus.


Asunto(s)
Copépodos/efectos de la radiación , Daño del ADN/efectos de la radiación , Rayos gamma , Estrés Oxidativo/efectos de la radiación , Animales , Femenino , Regulación de la Expresión Génica/efectos de la radiación , Estadios del Ciclo de Vida/efectos de la radiación , Oxidorreductasas/genética , Especies Reactivas de Oxígeno/análisis , Reproducción/efectos de la radiación , Análisis de Supervivencia
10.
Aquat Toxicol ; 152: 308-17, 2014 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-24813263

RESUMEN

In this study, we investigated the effects of the water-accommodated fraction (WAF) of crude oil on the development and reproduction of the intertidal copepod Tigriopus japonicus through life-cycle experiments. Furthermore, we investigated the mechanisms underlying the toxic effects of WAF on this benthic organism by studying expression patterns of cytochrome P450 (CYP) genes. Development of T. japonicus was delayed and molting was interrupted in response to WAF exposure. Hatching rate was also significantly reduced in response to WAF exposure. Activities of antioxidant enzymes such as glutathione S-transferase (GST), glutathione reductase (GR), and catalase (CAT) were increased by WAF exposure in a concentration-dependent manner. These results indicated that WAF exposure resulted in oxidative stress, which in turn was associated with dysfunctional development and reproduction. To evaluate the involvement of cytochrome P450 (CYP) genes, we cloned the entire repertoire of CYP genes in T. japonicus (n=52) and found that the CYP genes belonged to five different clans (i.e., Clans 2, 3, 4, mitochondrial, and 20). We then examined expression patterns of these 52 CYP genes in response to WAF exposure. Three TJ-CYP genes (CYP3024A2, CYP3024A3, and CYP3027C2) belonging to CYP clan 3 were significantly induced by WAF exposure in a time- and concentration-dependent manner. We identified aryl hydrocarbon responsive elements (AhRE), xenobiotic responsive elements (XREs), and metal response elements (MRE) in the promoter regions of these three CYP genes, suggesting that these genes are involved in detoxification of toxicants. Overall, our results indicate that WAF can trigger oxidative stress and thus induce dysfunctional development and reproduction in the copepod T. japonicus. Furthermore, we identified three TJ-CYP genes that represent potential biomarkers of oil pollution.


Asunto(s)
Copépodos/efectos de los fármacos , Sistema Enzimático del Citocromo P-450/genética , Regulación de la Expresión Génica/efectos de los fármacos , Estrés Oxidativo/efectos de los fármacos , Petróleo/toxicidad , Contaminantes Químicos del Agua/toxicidad , Animales , Copépodos/clasificación , Copépodos/enzimología , Copépodos/genética , Crecimiento y Desarrollo/efectos de los fármacos , Oxidorreductasas/genética , Filogenia , Regiones Promotoras Genéticas/genética , Reproducción/efectos de los fármacos
11.
Artículo en Inglés | MEDLINE | ID: mdl-24726801

RESUMEN

To investigate the effect of endocrine disrupting chemicals (EDCs) on the circadian rhythm pathway, we cloned clock and circadian rhythmic pathway-associated genes (e.g. Per2, Cry1, Cry2, and BMAL1) in the self-fertilizing mangrove killifish Kryptolebias marmoratus. The promoter region of Km-clock had 1 aryl hydrocarbon receptor element (AhRE, GTGCGTGACA) and 8 estrogen receptor (ER) half-sites, indicating that the AhRE and ER half sites would likely be associated with regulation of clock protein activity during EDCs-induced cellular stress. The Km-clock protein domains (bHLH, PAS1, PAS2) were highly conserved in five additional fish species (zebrafish, Japanese medaka, Southern platyfish, Nile tilapia, and spotted green pufferfish), suggesting that the fish clock protein may play an important role in controlling endogenous circadian rhythms. The promoter regions of Km-BMAL1, -Cry1, -Cry2, and -Per2 were found to contain several xenobiotic response elements (XREs), indicating that EDCs may be able to alter the expression of these genes. To analyze the endogenous circadian rhythm in K. marmoratus, we measured expression of Km-clock and other circadian rhythmic genes (e.g. Per2, Cry1, Cry2, and BMAL1) in different tissues, and found ubiquitous expression, although there were different patterns of transcript amplification during different developmental stages. In an estrogen (E2)-exposed group, Km-clock expression was down-regulated, however, a hydroxytamoxifen (TMX, nonsteroid estrogen antagonist)-exposed group showed an upregulated pattern of Km-clock expression, suggesting that the expression of Km-clock is closely associated with exposure to EDCs. In response to the exposure of bisphenol A (BPA) and 4-tert-octyphenol (OP), Km-clock expression was down-regulated in the pituitary/brain, muscle, and skin in both gender types (hermaphrodite and secondary male). In juvenile K. marmoratus liver tissue, expression of Km-clock and other circadian rhythmic pathway-associated genes showed a regular oscillation pattern over a period of approximately 24h during a 12L:12D cycle. However, the circadian rhythm of BPA-exposed juvenile K. marmoratus liver tissue was perturbed over a 12L:12D period. This study will aid in our understanding of how EDCs perturb endogenous circadian rhythms, particularly in BPA-exposed fish liver tissue.


