RESUMO
PURPOSE: To evaluate the efficacy of the modified superior inverted internal limiting (ILM) membrane flap technique in retinal reattachment, macular hole closure and external retinal layers restoration in macular hole associated retinal detachment compared to ILM peeling. METHODS: Retrospective case series of 10 patients that required pars plana vitrectomy for retinal detachment with macular hole followed for more than 12 months. Data from medical records were retrospectively collected and patients were divided into the superior inverted flap (5 patients) and ILM peeling group (5 patients). We compared best corrected visual acuity (BCVA) before and after surgery, retinal attachment, macular hole closure rate and external retinal layer restoration between groups. RESULTS: There were significant improvements in BCVA in both groups before and after surgery, with no differences between the two groups at 12 months after surgery (p=0.9). The macular hole closed in 100% of cases in the inverted flap group and 80% of the ILM peeling group with no significant differences between groups. The retina was reattached in 100% of cases in both groups. Only 2 patients in the inverted flap group (40%) had external retinal layer restoration and none in ILM peeling group (p=0.62). CONCLUSIONS: ILM peeling and superior inverted flap techniques are useful for treating retinal detachment with macular hole in myopic eyes.
Assuntos
Membrana Epirretiniana , Miopia , Descolamento Retiniano , Perfurações Retinianas , Membrana Epirretiniana/cirurgia , Humanos , Miopia/cirurgia , Retina , Descolamento Retiniano/cirurgia , Perfurações Retinianas/cirurgia , Estudos Retrospectivos , Acuidade Visual , Vitrectomia/métodosRESUMO
Plants have evolved different mechanisms to increase their tolerance to aluminum (Al) toxicity and low pH in the soil. The Zn finger transcription factor SENSITIVE TO PROTON RHIZOTOXICITY1 (STOP1) plays an essential role in the adaptation of plants to Al and low pH stresses. In this work, we isolated the ScSTOP1 gene from rye (Secale cereale L.), which is located on chromosome 3RS. The ectopic expression of ScSTOP1 complements the Arabidopsis stop1 mutation in terms of root growth inhibition due to Al and pH stress, as well as phosphate starvation tolerance, suggesting that rye ScSTOP1 is a functional ortholog of AtSTOP1. A putative STOP1 binding motif was identified in the promoter of a well-known STOP1 target from rye and Arabidopsis and was later corroborated by genomic DAP-seq analyses. Coexpression analyses verified that ScSTOP1 activated the promoter of ScALMT1. We have also identified a putative phosphorylatable serine in STOP1 that is phylogenetically conserved and critical for such activation. Our data indicated that ScSTOP1 also regulated Al and pH tolerance in rye.
Assuntos
Alumínio/toxicidade , Proteínas de Arabidopsis/metabolismo , Arabidopsis/efeitos dos fármacos , Arabidopsis/metabolismo , Transportadores de Ânions Orgânicos/metabolismo , Secale/metabolismo , Proteínas de Arabidopsis/genética , Cromossomos de Plantas/genética , Regulação da Expressão Gênica de Plantas/efeitos dos fármacos , Regulação da Expressão Gênica de Plantas/genética , Concentração de Íons de Hidrogênio , Mutação/genética , Transportadores de Ânions Orgânicos/genética , Regiões Promotoras Genéticas/efeitos dos fármacos , Regiões Promotoras Genéticas/genética , Secale/efeitos dos fármacos , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
After fertilization, microtubule (MT) sperm asters undergo long-range migration to accurately position pronuclei. Due to the large sizes of zygotes, the forces driving aster migration are considered to be from pulling on the astral MTs by dynein, with no significant contribution from pushing forces. Here, we re-investigate the forces responsible for sperm aster centration in sea urchin zygotes. Our quantifications of aster geometry and MT density preclude a pulling mechanism. Manipulation of aster radial lengths and growth rates, combined with quantitative tracking of aster migration dynamics, indicates that aster migration is equal to the length of rear aster radii, supporting a pushing model for centration. We find that dynein inhibition causes an increase in aster migration rates. Finally, ablation of rear astral MTs halts migration, whereas front and side ablations do not. Collectively, our data indicate that a pushing mechanism can drive the migration of asters in a large cell type.
