Regulation of Axon Initial Segment Diameter by COUP-TFI Fine-tunes Action Potential Generation / 神经科学通报·英文版

The axon initial segment (AIS) is a specialized structure that controls neuronal excitability via action potential (AP) generation. Currently, AIS plasticity with regard to changes in length and location in response to neural activity has been extensively investigated, but how AIS diameter is regulated remains elusive. Here we report that COUP-TFI (chicken ovalbumin upstream promotor-transcription factor 1) is an essential regulator of AIS diameter in both developing and adult mouse neocortex. Either embryonic or adult ablation of COUP-TFI results in reduced AIS diameter and impaired AP generation. Although COUP-TFI ablations in sparse single neurons and in populations of neurons have similar impacts on AIS diameter and AP generation, they strengthen and weaken, respectively, the receiving spontaneous network in mutant neurons. In contrast, overexpression of COUP-TFI in sparse single neurons increases the AIS diameter and facilitates AP generation, but decreases the receiving spontaneous network. Our findings demonstrate that COUP-TFI is indispensable for both the expansion and maintenance of AIS diameter and that AIS diameter fine-tunes action potential generation and synaptic inputs in mammalian cortical neurons.

MicroRNA-221 promotes cell proliferation, migration, and differentiation by regulation of ZFPM2 in osteoblasts

Bone fracture is a common medical condition, which may occur due to traumatic injury or disease-related conditions. Evidence suggests that microRNAs (miRNAs) can regulate osteoblast differentiation and function. In this study, we explored the effects and mechanism of miR-221 on the growth and migration of osteoblasts using MC3T3-E1 cells. The expression levels of miR-221 in the different groups were measured by qRT-PCR. Then, miR-221 mimic and inhibitor were transfected into MC3T3-E1 cells, and cell viability and migration were measured using the CCK-8 assay and the Transwell migration assay. Additionally, the expression levels of differentiation-related factors (Runx2 and Ocn) and ZFPM2 were measured by qRT-PCR. Western blot was used to measure the expression of cell cycle-related proteins, epithelial-mesenchymal transition (EMT)-related proteins, ZFPM2, and Wnt/Notch, and Smad signaling pathway proteins. miR-221 was significantly up-regulated in the patients with lumbar compression fracture (LCM) and trochanteric fracture (TF). miR-221 promoted ALP, Runx2, and OPN expressions in MC3T3-E1 cells. miR-221 overexpression significantly increased cell proliferation, migration, differentiation, and matrix mineralization, whereas suppression of miR-221 reversed these effects. Additionally, the results displayed that ZFPM2 was a direct target gene of miR-221, and overexpression of ZFPM2 reversed the promoting effects of miR-221 overexpression on osteoblasts. Mechanistic study revealed that overexpression of miR-221 inactivated the Wnt/Notch and Smad signaling pathways by regulating ZFPM2 expression. We drew the conclusions that miR-221 overexpression promoted osteoblast proliferation, migration, and differentiation by regulation of ZFPM2 expression and deactivating the Wnt/Notch and Smad signaling pathways.

Occurrence of Lutzomyia longipalpis (Phlebotominae) and canine visceral leishmaniasis in a rural area of Ilha Solteira, SP, Brazil / Ocorrência de Lutzomyia longipalpis (Phlebotominae) e leishmaniose visceral canina em uma área rural de Ilha Solteira, SP, Brasil

This study aimed to investigate the occurrence of Lutzomyia longipalpis and also the canine visceral leishmaniasis (CVL) in a rural area of Ilha Solteira, state of São Paulo. Blood samples were collected from 32 dogs from different rural properties (small farms) and were analyzed by ELISA and the indirect immunofluorescence antibody test (IFAT) in order to diagnose CVL. From these serological tests, 31.25% of the dogs were positive for CVL and these were distributed in 66.7% (8/12) of the rural properties, which were positive for L. longipalpis. CDC (Center for Disease Control and Prevention) light traps were installed in 12 properties (one per property) and insects were caught on three consecutive days per month for one year. L. longipalpis was present on 100% of the rural properties visited, at least once during the twelve-month interval, totaling 64 males and 25 females. The insects were more numerous after the peak of the rain, but the association between prevalence of peridomestic vectors and the climatic data (precipitation, relative air humidity and temperature) and the occurrences of CVL among dogs on each rural property were not statistical significant (p <0.05). However, the occurrence of CVL cases in dogs and the presence of L. longipalpis indicate that more attention is necessairy for the control of this disease in the rural area studied.