Asunto(s)
Ritmo Circadiano/efectos de los fármacos , Disruptores Endocrinos/toxicidad , Peces Killi/fisiología , Animales , Compuestos de Bencidrilo/toxicidad , Ritmo Circadiano/genética , Clonación Molecular , Trastornos del Desarrollo Sexual/genética , Femenino , Proteínas de Peces/genética , Regulación de la Expresión Génica/efectos de los fármacos , Peces Killi/genética , Hígado/efectos de los fármacos , Hígado/fisiología , Masculino , Datos de Secuencia Molecular , Fenoles/toxicidad , Filogenia , Estructura Terciaria de Proteína , Tamoxifeno/análogos & derivados , Tamoxifeno/toxicidad , Contaminantes Químicos del Agua/toxicidad
12.
Fish Shellfish Immunol ; 36(1): 240-51, 2014 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-24287371

RESUMEN

The crucian carp Carassius auratus (Cyprinidae) is one of the important fish species in aquaculture. Although the crucian carp has several economic benefits, their immune system and gene information have not been investigated in depth as yet. Here, we performed the transcriptome analysis of C. auratus using the pyrosequencing method and selected several immune-related genes. Of unigenes obtained in this species, we identified a number of immune system-related genes (e.g. adhesive protein, antimicrobial protein, apoptosis- and cell cycle-related protein, cellular defense effector, immune regulator, pattern recognition protein, protease, protease inhibitor, reduction/oxidation-related protein, signal transduction-related protein and stress protein) that are potentially useful for studies on fish immunity. To be of public and practical use, we designed primer pairs of each gene from the crucian carp for real-time RT-PCR application and tested the amplicon identity of entire gene sets with the total RNA sample. For comparative analysis, we measured tissue-preferential transcript profiles of selected genes. This study will be helpful to extend our knowledge on the immune system of the crucian carp in comparative aspects and to develop the crucian carp as a potential model organism for aquatic quality monitoring in fish farming.


Asunto(s)
Carpas/inmunología , Perfilación de la Expresión Génica/veterinaria , Ontología de Genes , Sistema Inmunológico/inmunología , Animales , Secuencia de Bases , Carpas/genética , Perfilación de la Expresión Génica/métodos , Datos de Secuencia Molecular , ARN/química , ARN/genética , Reacción en Cadena en Tiempo Real de la Polimerasa/veterinaria , Análisis de Secuencia de ADN
13.
J Sex Med ; 10(11): 2832-41, 2013 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-23937271

