Enhanced specialized metabolite, trichome density, and biosynthetic gene expression in Stevia rebaudiana (Bertoni) Bertoni plants inoculated with endophytic bacteria Enterobacter hormaechei.

Stevia rebaudiana (Bertoni) Bertoni is a plant of economic interest in the food and pharmaceutical industries due its steviol glycosides (SG), which are rich in metabolites that are 300 times sweeter than sucrose. In addition, S. rebaudiana plants contain phenolic compounds and flavonoids with antioxidant activity. Endophytic bacteria promote the growth and development and modulate the metabolism of the host plant. However, little is known regarding the role of endophytic bacteria in the growth; synthesis of SG, flavonoids and phenolic compounds; and the relationship between trichome development and specialized metabolites in S. rebaudiana, which was the subject of this study. The 12 bacteria tested did not increase the growth of S. rebaudiana plants; however, the content of SG increased with inoculation with the bacteria Enterobacter hormaechei H2A3 and E. hormaechei H5A2. The SG content in leaves paralleled an increase in the density of glandular, short, and large trichome. The image analysis of S. rebaudiana leaves showed the presence of SG, phenolic compounds, and flavonoids principally in glandular and short trichomes. The increase in the transcript levels of the KO, KAH, UGT74G1, and UGT76G1 genes was related to the SG concentration in plants of S. rebaudiana inoculated with E. hormaechei H2A3 and E. hormaechei H5A2. In conclusion, inoculation with the stimulating endophytes E. hormaechei H2A3 and E. hormaechei H5A2 increased SG synthesis, flavonoid content and flavonoid accumulation in the trichomes of S. rebaudiana plants.

AC-265347 Inhibits Neuroblastoma Tumor Growth by Induction of Differentiation without Causing Hypocalcemia.

Neuroblastoma is the most common extracranial solid tumor of childhood, with heterogeneous clinical manifestations ranging from spontaneous regression to aggressive metastatic disease. The calcium-sensing receptor (CaSR) is a G protein-coupled receptor (GPCR) that senses plasmatic fluctuation in the extracellular concentration of calcium and plays a key role in maintaining calcium homeostasis. We have previously reported that this receptor exhibits tumor suppressor properties in neuroblastoma. The activation of CaSR with cinacalcet, a positive allosteric modulator of CaSR, reduces neuroblastoma tumor growth by promoting differentiation, endoplasmic reticulum (ER) stress and apoptosis. However, cinacalcet treatment results in unmanageable hypocalcemia in patients. Based on the bias signaling shown by calcimimetics, we aimed to identify a new drug that might exert tumor-growth inhibition similar to cinacalcet, without affecting plasma calcium levels. We identified a structurally different calcimimetic, AC-265347, as a promising therapeutic agent for neuroblastoma, since it reduced tumor growth by induction of differentiation, without affecting plasma calcium levels. Microarray analysis suggested biased allosteric modulation of the CaSR signaling by AC-265347 and cinacalcet towards distinct intracellular pathways. No upregulation of genes involved in calcium signaling and ER stress were observed in patient-derived xenografts (PDX) models exposed to AC-265347. Moreover, the most significant upregulated biological pathways promoted by AC-265347 were linked to RHO GTPases signaling. AC-265347 upregulated cancer testis antigens (CTAs), providing new opportunities for CTA-based immunotherapies. Taken together, this study highlights the importance of the biased allosteric modulation when targeting GPCRs in cancer. More importantly, the capacity of AC-265347 to promote differentiation of malignant neuroblastoma cells provides new opportunities, alone or in combination with other drugs, to treat high-risk neuroblastoma patients.

Impact of Circulatory Assistance in the Early Evolution After Heart Transplantation. Unicentric Experience.