O objetivo desse trabalho foi o estudo da prevalência de Lutzomyia longipalpis e da leishmaniose visceral canina (LVC) em uma área rural do município de Ilha Solteira do estado de São Paulo. Amostras de sangue foram coletadas de 32 cães provenientes de pequenas propriedades rurais e analisadas por meio dos métodos sorológicos ELISA (imunoensaio enzimático indireto) e RIFI (reação de imunofluorescência indireta) para o diagnóstico da LVC. Pelos exames sorológicos, dos 32 cães avaliados, 31,25% foram diagnosticados positivos para LVC, os quais estavam diostribuídos em 66,67% (8/12) das propriedades positivas para Lutzomyia longipalpis. Armadilhas luminosas do tipo CDC (Center for Disease Control and Prevention) foram instaladas em 12 propriedades, sendo uma por propriedade, e as coletas dos insetos foram realizadas três dias consecutivos a cada mês, durante um ano. O inseto L. longipalpis foi encontrado em 100% das propriedades visitadas, pelo menos uma vez no ano, totalizando 65 machos e 25 fêmeas. A maior quantidade de insetos foi observada principalmente após a ocorrência dos maiores picos de precipitação pluvial, mas a associação entre a prevalência dos vetores peridomiciliares e os dados climáticos (precipitação, umidade relativa do ar e temperatura) assim como a ocorrência da CVL em cães em cada propriedade não foi estatisticamente significante (p<0.05). No entanto, alerta-se que pela presença dos casos de LVC nos cães amostrados e também de L. longipalpis, maior atenção deve ser dada durante as investigações epidemiológicas para o controle dessa doença nessa área rural estudada.

Medication adherence and quality of life among the elderly with diabetic retinopathy / Adesão medicamentosa e qualidade de vida em idosos com retinopatia diabética / Adhesión a la medicación y calidad de vida en ancianos con retinopatía diabética

METHOD: one hundred (n=100) elderly outpatients with diabetic retinopathy taking antihypertensives and/or oral antidiabetics/insulin were interviewed. Adherence was evaluated by the adherence proportion and its association with the care taken in administrating medications and by the Morisky Scale. The National Eye Institute Visual Functioning Questionnaire (NEI VFQ-25) was used to evaluate HRQoL. RESULTS: most (58%) reported the use of 80% or more of the prescribed dose and care in utilizing the medication. The item "stopping the drug when experiencing an adverse event", from the Morisky Scale, explained 12.8% and 13.5% of the variability of adherence proportion to antihypertensives and oral antidiabetics/insulin, respectively. CONCLUSION: there was better HRQoL in the Color Vision, Driving and Social Functioning domains of the NEI VFQ-25. Individuals with lower scores on the NEI VFQ-25 and higher scores on the Morisky Scale presented greater chance to be nonadherent to the pharmacological treatment of diabetes and hypertension. .

OBJETIVO: investigar os fatores relacionados à adesão medicamentosa e sua relação com a qualidade de vida relacionada à saúde em idosos com retinopatia diabética. MÉTODO: foram entrevistados 100 idosos, em acompanhamento ambulatorial, em uso de anti-hipertensivos e/ou antidiabéticos orais/insulina. A adesão foi avaliada pela proporção de adesão e sua associação com os cuidados no uso dos medicamentos e pela Escala de Morisky. O National Eye Institute Visual Funcioning Questionnaire foi utilizado para avaliar a qualidade de vida relacionada à saúde. RESULTADOS: A maioria (58%) relatou o uso de 80% ou mais das doses prescritas e os cuidados na tomada dos medicamentos. O item "interromper o uso dos medicamentos por se sentir pior", da Escala de Morisky, explicou 12,8 e 13,5% da variabilidade da proporção de adesão aos anti-hipertensivos e aos antidiabéticos orais/insulina, respectivamente. CONCLUSÃO: observou-se melhor qualidade de vida relacionada à saúde nos domínios visão de cores, dirigir automóvel e apectos sociais do National Eye Institute Visual Funcioning Questionnaire. Indivíduos com menor pontuação na National Eye Institute Visual Funcioning Questionnaire e maiores escores na Escala de Morisky apresentaram maiores chances de serem não aderentes aos medicamentos do diabetes e da hipertensão arterial. .