RESUMEN

INTRODUCTION: There is partial evidence to support the use of phophodiesterase-5 inhibitor (PDE5-I) for the treatment of premature ejaculation (PE). AIM: We compared on-demand dosing of dapoxetine alone and combined with mirodenafil in subjects with lifelong PE and without erectile dysfunction (ED). METHODS: Our prospective, randomized, double-blind, placebo-controlled, multicenter trial enrolled 118 subjects with lifelong PE without ED. PE was diagnosed using Diagnostic and Statistical Manual of Mental Disorders, fourth edition, text revision. Patients were divided into two groups: dapoxetine 30 mg plus placebo (group A, n=56) and dapoxetine 30 mg plus mirodenafil 50 mg (group B, n=62). MAIN OUTCOME MEASURES: During 12 weeks, intravaginal ejaculatory latency time (IELT) and the time from foreplay to beginning intercourse (FTIT) with a stopwatch, and Premature Ejaculation Profile (PEP) were measured. Overall sexual act time (OSAT; sum of FTIT and IELT) was calculated. Any treatment-emergent adverse events (TEAEs) were also recorded. RESULTS: Over 12 weeks, IELT, OSAT, and PEP index score significantly improved in group B compared with group A (increased geometric mean IELT in group A and B=3.6 and 6.1 minutes, P=0.026; increased geometric mean OSAT in group A and B=5.5 and 9.9 minutes, P=0.012; increased median PEP index score in group A and B=1.0 and 1.3, P=0.046). However, there was no significant difference between two groups with respect to improvement of FTIT (P=0.147). TEAEs did not differ between groups (all P>0.05), and there was no serious adverse event in any subjects. CONCLUSIONS: Low dose of dapoxetine combined with mirodenafil showed better results in terms of IELT, OSAT, and PEP index score, and similar TEAEs, compared with that of dapoxetine only. Our results support the suggestion that the PDE5-Is have a potential role in the treatment of PE without ED.


Asunto(s)
Bencilaminas/administración & dosificación , Naftalenos/administración & dosificación , Inhibidores de Fosfodiesterasa 5/administración & dosificación , Eyaculación Prematura/tratamiento farmacológico , Pirimidinonas/administración & dosificación , Inhibidores Selectivos de la Recaptación de Serotonina/administración & dosificación , Sulfonamidas/administración & dosificación , Adulto , Anciano , Método Doble Ciego , Humanos , Masculino , Persona de Mediana Edad , Placebos , Eyaculación Prematura/fisiopatología , Eyaculación Prematura/psicología , Estudios Prospectivos , Resultado del Tratamiento
14.
Mol Biosyst ; 8(2): 602-8, 2012 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-22048332

RESUMEN

A culture medium provides the major environmental conditions for cells in vitro. Replenishment of a culture medium causes an abrupt change in the extracellular environment for maintaining cells in a certain state. As a primitive form of a complex system, a stem cell is likely to be influenced by culture conditions that can change the destination of development. To understand how the change in extracellular environment can influence a biological system, we studied the effect of culture media replacement on the gene expression of differentiating neural progenitor cells. From time-series microarray gene expression data of neural progenitor cells, we observed a periodic wave that was synchronized with intermittent culture media replacement. We identified three modes that mostly contribute to the periodic patterns in gene expression and investigated mode-related genes that are sensitive to the changes in the extracellular environment. The biological significance of the three modes was explored, such as progressive development and cell fate decision, extracellular matrix reassembly, and cell growth regulation in response to stress. In addition, we explored systemic influences of media replacement on differentiating neural progenitor cells. Intermittent culture media replacement interrupts expression of genes that participate in the major processes of differentiating neural progenitor cells. This study shows how the abrupt changes in the cell environment influence gene expression systematically.


Asunto(s)
Diferenciación Celular/efectos de los fármacos , Medios de Cultivo/farmacología , Células-Madre Neurales/efectos de los fármacos , Técnicas de Cultivo de Célula/métodos , Proliferación Celular , Células Cultivadas , Matriz Extracelular/metabolismo , Expresión Génica , Humanos , Células-Madre Neurales/citología , Células-Madre Neurales/metabolismo , Análisis de Secuencia por Matrices de Oligonucleótidos , Estrés Fisiológico
15.
Stem Cells ; 29(11): 1861-73, 2011 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-21922608