Assuntos
Núcleo Celular/metabolismo , Microtúbulos/fisiologia , Animais , Ouriços-do-MarRESUMO
Posterior microphthalmos is a rare condition that can be found in paediatric patients with increased farsightedness and reduced vision. A retrospective study is presented of 5cases of posterior microphthalmia aged between 4 and 13 years. The following parameters were obtained: visual acuity, cycloplegic refractive error, optical biometry, slit lamp examination, intraocular pressure, and ocular ultrasound. The refraction, axial length and average visual acuity was+15.35 Dp, 16.20mm and 0.13, respectively. The fundus was examined, optical coherence tomography was performed, and also retinography and fluorescein angiography in one case. In all cases, the absence of foveal depression and different morphotypes of the papillo-macular fold were observed in the tomography. In the absence of a specific treatment, the appropriate detection, management, and monitoring of this disease is important to improve and maintain the vision of patients and recognise possible complications.
Assuntos
Microftalmia/diagnóstico por imagem , Tomografia de Coerência Óptica , Adolescente , Criança , Pré-Escolar , Humanos , Lactente , Microftalmia/diagnóstico , Estudos RetrospectivosAssuntos
Oftalmopatias Hereditárias/diagnóstico , Macula Lutea/patologia , Degeneração Retiniana/diagnóstico , Doenças Retinianas/diagnóstico , Idoso , Oftalmopatias Hereditárias/patologia , Feminino , Angiofluoresceinografia , Fundo de Olho , Humanos , Macula Lutea/diagnóstico por imagem , Degeneração Retiniana/patologia , Doenças Retinianas/etiologia , Doenças Retinianas/patologia , Baixa Visão/diagnóstico , Baixa Visão/etiologiaRESUMO
OBJECTIVE: To study elevated postoperative intraocular pressure (PIOP) after paediatric cataract surgery in a reference centre and to compare the results obtained with the most relevant world literature. MATERIALS AND METHODS: A retrospective observational study was conducted, collecting information from clinical records of 99 patients (142 eyes) operated on due to unilateral or bilateral paediatric cataracts between 2000 and 2008. RESULTS: The proportion of eyes with PIOP≥20mmHg and bilateral cataracts was higher, 79.3%, than the proportion of eyes with PIOP≥20mmHg and a unilateral cataract, 20.7% (P=.032). The proportion of eyes with PIOP≥20mmHg between the eyes with cataract surgery before one month of age was higher, 50%, than between the eyes in which the cataract surgery was performed after one month of age, 17.2% (P=.009). PIOP values≥20mmHg were associated with a lower best corrected visual acuity at 5 years after cataract surgery (P=.020). The proportion of eyes with PIOP≥20mmHg and without intraocular lenses was higher, 69%, than the proportion of eyes with PIOP≥20mmHg and intraocular lenses, 31% (P<.001). CONCLUSIONS: A PIOP>20mmHg was observed in 20.4% of eyes in a 5-year follow-up period, appearing more frequently in eyes with bilateral cataract than unilateral, and in aphakic eyes more than pseudophakic eyes. Elevation of the PIOP was associated with the performance of cataract surgery at early ages.