BACKGROUND: Heart transplantation (HT) is the reference treatment for patients with terminal heart failure. In recent years there has been a progressive increase in HT procedures in patients who have a circulatory support (CS). METHODS: This is a retrospective single-center study of 293 consecutive patients who underwent HT from 2009 to 2018, analyzing the evolution of the 2 cohorts: patients with and without CS as a bridge to HT. Baseline and evolutionary clinical data collected following the usual follow-up protocol were recorded, including clinical events observed during the follow-up 1 year after the procedure. RESULTS: The subgroup of patients transplanted with CS showed a higher incidence of primary graft failure, frequent infection, and mortality. A tendency toward lower cardiac allograft vasculopathy was observed in this subgroup. Mechanical ventilation added to the CS resulted in a higher incidence of primary graft failure, infection, and renal dysfunction. The CS variable as a bridge to HT was shown to be predictive of 1-year mortality in both univariate (odds ratio, 1.84; 95% confidence interval, 1.03-3.3; P = .038) and multivariate (odds ratio, 2.1; 95% confidence interval, 1.01-4.3; P = .047) analyses. CONCLUSIONS: In our experience, CS as a bridge to HT results in a higher incidence of primary graft failure, frequent infection, and mortality at 1-year follow-up. Mechanical ventilation added to CS has a clear unfavorable prognostic impact. CS as a bridge to HT was shown to be predictive of 1-year mortality in both univariate and multivariate analyses.

Chemical analysis of callus extracts from toxic and non-toxic varieties of Jatropha curcas L.

Jatropha curcas L. belongs to Euphorbiaceae family, and it synthesizes flavonoid and diterpene compounds that have showed antioxidant, anti-inflammatory, anticancer, antiviral, antimicrobial, antifungal and insecticide activity. Seeds of this plant accumulate phorbol esters, which are tigliane type diterpenes, reported as toxic and, depending on its concentration, toxic and non-toxic varieties has been identified. The aim of this work was to characterize the chemical profile of the extracts from seeds, leaves and callus of both varieties (toxic and non-toxic) of Jatropha curcas, to verify the presence of important compounds in dedifferentiated cells and consider the possibility of using these cultures for the massive production of metabolites. Callus induction was obtained using NAA (1.5 mg L-1) and BAP (1.5 mg L-1) after 21 d for both varieties. Thin layer chromatography analysis showed differences in compounds accumulation in callus from non-toxic variety throughout the time of culture, diterpenes showed an increase along the time, in contrast with flavonoids which decreased. Based on the results obtained through microQTOF-QII spectrometer it is suggested a higher accumulation of phorbol esters, derived from 12-deoxy-16-hydroxy-phorbol (m/z 365 [M+H]+), in callus of 38 d than those of 14 d culture, from both varieties. Unlike flavonoids accumulation, the MS chromatograms analysis allowed to suggest lower accumulation of flavonoids as the culture time progresses, in callus from both varieties. The presence of six glycosylated flavonoids is also suggested in leaf and callus extracts derived from both varieties (toxic and non-toxic), including: apigenin 6-C-α-L-arabinopyranosyl-8-C-ß -D-xylopyranoside (m/z 535 [M+H]+), apigenin 4'-O-rhamnoside (m/z 417 [M+H]+), vitexin (m/z 433 [M+H]+), vitexin 4'-O-glucoside-2″-O-rhamnoside (m/z 741 [M+H]+), vicenin-2 (m/z 595 [M+H]+), and vicenin-2,6″-O-glucoside (m/z 757 [M+H]+).

Parathyroid hormone-like hormone plays a dual role in neuroblastoma depending on PTH1R expression.

We have previously reported the expression of parathyroid hormone-like hormone (PTHLH) in well-differentiated, Schwannian stroma-rich neuroblastic tumors. The aim of this study was to functionally assess the role of PTHLH and its receptor, PTH1R, in neuroblastoma. Stable knockdown of PTHLH and PTH1R was conducted in neuroblastoma cell lines to investigate the succeeding phenotype induced both in vitro and in vivo. Downregulation of PTHLH reduced MYCN expression and subsequently induced cell cycle arrest, senescence, and migration and invasion impairment in a MYCN-amplified, TP53-mutated neuroblastoma cell line. These phenotypes were associated with reduced tumorigenicity in a murine model. We also show that PTHLH expression is not under the control of the calcium-sensing receptor in neuroblastoma. Conversely, its production is stimulated by epidermal growth factor receptor (EGFR). Accordingly, irreversible EGFR inhibition with canertinib abolished PTHLH expression. The oncogenic role of PTHLH appeared to be a consequence of its intracrine function, as downregulation of its receptor, PTH1R, increased anchorage-independent growth and induced a more undifferentiated, invasive phenotype. Respectively, high PTH1R mRNA expression was found in MYCN nonamplified primary tumors and also significantly associated with other prognostic factors of good outcome. This study provides the first evidence of the dual role of PTHLH in the behavior of neuroblastomas. Moreover, the identification of EGFR as a transcriptional regulator of PTHLH in neuroblastoma provides a novel therapeutic opportunity to promote a less aggressive tumor phenotype through irreversible inhibition of EGFR tyrosine kinase activity.