RESUMEN

Understanding midbrain dopamine (DA) neuron differentiation is of importance, because of physiological and clinical implications of this neuronal subtype. We show that prolonged membrane depolarization induced by KCl treatment promotes DA neuron differentiation from neural precursor cells (NPCs) derived from embryonic ventral midbrain (VM). Interestingly, the depolarization-induced increase of DA neuron yields was not abolished by L-type calcium channel blockers, along with no depolarization-mediated change of intracellular calcium level in the VM-derived NPCs (VM-NPCs), suggesting that the depolarization effect is due to a calcium-independent mechanism. Experiments with labeled DA neuron progenitors indicate that membrane depolarization acts at the differentiation fate determination stage and promotes the expression of DA phenotype genes (tyrosine hydroxylase [TH] and DA transporter [DAT]). Recruitment of Nurr1, a transcription factor crucial for midbrain DA neuron development, to the promoter of TH gene was enhanced by depolarization, along with increases of histone 3 acetylation (H3Ac) and trimethylation of histone3 on lysine 4 (H3K4m3), and decreases of H3K9m3 and H3K27m3 in the consensus Nurr1 binding regions of TH promoter. Depolarization stimuli on differentiating VM-NPCs also induced dissociation of methyl CpG binding protein 2 and related repressor complex molecules (repressor element-1 silencing transcription factor corepressor and histone deacetylase 1) from the CpG sites of TH and DAT promoters. Based on these findings, we suggest that membrane depolarization promotes DA neuron differentiation by opening chromatin structures surrounding DA phenotype genes and inhibiting the binding of corepressors, thus allowing transcriptional activators such as Nurr1 to access DA neuron differentiation gene promoter regions.


Asunto(s)
Diferenciación Celular/fisiología , Neuronas Dopaminérgicas/citología , Neuronas Dopaminérgicas/metabolismo , Histonas/metabolismo , Mesencéfalo/citología , Células-Madre Neurales/citología , Células-Madre Neurales/metabolismo , Animales , Calcio/metabolismo , Diferenciación Celular/genética , Proliferación Celular , Supervivencia Celular/genética , Supervivencia Celular/fisiología , Células Cultivadas , Metilación de ADN/genética , Epigénesis Genética/genética , Histonas/genética , Inmunohistoquímica , Proteína 2 de Unión a Metil-CpG/genética , Miembro 2 del Grupo A de la Subfamilia 4 de Receptores Nucleares/genética , Ratas , Reacción en Cadena en Tiempo Real de la Polimerasa
16.
J Clin Invest ; 121(6): 2326-35, 2011 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-21576821

RESUMEN

Parkinson disease (PD) involves the selective loss of midbrain dopamine (mDA) neurons and is a possible target disease for stem cell-based therapy. Human induced pluripotent stem cells (hiPSCs) are a potentially unlimited source of patient-specific cells for transplantation. However, it is critical to evaluate the safety of hiPSCs generated by different reprogramming methods. Here, we compared multiple hiPSC lines derived by virus- and protein-based reprogramming to human ES cells (hESCs). Neuronal precursor cells (NPCs) and dopamine (DA) neurons delivered from lentivirus-based hiPSCs exhibited residual expression of exogenous reprogramming genes, but those cells derived from retrovirus- and protein-based hiPSCs did not. Furthermore, NPCs derived from virus-based hiPSCs exhibited early senescence and apoptotic cell death during passaging, which was preceded by abrupt induction of p53. In contrast, NPCs derived from hESCs and protein-based hiPSCs were highly expandable without senescence. DA neurons derived from protein-based hiPSCs exhibited gene expression, physiological, and electrophysiological properties similar to those of mDA neurons. Transplantation of these cells into rats with striatal lesions, a model of PD, significantly rescued motor deficits. These data support the clinical potential of protein-based hiPSCs for personalized cell therapy of PD.


Asunto(s)
Reprogramación Celular , Dopamina/metabolismo , Células Madre Pluripotentes Inducidas/fisiología , Factores de Transcripción de Tipo Kruppel/fisiología , Neuronas/citología , Factor 3 de Transcripción de Unión a Octámeros/fisiología , Trastornos Parkinsonianos/cirugía , Proteínas Proto-Oncogénicas c-myc/fisiología , Factores de Transcripción SOXB1/fisiología , Animales , Apoptosis , Arginina , Diferenciación Celular , Línea Celular/trasplante , Linaje de la Célula , Senescencia Celular , Regulación del Desarrollo de la Expresión Génica , Genes p53 , Vectores Genéticos/farmacología , Humanos , Células Madre Pluripotentes Inducidas/trasplante , Factor 4 Similar a Kruppel , Factores de Transcripción de Tipo Kruppel/genética , Lentivirus/fisiología , Neuronas/metabolismo , Factor 3 de Transcripción de Unión a Octámeros/genética , Proteínas Proto-Oncogénicas c-myc/genética , Ratas , Retroviridae/fisiología , Factores de Transcripción SOXB1/genética , Proteína p53 Supresora de Tumor/biosíntesis
17.
Stem Cells ; 28(3): 501-12, 2010 Mar 31.
Artículo en Inglés | MEDLINE | ID: mdl-20049900

RESUMEN

Effective dopamine (DA) neuron differentiation from neural precursor cells (NPCs) is prerequisite for precursor/stem cell-based therapy of Parkinson's disease (PD). Nurr1, an orphan nuclear receptor, has been reported as a transcription factor that can drive DA neuron differentiation from non-dopaminergic NPCs in vitro. However, Nurr1 alone neither induces full neuronal maturation nor expression of proteins found specifically in midbrain DA neurons. In addition, Nurr1 expression is inefficient in inducing DA phenotype expression in NPCs derived from certain species such as mouse and human. We show here that Foxa2, a forkhead transcription factor whose role in midbrain DA neuron development was recently revealed, synergistically cooperates with Nurr1 to induce DA phenotype acquisition, midbrain-specific gene expression, and neuronal maturation. Thus, the combinatorial expression of Nurr1 and Foxa2 in NPCs efficiently yielded fully differentiated nigral (A9)-type midbrain neurons with clearly detectable DA neuronal activities. The effects of Foxa2 in DA neuron generation were observed regardless of the brain regions or species from which NPCs were derived. Furthermore, DA neurons generated by ectopic Foxa2 expression were more resistant to toxins. Importantly, Foxa2 expression resulted in a rapid cell cycle exit and reduced cell proliferation. Consistently, transplantation of NPCs transduced with Nurr1 and Foxa2 generated grafts enriched with midbrain-type DA neurons but reduced number of proliferating cells, and significantly reversed motor deficits in a rat PD model. Our findings can be applied to ongoing attempts to develop an efficient and safe precursor/stem cell-based therapy for PD.


Asunto(s)
Diferenciación Celular/genética , Factor Nuclear 3-beta del Hepatocito/genética , Neuronas/metabolismo , Miembro 2 del Grupo A de la Subfamilia 4 de Receptores Nucleares/genética , Trasplante de Células Madre/métodos , Células Madre/metabolismo , Animales , Proliferación Celular , Supervivencia Celular/genética , Tratamiento Basado en Trasplante de Células y Tejidos/métodos , Células Cultivadas , Dopamina/metabolismo , Humanos , Ratones , Neurogénesis/genética , Neuronas/citología , Neuronas/trasplante , Enfermedad de Parkinson/cirugía , Fenotipo , Ratas , Ratas Sprague-Dawley , Células Madre/citología , Sustancia Negra/citología , Sustancia Negra/metabolismo , Transfección/métodos , Resultado del Tratamiento
18.
Mol Ther ; 17(10): 1761-70, 2009 Oct.
Artículo en Inglés | MEDLINE | ID: mdl-19603007

RESUMEN

We have previously demonstrated derivation of neural precursor (NP) cells of a midbrain-type from human embryonic stem (hES) cells to yield an enriched population of dopamine (DA) neurons. These hES-derived NPs can be expanded in vitro through multiple passages without altering their DA neurogenic potential. Here, we studied two aspects of these hES-NP cells that are critical issues in cell therapeutic approaches for Parkinson's disease (PD): cell survival and tumorigenic potential. Neuroepithelial rosettes, a potentially tumorigenic structure, disappeared during hES-NP cell expansion in vitro. Although a minor population of cells positive for Oct3/4, a marker specific for undifferentiated hES cells, persisted in culture during hES-NP cell expansion, they could be completely eliminated by subculturing hES-NPs under differentiation-inducing conditions. Consistently, no tumors/teratomas are formed in rats grafted with multipassaged hES-NPs. However, extensively expanded hES-NP cells easily underwent cell death during differentiation in vitro and after transplantation in vivo. Transgenic expression of Bcl-XL and sonic hedgehog (SHH) completely overcame the cell survival problems without increasing tumor formation. These findings indicate that hES-NP cell expansion in conjunction with Bcl-XL+SHH transgene expression may provide a renewable and safe source of DA neurons for transplantation in PD.