Assuntos
Extração de Catarata/efeitos adversos , Catarata/congênito , Hipertensão Ocular/etiologia , Complicações Pós-Operatórias/etiologia , Fatores Etários , Catarata/patologia , Extração de Catarata/métodos , Extração de Catarata/estatística & dados numéricos , Pré-Escolar , Humanos , Lactente , Recém-Nascido , Pressão Intraocular , Implante de Lente Intraocular , Microftalmia/complicações , Hipertensão Ocular/diagnóstico , Estudos Retrospectivos , Acuidade VisualRESUMO
BACKGROUND: Regarding survival and quality of life recent mass casualty incidents again emphasize the importance of early identification of the correct degree of injury/illness to enable prioritization of treatment amongst patients and their transportation to an appropriate hospital. The present study investigated existing triage algorithms in terms of sensitivity (SE) and specificity (SP) as well as its process duration in a relevant emergency patient cohort. METHODS: In this study 500 consecutive air rescue missions were evaluated by means of standardized patient records. Classification of patients was accomplished by 19 emergency physicians. Every case was independently classified by at least 3 physicians without considering any triage algorithm. Existing triage algorithms Primary Ranking for Initial Orientation in Emergency Medical Services (PRIOR), modified Simple Triage and Rapid Treatment (mSTaRT), Field Triage Score (FTS), Amberg-Schwandorf Algorithm for Triage (ASAV), Simple Triage and Rapid Treatment (STaRT), Care Flight, and Triage Sieve were additionally carried out computer based on each case, to enable calculation of quality criteria. RESULTS: The analyzed cohort had an age of (mean ± SD) 59 ± 25 years, a NACA score of 3.5 ± 1.1 and consisted of 57% men. On arrival 8 patients were deceased. Consequently, 492 patients were included in the analysis. The distribution of triage categories T1/T2/T3 were 10%/47%/43%, respectively. The highest diagnostic quality was achieved with START, mSTaRT, and ASAV yielding a SE of 78% and a SP ranging from 80-83%. The subgroup of surgical patients reached a SE of 95% and a SP between 85-91%. The newly established algorithm PRIOR exerted a SE of 90% but merely a SP of 54% in the overall cohort thereby consuming the longest time for overall decision. CONCLUSION: Triage procedures with acceptable diagnostic quality exist to identify the most severely injured. Due to its high rate of false positive results (over-triage) the recently developed PRIOR algorithm will cause overload of available resources for the severely injured within mass casualty incident missions. Non-surgical patients still are poorly identified by the available algorithms.
Assuntos
Algoritmos , Incidentes com Feridos em Massa/estatística & dados numéricos , Triagem/métodos , Adulto , Idoso , Idoso de 80 Anos ou mais , Estudos de Coortes , Planejamento em Desastres/métodos , Serviços Médicos de Emergência , Serviço Hospitalar de Emergência , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Médicos , Qualidade de VidaRESUMO
BACKGROUND: Regarding survival and quality of life, recent mass casualty incidents have once more emphasized the importance of early identification of the correct degree of injury or illness, to enable prioritizing treatment of patients and transportation to an appropriate hospital. The present study investigated international triage algorithms in terms of sensitivity (SE) and specificity (SP) as well as the process duration in a relevant emergency patient cohort. METHODS: A total of 500 consecutive air rescue missions were evaluated by means of standardized patient records. Interdisciplinary classification of patients was accomplished by 19 emergency physicians. Every case was independently classified according to the triage category by at least three physicians without considering any triage algorithm. The available triage algorithms PRIOR (Primary Ranking for Initial Orientation in Emergency Medical Services), mSTaRT (modified Simple Triage and Rapid Treatment), FTS (Field Triage Score), ASAV (Amberg-Schwandorf Algorithm for Triage), STaRT (Simple Triage and Rapid Treatment), CareFlight triage and Triage Sieve were additionally carried out for each patient in a computer-based procedure, to enable calculation of test quality criteria for all procedures. RESULTS: The analyzed cohort had a mean age of 59 ± 25 years (±SD), a National Advisory Committee for Aeronautics (NACA) score of 3.5 ± 1.1 and consisted of 57% men. On arrival 8 patients were already deceased, consequently 492 patients were included in the analysis. The distributions of triage categories I/II/III were 10%/47%/43%, respectively. The highest diagnostic quality was achieved with START, mSTaRT, and ASAV with 78% SE and 80-83% SP. The subgroup of surgical patients achieved 95% SE and 85-91% SP. The newly established algorithm PRIOR exerted an SE of 90% but an SP of only 54% in the overall cohort thereby taking the longest overall time for decisions. CONCLUSION: Triage procedures with acceptable diagnostic quality exist to identify the most severely injured. Due to its high rate of false positive results (overtriage) in this study, the recently developed PRIOR algorithm could result in exhaustion of available resources for the severely injured and therefore to undertreatment of correctly assigned triage category I cases within mass casualty incidents. Non-surgical patients are still poorly allocated by the available algorithms. Contribution available free of charge by "Free Access".