Evaluación de la Escala EFD-66 como herramienta para predecir la adherencia al tratamiento en pacientes con enfermedades crónicas no transmisibles

Objetivo. Los objetivos del presente estudio son validar la construcción de la escala de fases de duelo (EFD-66) como instrumento para medir las etapas de duelo (ED) y evaluar su utilidad para discriminar entre pacientes con enfermedad crónica no transmisible (ECNT) que cumplen con adherencia al tratamiento farmacológico (ATF) en la consulta médica ambulatoria. Métodos. Se realizó un estudio transversal para determinar la asociación entre las ED y la ATF, en los meses de abril a octubre de 2015. La recogida de datos se realizó mediante un diseño prolectivo. Se aplicaron tres instrumentos: una cédula sociodemográfica, la escala EFD-66 y el cuestionario de Morisky-Green. Se reclutaron pacientes con antecedente de ECNT procedentes de la Clínica de Medicina Familiar “Gustavo A. Madero”, en la Ciudad de México. Los datos se analizaron con las pruebas estadísticas apropiadas. Resultados. Se incluyó un total de 165 pacientes. Se observó que altos puntajes en las subescalas de negación (razón de momios [RM]: 1,124; intervalo de confianza de 95% [IC95%]: 1,066-1,186; P < 0,001), de ira (RM: 1,157; IC95%: 1,080-1,240; P < 0,001) y de depresión (RM: 1,071; IC95%: 1,029-1,116; P = 0,001) se asocian con falta de ATF; sin embargo, un puntaje elevado en la subescala de aceptación (RM: 0,913; IC95%: 0,880-0,948; P < 0,001) se asocia con ATF. La mayor sensibilidad entre las subescalas se observó en las etapas de negación e ira (área bajo la curva [ABC]: 0,597 en ambas). Conclusiones. La EFD-66 es un instrumento que presenta una buena validez de construcción como instrumento para medir las ED y permite detectar pacientes con ECNT que cumplen con ATF, por lo que recomendamos su aplicación en la consulta médica ambulatoria. Además, nuestros hallazgos indican que el duelo es un factor de riesgo que incrementa la falta de ATF.

Objective. The objectives of this study are to validate the construct of the stages of grief scale (FD-66) as an instrument for measuring the stages of grief and to evaluate its usefulness in discriminating among patients with chronic non-communicable diseases in terms of adherence to the pharmacological treatment prescribed. Methods. A cross-sectional study was conducted from April to October 2015 to determine the association between the stages of grief and treatment adherence. Data were collected using a prolective design. Three instruments were applied: a sociodemographic document, the FD-66 scale, and the Morisky-Green questionnaire. Patients with a history of CNCDs were recruited from the Gustavo A. Madero Family Medicine Clinic in Mexico City. The data were analyzed using the appropriate statistical tests. Results. A total of 165 patients were included. It was observed that high scores on the subscales of denial (odds ratio [OR]: 1.124; confidence interval of 95% [CI95%]: 1.066-1.186; P < 0,001); anger (OR: 1.157; CI95%: 1.080-1.240; P < 0.001), and depression (OR: 1.071; CI95%: 1.029-1.116; P = 0.001) are associated with poor treatment adherence; however, a high score on the acceptance subscale (OR: 0.913; CI95%: 0.880-0.948; P < 0.001) is associated with good treatment adherence. The greatest sensitivity among the subscales was observed in the denial and anger stages (area under the [ABC] curve: 0.597 in both). Conclusions. The FD-66 is an instrument with good construct validity as a tool for measuring the stages of grief and makes it possible to identify patients with CNCD that will adhere to treatment. We therefore recommend its use in outpatient medical consultations. Furthermore, our findings indicate that grief is a risk factor that increases poor treatment adherence.