OBJETIVO: investigar los factores relacionados a la adhesión a la medicación y su relación con la Calidad de Vida Relacionada a la Salud (CVRS) de ancianos con retinopatía diabética. MÉTODO: fueron entrevistados cien (n=100) pacientes ancianos de ambulatorio con retinopatía diabética que toman medicamentos antihipertensivos y/o antidiabéticos orales/insulina. La adhesión fue evaluada mediante la proporción de adhesión y su asociación con el cuidado tomado en la administración de medicamentos y mediante la Escala de Morisky. El National Eye Institute Visual Functioning Questionnaire (NEI VFQ-25) fue usado para evaluar la CVRS. RESULTADOS: la mayoría (58%) relató el uso de 80% o más de la dosis prescrita y cuidado con el uso de la medicación. El ítem "suspender la droga cuando vivencia un evento adverso", de la Escala de Morisky, explicó 12.8% y 13.5% de la variabilidad en la proporción de adhesión a los antihipertensivos y antidiabéticos orales/insulina, respectivamente. CONCUSIÓN: fue encontrada mejor CVRS en los dominios de Visión Cromática, Dirección y Funcionamiento Social del NEI VFQ-25. Individuos con puntuaciones menores en el NEI VFQ-25 y puntuaciones mayores en la Escala de Morisky revelaron mayor chance de no adhesión al tratamiento farmacológico de la diabetes y hipertensión. .

Low dose hyper-radiosensitivity in human lung cancer cell line A549 and its possible mechanisms / 华中科技大学学报（医学）（英德文版）

The low dose hyper-radiosensitivity (HRS) in human lung cancer cell line A549 was investigated, the changes of ATM kinase, cell cycle and apoptosis of cells at different doses of radiation were observed, and the possible mechanisms were discussed. A549 cells in logarithmic growth phase were irradiated with (60)Co gamma-rays at doses of 0-2 Gy. Together with flow cytometry for precise cell sorting, cell survival fraction was measured by means of conventional colony-formation assay. The expression of ATM1981Ser-P protein was examined by Western blot 1 h after radiation. Apoptosis was detected by Hoechst 33258 fluorescent staining, and Annexin V-FITC/PI staining flow cytometry 24 h after radiation. Cell cycle distribution was observed by flow cytometry 6, 12 and 24 h after radiation. The results showed that the expression of ATM1981Ser-P protein was observed at 0.2 Gy, followed by an increase at >0.2 Gy, and reached the peak at 0.5 Gy, with little further increase as the dose exceeded 0.5 Gy. Twenty-four h after radiation, partial cells presented the characteristic morphological changes of apoptosis, and the cell apoptosis curve was coincident with the survival curve. As compared with control group, the cell cycle almost had no changes after exposure to 0.1 and 0.2 Gy radiation (P>0.05). After exposure to 0.3, 0.4 and 0.5 Gy radiation, G(2)/M phase arrest occurred 6 and 12 h after radiation (P<0.05), and the ratio of G(2)/M phase cells was decreased 24 h after radiation (P<0.05). It was concluded that A549 cells displayed the phenomenon of HRS/IRR. The mode of cell death was mainly apoptosis. The activity of ATM and cell cycle change may take an important role in HRS/IRR.

The functional role of the CARM1-SNF5 complex and its associated HMT activity in transcriptional activation by thyroid hormone receptor

We have investigated the function and mechanisms of the CARM1-SNF5 complex in T3-dependent transcriptional activation. Using specific small interfering RNAs (siRNA) to knock down coactivators in HeLa alpha2 cells, we found that coactivator associated arginine methyltransferase 1 (CARM1) and SWI/SNF complex component 5 (SNF5) are important for T3-dependent transcriptional activation. The CARM1- SWI/SNF chromatin remodeling complex serves as a mechanism for the rapid reversal of H3-K9 methylation. Importantly, siRNA treatment against CARM1 and/or SNF5 increased the recruitment of HMTase G9a to the type 1 deiodinase (D1) promoter even with T3. Knocking- down either CARM1 or SNF5 also inhibited the down- regulation of histone macroH2A, which is correlated with transcriptional activation. Finally, knocking down CARM1 and SNF5 by siRNA impaired the association of these coactivators to the D1 promoter, suggesting functional importance of CARM1- SNF5 complex in T3-dependent transcriptional activation.

The eukaryotic Pso2/Snm1/Artemis proteins and their function as genomic and cellular caretakers

DNA double-strand breaks (DSBs) represent a major threat to the genomic stability of eukaryotic cells. DNA repair mechanisms such as non-homologous end joining (NHEJ) are responsible for the maintenance of eukaryotic genomes. Dysfunction of one or more of the many protein complexes that function in NHEJ can lead to sensitivity to DNA damaging agents, apoptosis, genomic instability, and severe combined immunodeficiency. One protein, Pso2p, was shown to participate in the repair of DSBs induced by DNA inter-strand cross-linking (ICL) agents such as cisplatin, nitrogen mustard or photo-activated bi-functional psoralens. The molecular function of Pso2p in DNA repair is unknown, but yeast and mammalian cell line mutants for PSO2 show the same cellular responses as strains with defects in NHEJ, e.g., sensitivity to ICLs and apoptosis. The Pso2p human homologue Artemis participates in V(D)J recombination. Mutations in Artemis induce a variety of immunological deficiencies, a predisposition to lymphomas, and an increase in chromosomal aberrations. In order to better understand the role of Pso2p in the repair of DSBs generated as repair intermediates of ICLs, an in silico approach was used to characterize the catalytic domain of Pso2p, which led to identification of novel Pso2p homologues in other organisms. Moreover, we found the catalytic core of Pso2p fused to different domains. In plants, a specific ATP-dependent DNA ligase I contains the catalytic core of Pso2p, constituting a new DNA ligase family, which was named LIG6. The possible functions of Pso2p/Artemis/Lig6p in NHEJ and V(D)J recombination and in other cellular metabolic reactions are discussed.

A role for histone-like protein H1 (H-NS) in the regulation of hemolysin expression by Serratia marcescens

The histone-like protein H1 (H-NS) is an abundant structural component of the bacterial nucleoid and influences many cellular processes including recombination, transcription and transposition. Mutations in the hns gene encoding H-NS are highly pleiotropic, affecting the expression of many unrelated genes. We have studied the role of H-NS on the regulation of hemolysin gene expression in Serratia marcescens. The Escherichia coli hns mutant carrying S. marcescens hemolysin genes on a plasmid constructed by ligation of the 3.2-kb HindIII-SacI fragment of pR02 into pBluescriptIIKS, showed a high level of expression of this hemolytic factor. To determine the osmoregulation of wild-type and hns defective mutants the cells were grown to mid-logarithmic phase in LB medium with 0.06 or 0.3 M NaCl containing ampicillin and kanamycin, whereas to analyze the effect of pH on hemolysin expression, the cells were grown to late-logarithmic phase in LB medium buffered with 0.1 M Tris-HCl, pH 4.5 to 8.0. To assay growth phase-related hemolysin production, bacterial cells were grown in LB medium supplemented with ampicillin and kanamycin. The expression of S. marcescens hemolysin genes in wild-type E. coli and in an hns-defective derivative at different pH and during different growth phases indicated that, in the absence of H-NS, the expression of hemolysin did not vary with pH changes or growth phases. Furthermore, the data suggest that H-NS may play an important role in the regulation of hemolysin expression in S. marcescens and its effect may be due to changes in DNA topology influencing transcription and thus the amount of hemolysin expression. Implications for the mechanism by which H-NS influences gene expression are discussed.