Assuntos
Algoritmos , Incidentes com Feridos em Massa/estatística & dados numéricos , Triagem/métodos , Adulto , Idoso , Idoso de 80 Anos ou mais , Defesa Civil , Tomada de Decisão Clínica , Estudos de Coortes , Planejamento em Desastres , Serviços Médicos de Emergência , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Médicos , Valor Preditivo dos Testes , Reprodutibilidade dos Testes , Trabalho de Resgate , Triagem/estatística & dados numéricosRESUMO
OBJECTIVE: To determine the prevalence of five physical frailty phenotype components and to assess the relationship between them and other clinical factors. METHOD: A population-based cross-sectional study was performed. Subjects 75 years and older were randomly selected from primary care databases (with sampling stratified by gender). Physical frailty phenotypes were assessed using Fried's criteria. Sociodemographic data, comorbidities, nutritional status, and functional capacity were assessed. RESULTS: 126 subjects were recruited (47% women). Prevalence rates were poor muscle strength: 50%; low physical activity: 29%; slow gait: 28%; exhaustion: 27%; and weight loss: 5%. Prefrailty and frailty prevalence rates were 35.7% and 29.4%, respectively. Poor muscle strength and low physical activity showed a close relationship and concordance (kappa = 0.92). Most frailty components were associated with outdoor activity, hours walked daily, and certain comorbidities. CONCLUSIONS: Poor muscle strength was the most prevalent frailty component and was closely associated with physical activity, suggesting that training programs may revert or prevent the frailty process.
Assuntos
Exercício Físico/fisiologia , Idoso Fragilizado , Força Muscular/fisiologia , Atividades Cotidianas , Idoso , Idoso de 80 Anos ou mais , Comorbidade , Estudos Transversais , Fadiga/fisiopatologia , Feminino , Marcha/fisiologia , Humanos , Vida Independente , Masculino , Estado Nutricional , Redução de PesoRESUMO
Municipal water disinfection systems in some areas are not always able to meet water consumer needs, such as ensuring distributed water quality, because household water management can be a contributing factor in water re-contamination. This fact is related to the storage options that are common in places where water is scarce or is distributed over limited time periods. The aim of this study is to assess the removal capacity of a multiple-barrier water disinfection device for protozoa, bacteria, and viruses. Water samples were taken from households in Mexico City and spiked with a known amount of protozoa (Giardia cyst, Cryptosporidium oocyst), bacteria (Escherichia coli), and viruses (rotavirus, adenovirus, F-specific ribonucleic acid (FRNA) coliphage). Each inoculated sample was processed through a multiple-barrier device. The efficiency of the multiple-barrier device to remove E. coli was close to 100%, and more than 87% of Cryptosporidium oocysts and more than 98% of Giardia cysts were removed. Close to 100% of coliphages were removed, 99.6% of the adenovirus was removed, and the rotavirus was almost totally removed. An effect of site by zone was detected; this observation is important because the water characteristics could indicate the efficiency of the multiple-barrier disinfection device.
Assuntos
Água Potável , Microbiologia da Água , Purificação da Água/instrumentação , Adenoviridae/isolamento & purificação , Colífagos/isolamento & purificação , Cryptosporidium parvum/isolamento & purificação , Desenho de Equipamento , Escherichia coli/isolamento & purificação , Giardia lamblia/isolamento & purificação , Humanos , México , Rotavirus/isolamento & purificaçãoRESUMO
INTRODUCTION: The retinopathy of prematurity (ROP) is a potentially avoidable cause of blindness in children. The advances in neonatal care make the survival of extremely premature infants, who show a greater incidence of the disease, possible. The aim of the study is to evaluate the reliability of ROP screening using retinography imaging with the RetCam 3 wide-angle camera and also study the variability of ROP diagnosis depending on the evaluator. MATERIAL AND METHODS: The indirect ophthalmoscopy exam was performed by a Pediatric ROP-Expert Ophthalmologist. The same ophthalmologist and a technician specialized in digital image capture took retinal images using the RetCam 3 wide-angle camera. A total of 30 image sets were analyzed by 3 masked groups: group A (8 ophthalmologists), group B (5 experts in vision), and group C (2 ROP-expert ophthalmologists). RESULTS: According to the diagnosis using indirect ophthalmoscopy, the sensitivity (26-93), Kappa (0.24-0.80), and the percent agreement were statistically significant in group C for the diagnosis of ROP Type 1. In the diagnosis of ROP Type 1+Type 2, Kappa (0.17-0.33) and the percent agreement (58-90) were statistically significant, with higher values in group C. CONCLUSION: The diagnosis, carried out by ROP-expert ophthalmologists, using the wide-angle camera RetCam 3 has proved to be a reliable method.
Assuntos
Oftalmoscopia , Retinopatia da Prematuridade/diagnóstico , Humanos , Recém-Nascido , Variações Dependentes do Observador , Reprodutibilidade dos TestesRESUMO
Inflammation originating from infection of the vitreous cavity is called endophthalmitis. Attention has been focused on the epidemiologic, microbiologic reports, and treatment options; unfortunately, the role of the host immune reaction in the visual function damage is still not well understood. Endophthalmitis occurs most frequently after cataract surgery. In this paper we review the published literature regarding inflammatory mediators and apoptosis during the course of endophthalmitis. Toll-like receptors, cytokines, high-mobility group box 1 proteins, aB-crystallin and apoptosis have been studied during clinical and experimental cases of endophthalmitis. Further understanding of the host-immune reaction to vitreous infection is essential for the development of new therapies. The use of intravitreal antibiotics and corticosteroids, vitrectomy and systemic antibiotics for the preservation of visual function is still discouraging.
Assuntos
Endoftalmite/imunologia , Inflamação/metabolismo , Animais , Apoptose/efeitos dos fármacos , Endoftalmite/tratamento farmacológico , Endoftalmite/metabolismo , Endoftalmite/fisiopatologia , Humanos , Inflamação/tratamento farmacológicoRESUMO
CLINICAL CASE: A fair-skinned woman presented marked striate melanokeratosis in her left eye related to recurrent corneal erosion. The source of pigmentation was a conjunctival melanosis. The conjunctival melanosis responded to treatment with topical mitomycin, while the corneal pigmentation persisted. DISCUSSION: Striate melanokeratosis is a condition described in dark-skinned patients who show a well-defined pigmentation of the limbal area, with only one case of striate melanokeratosis reported previously in a Caucasian person. The stimuli for this proliferation are corneal lesions or melanosis close to the limbus. Avoiding both stimuli are the main steps in its management.
Assuntos
Doenças da Túnica Conjuntiva/etiologia , Doenças da Córnea/etiologia , Úlcera da Córnea/complicações , Limbo da Córnea/patologia , Melanose/etiologia , Doenças da Túnica Conjuntiva/tratamento farmacológico , Doenças da Túnica Conjuntiva/patologia , Doenças da Córnea/tratamento farmacológico , Doenças da Córnea/patologia , Cor de Olho , Feminino , Fluormetolona/administração & dosagem , Fluormetolona/uso terapêutico , Humanos , Melanose/tratamento farmacológico , Melanose/patologia , Pessoa de Meia-Idade , Mitomicina/administração & dosagem , Mitomicina/uso terapêutico , Soluções Oftálmicas , Recidiva , População BrancaRESUMO
CASE REPORT: A 56-year-old woman with a right optic nerve sheath meningioma and visual loss, was treated with fractionated stereotactic radiotherapy. This resulted in an almost complete recovery of her visual acuity. DISCUSSION: Currently there are many different alternatives available for the management of an optic nerve sheath meningioma: observation, surgery, conventional radiotherapy and fractionated stereotactic radiotherapy. The last of these treatments has been demonstrated to preserve or improve vision in 2 out of 3 treated patients, with fewer side effects, and has not been associated with recurrent disease or tumor progression.