Objetivo. Validar a construção da Escala facial de dor (EFD-66) como instrumento para medir as fases da dor e avaliar sua utilidade para discriminar, na consulta médica ambulatorial, os pacientes com doenças crônicas não transmissíveis (DCNT) que aderem ao tratamento medicamentoso. Métodos. Estudo transversal para determinar a associação entre as fases da dor e o tratamento medicamentoso conduzido de abril a outubro de 2015. A coleta de dados foi realizada prospectivamente. Foram aplicados três instrumentos: uma ficha sociodemográfica, a escala EFD-66 e o teste de Morisky-Green. Foram recrutados pacientes com história de DCNT provenientes da Clínica de Medicina da Família “Gustavo A. Madero”, na Cidade do México. Os dados foram analisados com os testes estatísticos apropriados. Resultados. Foi estudada uma amostra de 165 pacientes. Observou-se que uma pontuação alta nas subescalas de negação (odds ratio [OR] 1,124; intervalo de confiança de 95% [IC95%] 1,066–1,186; P < 0,001), raiva (OR 1,157; IC95% 1,080–1,240; P < 0,001) e depressão (OR 1,071; IC95% 1,029–1,116; P = 0,001) está associada à não adesão ao tratamento medicamentoso. No entanto, uma pontuação alta na subescala de aceitação (OR 0,913; IC95% 0,880–0,948; P < 0,001) está associada à adesão ao tratamento medicamentoso. As subescalas com maior sensibilidade foram as fases de negação e raiva (área sob a curva [ASC]: 0,597 em ambas). Conclusões. A escala EFD-66 tem boa validade de construção como instrumento para medir as fases da dor e permite discriminar os pacientes com DCNT que aderem ao tratamento medicamentosos. Assim, recomendamos que esta escala seja aplicada em consultas médicas ambulatoriais. Além disso, nossos achados indicam que a dor é um fator de risco que contribui para a não adesão ao tratamento medicamentoso.

Loss of the EPH receptor B6 contributes to colorectal cancer metastasis.

Although deregulation of EPHB signaling has been shown to be an important step in colorectal tumorigenesis, the role of EPHB6 in this process has not been investigated. We found here that manipulation of EPHB6 levels in colon cancer cell lines has no effect on their motility and growth on a solid substrate, soft agar or in a xenograft mouse model. We then used an EphB6 knockout mouse model to show that EphB6 inactivation does not efficiently initiate tumorigenesis in the intestinal tract. In addition, when intestinal tumors are initiated genetically or pharmacologically in EphB6+/+ and EphB6-/- mice, no differences were observed in animal survival, tumor multiplicity, size or histology, and proliferation of intestinal epithelial cells or tumor cells. However, reintroduction of EPHB6 into colon cancer cells significantly reduced the number of lung metastasis after tail-vein injection in immunodeficient mice, while EPHB6 knockdown in EPHB6-expressing cells increased their metastatic spread. Consistently, although EPHB6 protein expression in a series of 130 primary colorectal tumors was not associated with patient survival, EPHB6 expression was significantly lower in lymph node metastases compared to primary tumors. Our results indicate that the loss of EPHB6 contributes to the metastatic process of colorectal cancer.

Jatropha curcas and assisted phytoremediation of a mine tailing with biochar and a mycorrhizal fungus.

Soil pollution is an important ecological problem worldwide. Phytoremediation is an environmental-friendly option for reducing metal pollution. A greenhouse experiment was conducted to determine the growth and physiological response, metal uptake, and the phytostabilization potential of a nontoxic Jatropha curcas L. genotype when grown in multimetal-polluted conditions. Plants were established on a mine residue (MR) amended or not amended with corn biochar (B) and inoculated or not inoculated with the mycorrhizal fungus Acaulospora sp. (arbuscular mycorrhizal fungus, AMF). J. curcas was highly capable of growing in an MR and showed no phytotoxic symptoms. After J. curcas growth (105 days), B produced high desorption of Cd and Pb from the MR; however, no increases in metal shoot concentrations were observed. Therefore, Jatropha may be useful for phytostabilization of metals in mine tailings. The use of B is recommended because improved MR chemical properties conduced to plant growth (cation-exchange capacity, organic matter content, essential nutrients, electrical conductivity, water-holding capacity) and plant growth development (higher biomass, nutritional and physiological performance). Inoculation with an AMF did not improve any plant growth or physiological plant characteristic. Only higher Zn shoot concentration was observed, but it was not phytotoxic. Future studies of B use and its long-term effect on MR remediation should be conducted under field conditions.

[Evaluation of the FD-66 Scale as a tool for predicting treatment adherence in patients with chronic non-communicable diseasesAvaliação da escala EFD-66 como instrumento para predizer a adesão ao tratamento em pacientes com doenças crônicas não transmissíveis]. / Evaluación de la Escala EFD-66 como herramienta para predecir la adherencia al tratamiento en pacientes con enfermedades crónicas no transmisibles.