Asunto(s)
Dopamina/metabolismo , Células Madre Embrionarias/citología , Neuronas/citología , Animales , Apoptosis/efectos de los fármacos , Factor Neurotrófico Derivado del Encéfalo/farmacología , Diferenciación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , AMP Cíclico/farmacología , Células Madre Embrionarias/efectos de los fármacos , Femenino , Vectores Genéticos/genética , Factor Neurotrófico Derivado de la Línea Celular Glial/farmacología , Proteínas Hedgehog/genética , Proteínas Hedgehog/fisiología , Humanos , Inmunohistoquímica , Neuronas/efectos de los fármacos , Reacción en Cadena de la Polimerasa , Ratas , Ratas Sprague-Dawley , Retroviridae/genética , Proteína bcl-X/genética , Proteína bcl-X/fisiología
19.
Stem Cells ; 27(9): 2238-46, 2009 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-19522012

RESUMEN

Nurr1 is a transcription factor specific for the development and maintenance of the midbrain dopamine (DA) neurons. Exogenous Nurr1 in neural precursor (NP) cells induces the differentiation of DA neurons in vitro that are capable of reversing motor dysfunctions in a rodent model for Parkinson disease. The promise of this therapeutic approach, however, is unclear due to poor cell survival and phenotype loss of DA cells after transplantation. We herein demonstrate that Nurr1 proteins undergo ubiquitin-proteasome-system-mediated degradation in differentiating NP cells. The degradation process is activated by a direct Akt-mediated phosphorylation of Nurr1 proteins and can be prevented by abolishing the Akt-target sequence in Nurr1 (Nurr1(Akt)). Overexpression of Nurr1(Akt) in NP cells yielded DA neurons in which Nurr1 protein levels were maintained for prolonged periods. The sustained Nurr1 expression endowed the Nurr1(Akt)-induced DA neurons with resistance to toxic stimuli, enhanced survival, and sustained DA phenotypes in vitro and in vivo after transplantation.


Asunto(s)
Dopamina/metabolismo , Neuronas/citología , Neuronas/metabolismo , Miembro 2 del Grupo A de la Subfamilia 4 de Receptores Nucleares/fisiología , Animales , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/farmacología , Western Blotting , Butadienos/farmacología , Proteínas Quinasas Dependientes de Calcio-Calmodulina/antagonistas & inhibidores , Proteínas Quinasas Dependientes de Calcio-Calmodulina/fisiología , Diferenciación Celular/genética , Diferenciación Celular/fisiología , Línea Celular , Supervivencia Celular/genética , Supervivencia Celular/fisiología , Cromonas/farmacología , Inhibidores Enzimáticos/farmacología , Flavonoides/farmacología , Humanos , Inmunoprecipitación , Mesencéfalo/citología , Morfolinas/farmacología , Nitrilos/farmacología , Miembro 2 del Grupo A de la Subfamilia 4 de Receptores Nucleares/genética , Fosfatidilinositol 3-Quinasas/fisiología , Inhibidores de las Quinasa Fosfoinosítidos-3 , Estabilidad Proteica/efectos de los fármacos , Ratas , Ratas Sprague-Dawley , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa
20.
Biosystems ; 95(1): 17-25, 2009 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-18640237

RESUMEN

A time-series microarray experiment is useful to study the changes in the expression of a large number of genes over time. Many methods for clustering genes using gene expression profiles have been suggested, but it is not easy to interpret the biological significance of the results or utilize these methods for understanding the dynamics of gene regulatory systems. In this study, we introduce an algorithm for readjusting the boundaries of clusters by adopting the advantages of both k-means and singular value decomposition (SVD). In addition, we suggest a methodology for searching the principal genes that can be the most crucial genes in regulation of clusters. We found 34 principal genes from 171 clusters having strong concentratedness in their expression patterns and distinct ranges of oscillatory phases, by using a time-series microarray dataset of mouse embryonic stem (ES) cells after induction of dopaminergic neural differentiation. The biological significance of the principal genes examined in the literature supports the feasibility of our algorithms in that the hierarchy of clusters may lead the manifestation of the phenotypes, e.g., the development of the nervous system.


Asunto(s)
Diferenciación Celular/genética , Células Madre Embrionarias/citología , Familia de Multigenes , Sistema Nervioso/citología , Algoritmos , Animales , Ratones
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