Assuntos
Meningioma/cirurgia , Neoplasias do Nervo Óptico/cirurgia , Radiocirurgia , Diplopia/etiologia , Fracionamento da Dose de Radiação , Exoftalmia/etiologia , Feminino , Humanos , Imageamento por Ressonância Magnética , Meningioma/complicações , Pessoa de Meia-Idade , Neoplasias do Nervo Óptico/complicações , Escotoma/etiologiaRESUMO
AIMS: To analyze the validity of the single deck 64-card Wisconsin Card Sorting Test (WCST-64) compared to the standard version (WCST-128), and to study the sensibility of both versions to identify changes over time in patients with traumatic brain injury (TBI). PATIENTS AND METHODS: The WCST was administered twice across a 6-month period to a sample of 50 patients with TBI. Pearson correlation coefficients were calculated to examine bivariate associations between WCST-128 scores and the corresponding WCST-64 scores at inclusion and at follow-up. Agreement in classification of impairment (z = -1) or normal performance was calculated for the two tests (kappa). Significant change over time was also analyzed for both versions of the test (paired-samples t test). RESULTS: The results revealed positive and significant correlations between both measures as well as a significant agreement in the classification of patients as having deficits or not. Moreover, the WCST-128 and the WCST-64 showed similar ability to identify changes over time. CONCLUSIONS: Our findings showed strong associations between scores derived from the two test and support the comparability of both versions when analyzing cross-sectional or longitudinal data. The findings support the use of the WCST-64 in evaluations of executive deficits of patients with moderate and severe TBI.
Assuntos
Lesões Encefálicas/diagnóstico , Testes Neuropsicológicos , Adolescente , Adulto , Feminino , Humanos , Masculino , Pessoa de Meia-IdadeRESUMO
AIM: Longitudinal study of verbal learning and memory processes after traumatic brain injury (TBI). PATIENTS AND METHODS: Twenty-six patients who had sustained a moderate-severe TBI were assessed with a Spanish version of the California Verbal Learning Test at the time of admission and 6-months after inclusion in a multidisciplinary rehabilitation program that comprised rehabilitation strategies for memory impairments. Global memory indexes were determined, including retroactive interference, proactive interference and recall indexes controlled for level of verbal acquisition. Memory change over time was correlated to demographic and clinical relevant variables, including the level of patients' self-awareness. RESULTS: More than 75% of our patients presented learning, immediate memory and delayed memory deficits at baseline, with an important effect of retroactive interference (69%). At 6-month follow-up, 34.6% showed learning difficulties, 46.2% immediate memory deficits, and 53% delayed memory problems, with 34.6% of the patients showing retroactive interference. Chronicity, level of self-awareness and premorbid intelligence correlated to the degree of memory change over time. CONCLUSIONS: Prominent verbal memory problems developed, not only during the first months after TBI but also over time are mostly due to impaired consolidation related to an intense retroactive interference. These data should be considered when developing memory rehabilitation strategies.
Assuntos
Lesões Encefálicas/complicações , Lesões Encefálicas/reabilitação , Transtornos da Linguagem/etiologia , Transtornos da Linguagem/reabilitação , Aprendizagem Verbal , Adolescente , Adulto , Cognição , Feminino , Seguimentos , Humanos , Estudos Longitudinais , Masculino , Pessoa de Meia-IdadeRESUMO
AIM: To establish the effectiveness of an educational board game for improving self-awareness following acquired brain injury. PATIENTS AND METHODS: The 'awareness climbing' is a board game format intervention for improving awareness of patients with acquired brain injury (ABI) and for facilitating the use of anticipatory and compensatory strategies. 17 patients with ABI (traumatic brain injury, 10; stroke, 5; others, 2) were included in a self-awareness rehabilitation program using the 'awareness climbing'. Self-awareness was assessed with the Self-Awareness of Deficit Interview (SADI) before and after game intervention. According to the SADI self-awareness consists of three-interdependent levels: intellectual awareness (impairments), emergent awareness (disability) and anticipatory awareness, which involve the client predicting when impairments will affect his or her performance. RESULTS: Initially five patients showed impaired intellectual awareness, four patients had difficulties on emergent awareness and nine patients showed deficits on anticipatory awareness. Intellectual awareness improved in 10 patients, while six patients experienced improvements in emergent awareness over the intervention period. The most striking change was an improvement in 70% of the sample (n = 11) in anticipatory awareness after game sessions. CONCLUSION: According to SADI scores we suggest that the 'awareness climbing' may be a useful tool for improving self-awareness after acquired brain injury.