OBJECTIVE: The objectives of this study are to validate the construct of the stages of grief scale (FD-66) as an instrument for measuring the stages of grief and to evaluate its usefulness in discriminating among patients with chronic non-communicable diseases in terms of adherence to the pharmacological treatment prescribed. METHODS: A cross-sectional study was conducted from April to October 2015 to determine the association between the stages of grief and treatment adherence. Data were collected using a prolective design. Three instruments were applied: a sociodemographic document, the FD-66 scale, and the Morisky-Green questionnaire. Patients with a history of CNCDs were recruited from the Gustavo A. Madero Family Medicine Clinic in Mexico City. The data were analyzed using the appropriate statistical tests. RESULTS: A total of 165 patients were included. It was observed that high scores on the subscales of denial (odds ratio [OR]: 1.124; confidence interval of 95% [CI95%]: 1.066-1.186; P < 0,001); anger (OR: 1.157; CI95%: 1.080-1.240; P < 0.001), and depression (OR: 1.071; CI95%: 1.029-1.116; P = 0.001) are associated with poor treatment adherence; however, a high score on the acceptance subscale (OR: 0.913; CI95%: 0.880-0.948; P < 0.001) is associated with good treatment adherence. The greatest sensitivity among the subscales was observed in the denial and anger stages (area under the [ABC] curve: 0.597 in both). CONCLUSIONS: The FD-66 is an instrument with good construct validity as a tool for measuring the stages of grief and makes it possible to identify patients with CNCD that will adhere to treatment. We therefore recommend its use in outpatient medical consultations. Furthermore, our findings indicate that grief is a risk factor that increases poor treatment adherence.

OBJETIVO: Validar a construção da Escala facial de dor (EFD-66) como instrumento para medir as fases da dor e avaliar sua utilidade para discriminar, na consulta médica ambulatorial, os pacientes com doenças crônicas não transmissíveis (DCNT) que aderem ao tratamento medicamentoso. MÉTODOS: Estudo transversal para determinar a associação entre as fases da dor e o tratamento medicamentoso conduzido de abril a outubro de 2015. A coleta de dados foi realizada prospectivamente. Foram aplicados três instrumentos: uma ficha sociodemográfica, a escala EFD-66 e o teste de Morisky-Green. Foram recrutados pacientes com história de DCNT provenientes da Clínica de Medicina da Família "Gustavo A. Madero", na Cidade do México. Os dados foram analisados com os testes estatísticos apropriados. RESULTADOS: Foi estudada uma amostra de 165 pacientes. Observou-se que uma pontuação alta nas subescalas de negação (odds ratio [OR] 1,124; intervalo de confiança de 95% [IC95%] 1,066­1,186; P < 0,001), raiva (OR 1,157; IC95% 1,080­1,240; P < 0,001) e depressão (OR 1,071; IC95% 1,029­1,116; P = 0,001) está associada à não adesão ao tratamento medicamentoso. No entanto, uma pontuação alta na subescala de aceitação (OR 0,913; IC95% 0,880­0,948; P < 0,001) está associada à adesão ao tratamento medicamentoso. As subescalas com maior sensibilidade foram as fases de negação e raiva (área sob a curva [ASC]: 0,597 em ambas). CONCLUSÕES: A escala EFD-66 tem boa validade de construção como instrumento para medir as fases da dor e permite discriminar os pacientes com DCNT que aderem ao tratamento medicamentosos. Assim, recomendamos que esta escala seja aplicada em consultas médicas ambulatoriais. Além disso, nossos achados indicam que a dor é um fator de risco que contribui para a não adesão ao tratamento medicamentoso.

Cell Culture of Bursera linanoe in a Stirred Tank Bioreactor for Production of Linalool and Linalyl Acetate.

Bursera linanoe cell suspension cultures were initiated from callus grown in Murashige and Skoog medium supplemented with naphthalene acetic acid (3.0 mg L⁻¹) and 6-benzylaminopurine (0.5 mg L⁻¹). In flasks, B. linanoe cell cultures grew over a 9 day period, reaching a maximum biomass of 11.16 g DW L⁻¹. Throughout the growth phase, cell viability was constant at 60 - 70%. In contrast, B. linanoe cells growing in a bioreactor achieved a maximum biomass of 22.26 g DW L⁻¹ (after 7 days), and cell viability was constant at 75 - 85%. Production of linalool and linalyl acetate in the bioreactor (3.02 and 2.40 mg g⁻¹ DW, respectively) was significantly greater than that achieved from cells in flask cultures (1.05 and 0.97 mg g⁻¹ DW, respectively). B. linanoe cell suspension culture has potential as an alternative method for the production of essential oils.