The maternal JAK/STAT pathway of Drosophila regulates embryonic dorsal-ventral patterning

Activation of NFkappaB plays a pivotal role in many cellular processes such as inflammation, proliferation and apoptosis. In Drosophila, nuclear translocation of the NFkappaB-related transcription factor Dorsal is spatially regulated in order to subdivide the embryo into three primary dorsal-ventral (DV) domains: the ventral presumptive mesoderm, the lateral neuroectoderm and the dorsal ectoderm. Ventral activation of the Toll receptor induces degradation of the IkappaB-related inhibitor Cactus, liberating Dorsal for nuclear translocation. In addition, other pathways have been suggested to regulate Dorsal. Signaling through the maternal BMP member Decapentaplegic (Dpp) inhibits Dorsal translocation along a pathway parallel to and independent of Toll. In the present study, we show for the first time that the maternal JAK/STAT pathway also regulates embryonic DV patterning. Null alleles of loci coding for elements of the JAK/STAT pathway, hopscotch (hop), marelle (mrl) and zimp (zimp), modify zygotic expression along the DV axis. Genetic analysis suggests that the JAK kinase Hop, most similar to vertebrate JAK2, may modify signals downstream of Dpp. In addition, an activated form of Hop results in increased levels of Cactus and Dorsal proteins, modifying the Dorsal/Cactus ratio and consequently DV patterning. These results indicate that different maternal signals mediated by the Toll, BMP and JAK/STAT pathways may converge to regulate NFkappaB activity in Drosophila.

Zinc fingers: DNA binding and protein-protein interactions

The zinc finger domain is a very ubiquitous structural element whose hallmark is the coordination of a zinc atom by several amino acid residues (cysteines and histidines, and occasionally aspartate and glutamate). These structural elements are associated with protein-nucleic acid recognition as well as protein-protein interactions. The purpose of this review is to examine recent data on the DNA and protein binding properties of a few zinc fingers whose three dimensional structure is known.

Autoantigen Ku and its role in multiple cellular processes.

Ku is a DNA binding protein composed of 70 and 80 kDa subunits which was discovered as autoantigen in a patient with scleroderma-polymyositis overlap syndrome. Ku can bind to the end of DNA and also to some internal sequences. Ku-autoantigen acts as a potential transcription factor for several RNA polymerase II genes and RNA polymerase I gene. Ku is also associated with DNA-dependent protein kinase and involved in V(D)J recombination and DNA break repair mechanisms. Ku may be involved in replication, helicase activity and cell signaling. Therefore, Ku-autoantigen is a very important cellular factor which plays important role in the multiple cellular processes.

IRF-1 and IRF-2 as regulators of the interferon system and cell growth.

Interferons (IFNs) are families of cytokines which have been discovered and extensively characterized in the context of host defense against viral infections. We have discovered two structurally related transcription factors, Interferon regulatory factor-1 (IRF-1) and IRF-2. These two factors, however, function not only as regulators of the IFN system, but are also key transcription factors in the regulation of cell cycle and apoptosis. These studies uncover a complex gene transcription network, by which the fate of cellular responses are determined depending on how the IRF transcription factors function in conjunction with other factors, and on the promoters of distinct genes under different conditions of the cells.

Relationship of single-stranded DNA-binding proteins of Ehrlich ascites tumour to cell growth phase and DNA replications.

The possible involvement of SSB-proteins in DNA replication in Ehrlich ascites tumour (EAT) has been investigated. A direct relation (the computer-generated correlation coefficient was 0.9) between the SSB-proteins content in chromatin and intensity of the replicative synthesis of DNA in various preparation of EAT in vivo and in vitro is observed. Addition of exogenous SSB-proteins to the permeable EAT cells has been found to increase the replicative synthesis. Although eukaryotic SSB-proteins are not complete analogs of the prokaryotic SSB-proteins, they evidently participate in DNA replication in eukaryotic cells and possibly are intracellular regulators of proliferation.