Assuntos
Lesões Encefálicas , Recreação , Autoavaliação (Psicologia) , Adolescente , Adulto , Idoso , Conscientização , Lesões Encefálicas/patologia , Lesões Encefálicas/fisiopatologia , Lesões Encefálicas/terapia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Testes Neuropsicológicos , Reabilitação , AutoimagemRESUMO
The KCNQ family of voltage-dependent non-inactivating K+ channels is composed of five members, four of which (KCNQ2-5) are expressed in the CNS and are responsible for the M-current. Mutations in either KCNQ2 or KCNQ3 lead to a hereditary form of dominant generalized epilepsy. Using specific antisera to the KCNQ2, KCNQ3 and KCNQ5 subunits, we found that KCNQ3 co-immunoprecipitated with KCNQ2 and KCNQ5 subunits, but no association was detected between KCNQ2 and KCNQ5. Intense KCNQ5 immunoreactivity was found to be widely distributed throughout the temporal neocortex and the hippocampal formation. In these structures, both pyramidal and non-pyramidal neurons and a population of glial cells in the white matter expressed the KCNQ5 subunit. In the sclerotic areas of the CA fields of epileptic patients, a marked loss of KCNQ5 immunoreactive pyramidal neurons was found in relation with the loss of neurons in these regions. However, in the regions adjacent to the sclerotic areas, the distribution and intensity of KCNQ5 immunostaining was apparently normal. The widespread distribution of KCNQ5 subunits, its persistence in pharmacoresistant epilepsy, along with the significant role of the M-current in the control of neuronal excitability, makes this protein a possible target for the development of anticonvulsant drugs.
Assuntos
Epilepsia/metabolismo , Hipocampo/metabolismo , Canais de Potássio/metabolismo , Lobo Temporal/metabolismo , Adolescente , Adulto , Idoso , Western Blotting , Linhagem Celular , Epilepsia/patologia , Feminino , Feto , Imunofluorescência/métodos , Hipocampo/citologia , Humanos , Imuno-Histoquímica/métodos , Canais de Potássio KCNQ , Canal de Potássio KCNQ2 , Canal de Potássio KCNQ3 , Rim , Masculino , Pessoa de Meia-Idade , Canais de Potássio/genética , Canais de Potássio/imunologia , Canais de Potássio de Abertura Dependente da Tensão da Membrana , Testes de Precipitina/métodos , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/imunologia , Proteínas Recombinantes de Fusão/metabolismo , Lobo Temporal/citologia , Transfecção/métodosRESUMO
We have analyzed the folding state of cytosolic proteins imported in vitro into lysosomes, using an approach originally developed by Eilers and Schatz, (Eilers, M., and Schatz, G. (1986) Nature 322, 228-232) to investigate protein import into mitochondria. The susceptibility toward proteases of mouse dihydrofolate reductase (DHFR), synthesized in a coupled transcription-translation system with rabbit reticulocytes, decreased in the presence of its substrate analogue, methotrexate. This analogue complexes with high affinity with the in vitro synthesized DHFR and locks it into a protease-resistant folded conformation. DHFR was taken up by freshly isolated rat liver lysosomes and methotrexate reduced this uptake by about 80%. A chimeric DHFR protein, which carries the N-terminal presequence of subunit 9 of ATP synthase preprotein from Neurospora crassa fused to its N terminus, was taken up by lysosomes more efficiently. Again, methotrexate abolished the lysosomal uptake of the fusion protein, which was partially restored by washing of methotrexate from DHFR or by adding together methotrexate and dihydrofolate, the natural substrate of DHFR. Immunoblot analysis with anti-DHFR of liver lysosomes and of other fractions, isolated from rats starved for 88 h and treated with lysosomal inhibitors, suggests that DHFR is degraded by chaperone-mediated autophagy. Competition with ribonuclease A and stimulation by ATP/Mg(2+) and the heat shock cognate protein of 73 kDa show that the lysosomal uptake of the fusion protein also occurs by this pathway. It is concluded that the lysosomal uptake of cytosolic proteins by chaperone-mediated autophagy mainly occurs by passage of the unfolded proteins through the lysosomal membrane. Therefore, this mechanism is different from protein transport into peroxisomes, but similar to the import of proteins into the endoplasmic reticulum and mitochondria.