Evaluación de la Escala EFD-66 como herramienta para predecir la adherencia al tratamiento en pacientes con enfermedades crónicas no transmisibles / Evaluation of the FD-66 Scale as a tool for predicting treatment adherence in patients with chronic non-communicable diseases / Avaliação da escala EFD-66 como instrumento para predizer a adesão ao tratamento em pacientes com doenças crônicas não transmissíveis

RESUMEN Objetivo Los objetivos del presente estudio son validar la construcción de la escala de fases de duelo (EFD-66) como instrumento para medir las etapas de duelo (ED) y evaluar su utilidad para discriminar entre pacientes con enfermedad crónica no transmisible (ECNT) que cumplen con adherencia al tratamiento farmacológico (ATF) en la consulta médica ambulatoria. Métodos Se realizó un estudio transversal para determinar la asociación entre las ED y la ATF, en los meses de abril a octubre de 2015. La recogida de datos se realizó mediante un diseño prolectivo. Se aplicaron tres instrumentos: una cédula sociodemográfica, la escala EFD-66 y el cuestionario de Morisky-Green. Se reclutaron pacientes con antecedente de ECNT procedentes de la Clínica de Medicina Familiar "Gustavo A. Madero", en la Ciudad de México. Los datos se analizaron con las pruebas estadísticas apropiadas. Resultados Se incluyó un total de 165 pacientes. Se observó que altos puntajes en las subescalas de negación (razón de momios [RM]: 1,124; intervalo de confianza de 95% [IC95%]: 1,066-1,186; P < 0,001), de ira (RM: 1,157; IC95%: 1,080-1,240; P < 0,001) y de depresión (RM: 1,071; IC95%: 1,029-1,116; P = 0,001) se asocian con falta de ATF; sin embargo, un puntaje elevado en la subescala de aceptación (RM: 0,913; IC95%: 0,880-0,948; P < 0,001) se asocia con ATF. La mayor sensibilidad entre las subescalas se observó en las etapas de negación e ira (área bajo la curva [ABC]: 0,597 en ambas). Conclusiones La EFD-66 es un instrumento que presenta una buena validez de construcción como instrumento para medir las ED y permite detectar pacientes con ECNT que cumplen con ATF, por lo que recomendamos su aplicación en la consulta médica ambulatoria. Además, nuestros hallazgos indican que el duelo es un factor de riesgo que incrementa la falta de ATF.

ABSTRACT Objective The objectives of this study are to validate the construct of the stages of grief scale (FD-66) as an instrument for measuring the stages of grief and to evaluate its usefulness in discriminating among patients with chronic non-communicable diseases in terms of adherence to the pharmacological treatment prescribed. Methods A cross-sectional study was conducted from April to October 2015 to determine the association between the stages of grief and treatment adherence. Data were collected using a prolective design. Three instruments were applied: a sociodemographic document, the FD-66 scale, and the Morisky-Green questionnaire. Patients with a history of CNCDs were recruited from the Gustavo A. Madero Family Medicine Clinic in Mexico City. The data were analyzed using the appropriate statistical tests. Results A total of 165 patients were included. It was observed that high scores on the subscales of denial (odds ratio [OR]: 1.124; confidence interval of 95% [CI95%]: 1.066-1.186; P < 0,001); anger (OR: 1.157; CI95%: 1.080-1.240; P < 0.001), and depression (OR: 1.071; CI95%: 1.029-1.116; P = 0.001) are associated with poor treatment adherence; however, a high score on the acceptance subscale (OR: 0.913; CI95%: 0.880-0.948; P < 0.001) is associated with good treatment adherence. The greatest sensitivity among the subscales was observed in the denial and anger stages (area under the [ABC] curve: 0.597 in both). Conclusions The FD-66 is an instrument with good construct validity as a tool for measuring the stages of grief and makes it possible to identify patients with CNCD that will adhere to treatment. We therefore recommend its use in outpatient medical consultations. Furthermore, our findings indicate that grief is a risk factor that increases poor treatment adherence.

RESUMO Objetivo Validar a construção da Escala facial de dor (EFD-66) como instrumento para medir as fases da dor e avaliar sua utilidade para discriminar, na consulta médica ambulatorial, os pacientes com doenças crônicas não transmissíveis (DCNT) que aderem ao tratamento medicamentoso. Métodos Estudo transversal para determinar a associação entre as fases da dor e o tratamento medicamentoso conduzido de abril a outubro de 2015. A coleta de dados foi realizada prospectivamente. Foram aplicados três instrumentos: uma ficha sociodemográfica, a escala EFD-66 e o teste de Morisky-Green. Foram recrutados pacientes com história de DCNT provenientes da Clínica de Medicina da Família "Gustavo A. Madero", na Cidade do México. Os dados foram analisados com os testes estatísticos apropriados. Resultados Foi estudada uma amostra de 165 pacientes. Observou-se que uma pontuação alta nas subescalas de negação (odds ratio [OR] 1,124; intervalo de confiança de 95% [IC95%] 1,066-1,186; P < 0,001), raiva (OR 1,157; IC95% 1,080-1,240; P < 0,001) e depressão (OR 1,071; IC95% 1,029-1,116; P = 0,001) está associada à não adesão ao tratamento medicamentoso. No entanto, uma pontuação alta na subescala de aceitação (OR 0,913; IC95% 0,880-0,948; P < 0,001) está associada à adesão ao tratamento medicamentoso. As subescalas com maior sensibilidade foram as fases de negação e raiva (área sob a curva [ASC]: 0,597 em ambas). Conclusões A escala EFD-66 tem boa validade de construção como instrumento para medir as fases da dor e permite discriminar os pacientes com DCNT que aderem ao tratamento medicamentosos. Assim, recomendamos que esta escala seja aplicada em consultas médicas ambulatoriais. Além disso, nossos achados indicam que a dor é um fator de risco que contribui para a não adesão ao tratamento medicamentoso.

Caveolin-1 promotes Ewing sarcoma metastasis regulating MMP-9 expression through MAPK/ERK pathway.

Ewing sarcoma (ES) is a bone and soft tissue sarcoma affecting mostly children and young adults. Caveolin-1 (CAV1) is a well-known target of EWS/FLI1, the main driver of ES, with an oncogenic role in ES. We have previously described how CAV1 is able to induce metastasis in ES via matrix metalloproteinase-9 (MMP-9). In the present study we showed how CAV1 silencing in ES reduced MEK1/2 and ERK1/2 phosphorylation. Accordingly, chemical inhibition of MEK1/2 resulted in reduction in MMP-9 expression and activity that correlated with reduced migration and invasion. IQ Motif Containing GTPase Activating Protein 1 (IQGAP1) silencing reduced MEK1/2 and ERK1/2 phosphorylation and MMP-9 expression. Furthermore, IQGAP1 silenced cells showed a marked decrease in their migratory and invasive capacity. We demonstrated that CAV1 and IQGAP1 localize in close proximity at the cellular edge, thus IQGAP1 could be the connecting node between CAV1 and MEK/ERK in ES metastatic phenotype. Analysis of the phosphorylation profile of CAV1-silenced cells showed a decrease of p-ribosomal protein S6 (RPS6). RPS6 can be phosphorylated by p90 ribosomal S6 kinases (RSK) proteins. CAV1-silenced cells showed reduced levels of p-RSK1 and treatment with U0126 provoked the same effect. Despite not affecting ERK1/2 and RPS6 phosphorylation status neither MMP-9 expression nor activity, RSK1 silencing resulted in a reduced migratory and invasive capacity in vitro and reduced incidence of metastases in vivo in a novel orthotopic model. The present work provides new insights into CAV1-driven metastatic process in ES unveiling novel key nodes.

Regulation of Differentiation by Calcium-Sensing Receptor in Normal and Tumoral Developing Nervous System.

During normal development of the nervous system (NS), neural progenitor cells (NPCs) produce specialized populations of neurons and glial cells upon cell fate restriction and terminal differentiation. These sequential processes require the dynamic regulation of thousands of genes. The calcium-sensing receptor (CaSR) is temporally and spatially regulated in both neurons and glial cells during development of the NS. In particular, CaSR expression and function have been shown to play a significant role during differentiation of NPCs toward the oligodendrocyte lineage and also in maturation of cerebellar granule cell precursors (GCPs). Moreover, CaSR regulates axonal and dendritic growth in both central and peripheral nervous systems (PNSs), a process necessary for proper construction of mature neuronal networks. On the other hand, several lines of evidence support a role for CaSR in promotion of cell differentiation and inhibition of proliferation in neuroblastoma, a tumor arising from precursor cells of developing PNS. Thus, among the variety of NS functions in which the CaSR participates, this mini-review focuses on its role in differentiation of normal and tumoral cells. Current knowledge of the mechanisms responsible for CaSR regulation and function in these contexts is also discussed, together with the therapeutic opportunities provided by CaSR allosteric modulators.

Cinacalcet inhibits neuroblastoma tumor growth and upregulates cancer-testis antigens.

The calcium-sensing receptor is a G protein-coupled receptor that exerts cell-type specific functions in numerous tissues and some cancers. We have previously reported that this receptor exhibits tumor suppressor properties in neuroblastoma. We have now assessed cinacalcet, an allosteric activator of the CaSR approved for clinical use, as targeted therapy for this developmental tumor using neuroblastoma cell lines and patient-derived xenografts (PDX) with different MYCN and TP53 status. In vitro, acute exposure to cinacalcet induced endoplasmic reticulum stress coupled to apoptosis via ATF4-CHOP-TRB3 in CaSR-positive, MYCN-amplified cells. Both phenotypes were partially abrogated by phospholipase C inhibitor U73122. Prolonged in vitro treatment also promoted dose- and time-dependent apoptosis in CaSR-positive, MYCN-amplified cells and, irrespective of MYCN status, differentiation in surviving cells. Cinacalcet significantly inhibited tumor growth in MYCN-amplified xenografts and reduced that of MYCN-non amplified PDX. Morphology assessment showed fibrosis in MYCN-amplified xenografts exposed to the drug. Microarrays analyses revealed up-regulation of cancer-testis antigens (CTAs) in cinacalcet-treated MYCN-amplified tumors. These were predominantly CTAs encoded by genes mapping on chromosome X, which are the most immunogenic. Other modulated genes upon prolonged exposure to cinacalcet were involved in differentiation, cell cycle exit, microenvironment remodeling and calcium signaling pathways. CTAs were up-regulated in PDX and in vitro models as well. Moreover, progressive increase of CaSR expression upon cinacalcet treatment was seen both in vitro and in vivo. In summary, cinacalcet reduces neuroblastoma tumor growth and up-regulates CTAs. This effect represents a therapeutic opportunity and provides surrogate circulating markers of neuroblastoma response to this treatment.

RHOA inactivation enhances Wnt signalling and promotes colorectal cancer.

Activation of the small GTPase RHOA has strong oncogenic effects in many tumour types, although its role in colorectal cancer remains unclear. Here we show that RHOA inactivation contributes to colorectal cancer progression/metastasis, largely through the activation of Wnt/ß-catenin signalling. RhoA inactivation in the murine intestine accelerates the tumorigenic process and in human colon cancer cells leads to the redistribution of ß-catenin from the membrane to the nucleus and enhanced Wnt/ß-catenin signalling, resulting in increased proliferation, invasion and de-differentiation. In mice, RHOA inactivation contributes to colon cancer metastasis and reduced RHOA levels were observed at metastatic sites compared with primary human colon tumours. Therefore, we have identified a new mechanism of activation of Wnt/ß-catenin signalling and characterized the role of RHOA as a novel tumour suppressor in colorectal cancer. These results constitute a shift from the current paradigm and demonstrate that RHO GTPases can suppress tumour progression and metastasis.

Caveolin-1 is down-regulated in alveolar rhabdomyosarcomas and negatively regulates tumor growth.

Rhabdomyosarcoma is the most common soft tissue sarcoma of childhood and adolescence. Despite advances in therapy, patients with histological variant of rhabdomyosarcoma known as alveolar rhabdomyosarcoma (ARMS) have a 5-year survival of less than 30%. Caveolin-1 (CAV1), encoding the structural component of cellular caveolae, is a suggested tumor suppressor gene involved in cell signaling. In the present study we report that compared to other forms of rhabdomyosarcoma (RMS) CAV1 expression is either undetectable or very low in ARMS cell lines and tumor samples. DNA methylation analysis of the promoter region and azacytidine-induced re-expression suggest the involvement of epigenetic mechanisms in the silencing of CAV1. