Participation of leukaemia inhibitory factor in follicular development and steroidogenesis in rat ovaries.

Leukaemia inhibitory factor (LIF) is a cytokine belonging to the interleukin-6 family that is important at the reproductive level in the uterine implantation process. However, there is very little evidence regarding its effect at the ovarian level. The aim of this work was to study the local involvement of the LIF/LIFRß system in follicular development and steroidogenesis in rat ovaries. To carry out this research, LIF/LIFR/GP130 transcript and protein levels were measured in fertile and sub-fertile rat ovaries, and in vitro experiments were performed to assess STAT3 activation. Then, in in vivo experiments, LIF was administered chronically and locally for 28 days to the ovaries of rats by means of an osmotic minipump to enable us to evaluate the effect on folliculogenesis and steroidogenesis. It was determined by quantitative polymerase chain reaction and western blot that LIF and its receptors are present in fertile and sub-fertile ovaries and that LIF varies during the oestrous cycle, being higher during the oestrus and meta/dioestrus stages. In addition to this, it was found that LIF can activate STAT3 pathways and cause pSTAT3 formation. It was also observed that LIF decreases the number and size of preantral and antral follicles without altering the number of atretic antral follicles and can increase the number of corpora lutea, with a notable increase in the levels of progesterone (P4). It is therefore possible to infer that LIF exerts an important effect in vivo on folliculogenesis, ovulation and steroidogenesis, specifically the synthesis of P4.

IL-6 Cytokine Family: A Putative Target for Breast Cancer Prevention and Treatment.

The IL-6 cytokine family is a group of signaling molecules with wide expression and function across vertebrates. Each member of the family signals by binding to its specific receptor and at least one molecule of gp130, which is the common transmembrane receptor subunit for the whole group. Signal transduction upon stimulation of the receptor complex results in the activation of multiple downstream cascades, among which, in mammary cells, the JAK-STAT3 pathway plays a central role. In this review, we summarize the role of the IL-6 cytokine family-specifically IL-6 itself, LIF, OSM, and IL-11-as relevant players during breast cancer progression. We have compiled evidence indicating that this group of soluble factors may be used for early and more precise breast cancer diagnosis and to design targeted therapy to treat or even prevent metastasis development, particularly to the bone. Expression profiles and possible therapeutic use of their specific receptors in the different breast cancer subtypes are also described. In addition, participation of these cytokines in pathologies of the breast linked to lactation and involution of the gland, as post-partum breast cancer and mastitis, is discussed.

Leukemia Inhibitory Factor (LIF) Overexpression Increases the Angiogenic Potential of Bone Marrow Mesenchymal Stem/Stromal Cells.

Mesenchymal stem/stromal cells (MSCs) have the ability to secrete bioactive molecules, exerting multiple biological effects, such as tissue regeneration, reduction of inflammation, and neovascularization. The therapeutic potential of MSCs can be increased by genetic modification to overexpress cytokines and growth factors. Here we produced mouse MSCs overexpressing human leukemia inhibitory factor (LIF) to assess their proangiogenic potential in vitro and in vivo. Mouse bone marrow-derived MSCs were transduced by using a second-generation lentiviral system to express human LIF. Leukemia inhibitory factor expression was confirmed by RT-qPCR and by ELISA, allowing the quantification of the transcript and secreted protein, respectively. Flow cytometry analysis and trilineage differentiation assay showed that the MSC_LIF cell line maintained the immunophenotype and a multipotency characteristic of MSCs. The immunosuppressive activity of MSC_LIF was confirmed using a lymphoproliferation assay. Moreover, gene expression analysis demonstrated upregulation of genes coding for strategic factors in the neovascularization process, such as angiogenin, IL-8, MCP-1, and VEGF, and for the perivascular cell markers αSMA, Col4a1, SM22, and NG2. To evaluate the pro-angiogenic potential of MSC_LIF, we first tested its effects on endothelial cells obtained from umbilical vein in a scratch wound healing assay. Conditioned medium (CM) from MSC_LIF promoted a significant increase in cell migration compared to CM from control MSC. Additionally, in vitro tube formation of endothelial cells was increased by the presence of MSC_LIF, as shown in microvessel sprouting in aortic ring cultures. Finally, an in vivo Matrigel plug assay was performed, showing that MSC_LIF were more potent in promoting in vivo angiogenesis and tissue vascularization than control MSCs. In conclusion, LIF overexpression is a promising strategy to increase the proangiogenic potential of MSCs and sets precedents for future investigations of their potential applications for the treatment of ischemic diseases and tissue repair.

A dose-dependent response to MEK inhibition determines hypoblast fate in bovine embryos.

BACKGROUND: The segregation of the hypoblast and the emergence of the pluripotent epiblast mark the final stages of blastocyst formation in mammalian embryos. In bovine embryos the formation of the hypoblast has been partially studied, and evidence shows that MEK signalling plays a limited role in the segregation of this lineage. Here we explored the role of different signalling pathways during lineage segregation in the bovine embryo using immunofluorescence analysis of NANOG and SOX17 as readouts of epiblast and hypoblast, respectively. RESULTS: We show that SOX17 starts to be expressed in 16-32-cell stage embryos, whereas NANOG is first detected from 8-cell stage. SOX17 is first co-expressed with NANOG, but these markers become mutually exclusive by the late blastocyst stage. By assessing the expression kinetics of NANOG/SOX17 we show that inhibition of MEK signalling can eliminate SOX17 expression in bovine blastocysts, without altering NANOG expression. Modulation of WNT, PKC and LIF did not affect NANOG expression in the epiblast when used in combination with the ERK inhibitor. CONCLUSIONS: This study shows that SOX17 can be used as a reliable early marker of hypoblast in the bovine, and based on its expression profile we show that the hypoblast segregates in day 7 blastocysts. Furthermore, SOX17 expression is abolished using 1 µM of PD0325901, without affecting the NANOG population in the epiblast. Modulation of WNT, PKC and LIF are not sufficient to support enhanced NANOG expression in the epiblast when combined with ERK inhibitor, indicating that additional signalling pathways should be examined to determine their potential roles in epiblast expansion.

Alveolar bone pattern and salivary leptin levels among premenopausal obese women / Padrão ósseo alveolar e níveis salivares de leptina entre mulheres obesas na pré-menopausa

ABSTRACT Background: Systemic bone loss may lead to more severe periodontal destruction, decreasing local bone mineral density. Aim: A cross-sectional designed was performed to study associations among alveolar bone pattern, salivary leptin concentrations, and clinical periodontal status in premenopausal obese and eutrophic women. Methods: Thirty morbid obese (G1) and 30 normal-weight (G2) women were included. Anthropometric and periodontal measurements (probing depth - PD, clinical attachment levels - CAL, presence of calculus, bleeding on probing -BOP, and plaque accumulation) were assessed. OHIP-14 was used for assessment of oral health impact on quality of life. Panoramic radiography was used to obtain the panoramic mandibular index (PMI), mandibular cortical index (MCI), and mental index (MI). Intraoral periapical (PA) radiography was taken to measure the total trabecular bone volume. Leptin was measured in saliva of fasted overnight women. Results: Groups 1 and 2 differed in all anthropometric aspects, but height. Pocket depth, calculus, BOP, and plaque index were worse in G1. No differences between groups were found considering OHIP. Normal-weight subjects showed higher proportion of dense bone trabeculae than obese subjects for pre-molars, but not for molars. Mental and panoramic mandibular indexes did not differ and were in normal level. Leptin concentration was dependent only on BMI. Conclusion: Obesity affected the periodontal conditions, the alveolar bone pattern, and the salivary leptin concentration.

RESUMO Racional: A perda óssea sistêmica pode levar à destruição periodontal mais severa, diminuindo a densidade mineral óssea local. Objetivo: Investigar as associações entre padrão ósseo alveolar, concentrações de leptina salivar e estado periodontal em mulheres obesas na pré-menopausa e eutróficas. Métodos: Foram avaliadas 30 mulheres com obesidade mórbida (G1) e 30 com peso normal (G2). Foram analisadas as medidas antropométricas e periodontais (profundidade de sondagem - PS, nível clínico de inserção - NCI, presença de cálculo, sangramento à sondagem - SS e acúmulo de placa). O impacto da saúde bucal na qualidade de vida foi mensurado por meio do questionário OHIP-14. Radiografia panorâmica foi utilizada para obter o índice mandibular panorâmico (PMI), índice cortical mandibular (MCI) e índice mental (MI); já a radiografia periapical intraoral (AF) para medir o volume ósseo trabecular total. A leptina salivar foi coletada no período da manhã com a paciente em jejum. Resultados: Os grupos 1 e 2 diferiram em todos os aspectos antropométricos, exceto em estatura. Profundidade de bolsa, cálculo, SS e índice de placa foram piores no G1. Não foram encontradas diferenças entre os grupos considerando o OHIP. Indivíduos com peso normal apresentaram maior proporção de trabéculas ósseas densas do que os obesos para pré-molares, mas não para molares. Índices radiomorfométricos não diferiram entre os grupos e estavam dentro de valores normais. A concentração de leptina esteve associada ao IMC. Conclusão: A obesidade afetou as condições periodontais, o padrão ósseo alveolar e a concentração de leptina salivar.

Leukemia Inhibitory Factor Increases Survival of Pluripotent Stem Cell-Derived Cardiomyocytes.

Leukemia inhibitory factor (LIF) is a growth factor with pleiotropic biological functions. It has been reported that LIF acts at different stages during mesoderm development. Also, it has been shown that LIF has a cytoprotective effect on neonatal murine cardiomyocytes (CMs) in culture, but little is known about the role of LIF during human cardiogenesis. Thus, we analyzed the effects of LIF on human pluripotent stem cells (PSC) undergoing cardiac differentiation. We first showed that LIF is expressed in the human heart during early development. We found that the addition of LIF within a precise time window during the in vitro differentiation process significantly increased CMs viability. This finding was associated to a decrease in the expression of pro-apoptotic protein Bax, which coincides with a reduction of the apoptotic rate. Therefore, the addition of LIF may represent a promising strategy for increasing CMs survival derived from PSCs.

Alterations in expression of endometrial milk fat globule-EGF factor 8 (MFG-E8) and leukemia inhibitory factor (LIF) in patients with infertility and endometriosis.

OBJECTIVE: The aim of this study was to compare the endometrial expression of milk fat globule-EGF factor 8 (MFG-E8), its receptor integrin αvß3, and leukemia inhibitory factor (LIF) in patients with endometriosis and infertility and in healthy fertile patients during the window of implantation. METHODS: Five patients with peritoneal endometriosis and infertility (case group) and four healthy fertile patients (control group) were recruited. All patients were either diagnosed with or ruled out for endometriosis by laparoscopic surgery; the case group underwent surgery for infertility investigation and the control group for tubal ligation. Endometrial biopsies were performed in all patients during the window of implantation (LH+8 to LH+10), and then the samples were analyzed by immunochemistry for MFG-E8, integrin αvß3, and LIF. RESULTS: In patients with endometriosis and infertility, expression of MFG-E8 was significantly increased in the glandular epithelium when compared to healthy fertile patients (p<0.001). Moreover, LIF expression was lower in patients with endometriosis and infertility (p<0.05). Nevertheless, we found no difference in integrin αvß3 expression between the groups (p=0.084). CONCLUSION: This study showed for the first time that MFG-E8 expression is impaired in the endometrium of patients with endometriosis and infertility during the window of implantation. Moreover, LIF is also diminished in the endometrium of these patients as shown before.

Colony stimulating factor-1 and leukemia inhibitor factor expression from current-cycle cannula isolated endometrial cells are associated with increased endometrial receptivity and pregnancy.

BACKGROUND: Poor endometrial quality is associated with more than a third of embryo implantation failures. Current ultrasonography technology lacks the capacity to determine efficiently the endometrial receptivity during ongoing cycle transfers. We analyzed the relationship between the gene expression profile associated with implantation and clinical pregnancy from endometrial cells taken during embryo transfer. METHODS: Seventy-six patients submitted to a standard ovarian stimulation protocol, in vitro fertilization, and good quality embryos were collected (morphological assessment). Endometrial samples were taken with ultrasonography guidance and cells were Hematoxylin and Eosin stained for morphological identification. Total RNA was extracted and the expression of Mucin 1 (MUC1), Homeobox A10 (HOXA-10), Leukemia Inhibitor Factor (LIF), Colony Stimulating Factor-1 (CSF-1), and ribosomal 18 s (endogenous control) were analyzed using RT-qPCR. Presence of a gestational sac, ß-hGC (≥10 mIU/mL on Day 20), and a fetal heartbeat were used to determine a positive embryo implantation and pregnancy. RESULTS: Samples collected from same cycle embryo transfer showed clear morphological staining for endometrial cells (80-90% of the cells). Cells in the sample were molecularly identified as the endometrium (HOXA-10 positive and MUC-1 negative). CSF-1 expression was 4.55-fold and LIF expression was 12.25-fold higher in patients who became pregnant. Both increases were statistically significant (p < 0.05). CONCLUSIONS: Here, we provide evidence of a new method to assess endometrial receptivity. Furthermore, we demonstrate that the expression profile, based on LIF and CSF-1, showed a difference between a receptive and a non-receptive endometrium.

El desafío de construir una Psicología del desarrollo crítica en sociedades inhóspitas / The Challenge of Constructing a Critical Developmental Psychology in Inhospitable Societies

Resumen La Psicología del desarrollo constituye un campo de conocimiento cuya existencia se da por descontada en el imaginario académico de la psicología salvadoreña. No obstante, existe una Psicología del desarrollo dominante que se ve reproducida en textos, clases y discursos, mientras que otra distinta, consecuente con la realidad histórica donde su estudio tiene lugar, aún es una tarea pendiente. Al considerar la descontextualización como uno de los presupuestos fundamentales de la Psicología del desarrollo dominante al uso, en este artículo de carácter teórico se reflexiona sobre esta desde la perspectiva del caso salvadoreño en tanto sociedad inhóspita: aquella que compromete el avance del ciclo vital de la mayoría de la población por, entre otras características, ser violenta y desigual. Son sometidas a escrutinio las conocidas categorías analíticas de la Psicología del desarrollo definidas como influencias normativas y no normativas para mostrar la relatividad y los sesgos de su aplicación usual. Se sostiene que, en el marco de una sociedad inhóspita, una Psicología del desarrollo debe considerar, al menos, procesos de adultización prematura en la niñez y la juventud, la improbabilidad de biografías estables, la vejez precaria así como las características típicas que muestran los miembros del contexto de interés. Se propone avanzar a una Psicología del desarrollo crítica: la necesidad de reacomodar su quehacer y reinventarse epistemológicamente para construir un conocimiento situado, propio y multidisciplinario que supere cegueras, provincianismos y reduccionismos académicos.

Abstract Developmental Psychology is a field of knowledge whose existence is taken for granted in the academic imagination of Salvadoran psychology. However, there is a mainstream Developmental Psychology that is reproduced in texts, lectures and discourses, while a different one, consistent with the historical reality where its study takes place, and is still a pending task. Considering de-contextualization as one of its fundamental assumptions, this theoretical article deals with the situation of Developmental Psychology from the perspective of the Salvadoran case as an Inhospitable Society: one that distorts the progress of most of its people´s life cycle, due to, among other features, its violence and social inequality. Very well known analytical categories such as normative and non-normative developmental trends are subjected to scrutiny to show the relativity and biases of their usual applications. It is argued that, in the context of an inhospitable society, Developmental Psychology should consider at least: Early Childhood and Youth Adultisation Processes, the improbability of stable biographies, Precarious Old Age and the typical features of individuals living in the context of interest. It aims to advance a Critical Developmental Psychology: the need to rearrange the work and to reinvent itself epistemologically in order to construct a situated, appropriated and multidisciplinary knowledge that overcomes blindness, provincialism and academic reductionism.

Association between metabolic syndrome and quality of life / Associação entre síndrome metabólica e qualidade de vida

AIMS: Despite all progress made in understanding and treating metabolic syndrome, the study of its impact on quality of life is still controversial and not well understood. The aim of this study was to test the hypothesis that metabolic syndrome can be associated with a worse quality of life. MESTHODS: A controlled cross-sectional study included individuals with metabolic syndrome, from the sub-analysis of a randomized clinical trial about lifestyle modification and cardiovascular risk factors, as well as individuals with no metabolic syndrome, attended as outpatients in several clinics at a general university hospital in Southern Brazil. Measurements were made in individual interviews and included data collection, laboratory tests, and application of general scales such as Mini-Mental State Examination and Medical Outcomes Study Short Form, General Health Survey (SF-36). Comparisons of quantitative data used the Student's t test, followed by analysis of covariance or multiple linear regression for adjustment, and correlation coefficient. For categorical data, the Chi-square or Fisher's exact test were used. RESULTS: The study included 229 individuals, 118 metabolic syndrome and 111 no metabolic syndrome. In univariate analysis, metabolic syndrome was significantly associated with lower scores in the social functioning (p<0.001) and role emotional (p=0.019) quality of life domains, and with the Mental Component Summary score of the SF-36 (p=0.013). However, after adjustments for confounding factors, especially body mass index, the significance of these associations was lost. The only significant association between metabolic syndrome and quality of life that has remained after the adjustments was with worse QOL in the role emotional domain, but only in men (p=0.049). CONCLUSIONS: After adjusting for multiple variables, metabolic syndrome was significantly associated with decreased quality of life scores in men in the role emotional domain.

OBJETIVOS: Apesar dos avanços no entendimento e tratamento da síndrome metabólica, o estudo do seu impacto sobre a qualidade de vida é ainda controverso e inconclusivo. O objetivo deste estudo foi testar a hipótese de que a síndrome metabólica estaria associada à piora da qualidade de vida. MÉTODOS: Um estudo transversal controlado incluiu indivíduos com síndrome metabólica, advindos da subanálise de um estudo controlado randomizado sobre modificação de estilo de vida e fatores de risco cardiovascular; e indivíduos sem síndrome metabólica, provenientes de diversos ambulatórios de um hospital geral no sul do Brasil. As medidas foram realizadas em entrevistas individuais, que incluíram coleta de dados demográficos, testes laboratoriais e aplicação do Exame do Estado Mental e o Medical Outcome Study Short Form, General Health Survey (SF-36). Para comparações entre dados quantitativos, foram utilizados teste t de Student, análise de covariância ou regressão linear múltipla para os ajustes dos fatores confundidores, e coeficiente de correlação. Para dados categóricos foi utilizado o qui quadrado ou o Exato de Fisher quando necessário. RESULTADOS: O estudo incluiu 229 indivíduos, sendo 118 com síndrome metabólica e 111 sem síndrome metabólica. A síndrome metabólica foi significativamente associada com baixos escores de qualidade de vida nos domínios funcionamento social (p<0,001) e aspectos emocionais (p=0,019); e com baixos escores no Componente Sumário da Saúde Mental do SF-36 (p=0,013). Entretanto, após ajustes para os fatores confundidores, especialmente índice de massa corporal, a significância dessas associações foi perdida. A única associação significativa que foi mantida entre síndrome metabólica e qualidade de vida após a análise ajustada foi com o domínio aspectos emocionais, embora somente em homens (p=0,049). CONCLUSÕES: Após o ajuste para múltiplas variáveis, a síndrome metabólica foi significativamente associada com diminuição da pontuação para qualidade de vida em homens, no domínio aspectos emocionais.

Association between leukaemia inhibitory factor gene polymorphism and pregnancy outcomes after assisted reproduction techniques.

Certain gene polymorphisms are associated with implantation failure and pregnancy loss. Studies of leukaemia inhibitory factor (LIF) gene polymorphisms are scarce. The LIF single nucleotide polymorphism (SNP) thymine (T)/guanine (G) (rs929271) was studied in women to determine whether an association existed with pregnancy outcomes after intracytoplasmic sperm injection (ICSI); 411 women who underwent ICSI were recruited. DNA was extracted from the peripheral blood, and the LIF gene SNP T/G (rs929271) was genotyped using real-time polymerase chain reaction. Participants were divided into three groups according to their LIF genotype: T/T (n = 168), T/G (n = 202) and G/G (n = 41). All IVF and ICSI procedures were carried out under the same clinical and laboratory conditions. The ICSI cumulative results (from fresh plus frozen cycles) of each genotype group were analysed. The G/G genotype in women was associated with a higher implantation rate (T/T: 15.9%, T/G: 16.2%, G/G: 27.0%; P < 0.05), ongoing pregnancy rate/patient (T/T: 31.5%, T/G: 36.1%, G/G: 53.7%; P < 0.05) and ongoing pregnancy rate/transfer (T/T: 18.5%, T/G: 20.2%, G/G: 36.7%; P < 0.05). LIF SNP T/G (rs929271) seems to be a susceptibility biomarker capable of predicting implantation efficiency and pregnancy outcomes.

Hepatitis B monovalent vaccines produced by different manufacturers: comparative study on the quality of vaccine in period before and after the shelf life / Vacinas de Hepatite B monovalente produzidas por diferentes fabricantes: estudo comparativo da qualidade das vacinas no período pré e após o prazo de validade

For over 20 years, the hepatitis B (HB) vaccine has been produced by the expression of the viral gene encoding the hepatitis B surface antigen (HBsAg) in yeast. According to the data from WHO, the hepatitis B vaccines are generally stable for up to three years when stored at 2 ºC to 8 ºC. The purpose of this study was to evaluate whether the hepatitis B vaccine, at the time of their release, the quality criteria of this product were maintained seven years after the expiration date. Vaccine vials in multi-dose (10 and 05 doses) and three lots from each manufacturer (A, B and C) were analyzed. All batches were assayed for visual appearance, potency, bacterial endotoxin, thiomersal amount, aluminum hidroxyde contents and pH by means of validated tests. The nine lots evaluated seven years after the expiration date showed similar concentrations when compared to those demonstrated at the time of batches release by the National Institute for Quality Control in Health (INCQS). No significant change in the quality of the hepatitis B vaccine after the expiration date was confirmed. These data might be useful to subsidize a future evaluation for reviewing an extension of the vaccines shelf life.

As vacinas contra a hepatite B são produzidas pela expressão do gene viral codificado para o antígeno de superfície do vírus da hepatite B (HBsAg) em levedura, há mais de 20 anos. De acordo com os dados da OMS, a vacina de hepatite B tem até três anos de estabilidade quando armazenada entre 2 ºC e 8 ºC. O objetivo deste estudo foi de avaliar se, no momento da liberação, os critérios de qualidade da vacina de hepatite B foram mantidos após sete anos da data de validade. Foram analisados frascos de vacinas multi-dose (10 e 5 doses), sendo três lotes de cada produtor (A, B e C). Todos os lotes foram avaliados quanto às características de aparência visual, potência, endotoxina bacteriana, presença de timerosal, conteúdo de hidróxido de alumínio e pH por meio de testes validados. Os nove lotes avaliados sete anos após a data de expiração tiveram resultados similares quando comparados às concentrações na época de liberação dos lotes, realizada pelo Instituto Nacional de Controle de Qualidade em Saúde (INCQS). Os estudos confirmaram a manutenção da qualidade da vacina após o período de expiração. Estes dados podem subsidiar uma futura avaliação para extensão do prazo de validade das vacinas.

Silica nanoparticle-based microfluidic immunosensor with laser-induced fluorescence detection for the quantification of immunoreactive trypsin.

The purpose of this study was to develop a silica nanoparticle-based immunosensor with laser-induced fluorescence (LIF) as a detection system. The proposed device was applied to quantify the immunoreactive trypsin (IRT) in cystic fibrosis (CF) newborn screening. A new ultrasonic procedure was used to extract the IRT from blood spot samples collected on filter papers. After extraction, the IRT reacted immunologically with anti-IRT monoclonal antibodies immobilized on a microfluidic glass chip modified with 3-aminopropyl functionalized silica nanoparticles (APSN-APTES-modified glass chips). The bounded IRT was quantified by horseradish peroxidase (HRP)-conjugated anti-IRT antibody (anti-IRT-Ab) using 10-acetyl-3,7-dihydroxyphenoxazine (ADHP) as enzymatic mediator. The HRP catalyzed the oxidation of nonfluorescent ADHP to highly fluorescent resorufin, which was measured by LIF detector, using excitation lambda at 561nm and emission at 585nm. The detection limits (LODs) calculated for LIF detection and for a commercial enzyme-linked immunosorbent assay (ELISA) test kit were 0.87 and 4.2ngml(-1), respectively. The within- and between-assay variation coefficients for the LIF detection procedure were below 6.5%. The blood spot samples collected on filter papers were analyzed with the proposed method, and the results were compared with those of the reference ELISA method, demonstrating a potential usefulness for the clinical assessment of IRT during the early neonatal period.

Variation in neuronal differentiation of a newly isolated mouse embryonic stem cell line: a detailed immunocytochemistry study.

Neural precursor differentiation from mouse ES (embryonic stem) cells have been demonstrated using EB (embryoid body), co-culture on stromal feeder layers, and in the absence of external inducing signals. Most of available mouse ES cell original research articles have worked with only six different cell lines. Our goals were to isolate one new mouse ES lineage, and perform a detailed immunocytochemistry study during neural differentiation, making use of an EB strategy protocol following the generation of neural progenitors, glial cells and postmitotic neurons. The dynamics of differentiation of ES cell derived neuronal precursors into differentiated glia cells and neurons were followed in vitro and correlated to exposure to specific elements of feeder medium. Morphological aspects of generated cellular types, including its immunocytochemical expression of differentiation markers were studied. Immuno-positivity against ß-III tubulin, PGP and TH (tyrosine hydroxylase) was observed from stage I. Approximately 80% of cells were positive for TH at stage I. The first glial cell type appears in stage III. TH, PGP or ß-III tubulin-positive cells with neuronal typical morphology only being seen in stage III when TH-positive cells corresponded to approximately 12% of total cells. Variations among other literature findings can be explained by the choice we made to use a newly isolated ES cell line. As colonies may behave differently during neuronal differentiation, it reinforces the necessity of studying original ES cell lines.

Secretory TAT-peptide-mediated protein transduction of LIF receptor α-chain distal cytoplasmic motifs into human myeloid HL-60 cells

The distal cytoplasmic motifs of leukemia inhibitory factor receptor α-chain (LIFRα-CT3) can independently induce intracellular myeloid differentiation in acute myeloid leukemia (AML) cells by gene transfection; however, there are significant limitations in the potential clinical use of these motifs due to liposome-derived genetic modifications. To produce a potentially therapeutic LIFRα-CT3 with cell-permeable activity, we constructed a eukaryotic expression pcDNA3.0-TAT-CT3-cMyc plasmid with a signal peptide (ss) inserted into the N-terminal that codes for an ss-TAT-CT3-cMyc fusion protein. The stable transfection of Chinese hamster ovary (CHO) cells via this vector and subsequent selection by Geneticin resulted in cell lines that express and secrete TAT-CT3-cMyc. The spent medium of pcDNA3.0-TAT-CT3-cMyc-transfected CHO cells could be purified using a cMyc-epitope-tag agarose affinity chromatography column and could be detected via SDS-PAGE, with antibodies against cMyc-tag. The direct administration of TAT-CT3-cMyc to HL-60 cell culture media caused the enrichment of CT3-cMyc in the cytoplasm and nucleus within 30 min and led to a significant reduction of viable cells (P < 0.05) 8 h after exposure. The advantages of using this mammalian expression system include the ease of generating TAT fusion proteins that are adequately transcripted and the potential for a sustained production of such proteins in vitro for future AML therapy.

Caracterização, mecanismo de ação e papel do fator inibidor de leucemia na fisiologia ovariana e implantação embrionária / Characterization, mechanisms of action and role of leukemia inhibitory factor in ovarian physiology and embryonic implantation

O fator inibidor de leucemia (LIF) é uma glicoproteína pertencente à família das interleucinas-6 que ligados aos seus receptores intermembranários atua em diferentes processos biológicos. Estudos têm relatado a participação deste fator durante o desenvolvimento folicular ovariano e implantação em mamíferos. Esta revisão resume os principais aspectos da estrutura molecular do LIF, seus receptores, mecanismos de ação, expressão e funções focando na fisiologia ovariana e implantação embrionária.

The leukemia inhibitory factor (LIF) is a glycoprotein from the interleukin-6 family, which acts with its membrane receptors in different biological processes. Studies have reported the involvement of this factor during ovarian follicular development and embryo implantation in mammals. This review summarizes the main aspects of the molecular structure of LIF, its receptors, mechanisms of action, expression and functions focusing on ovarian physiology and embryo implantation.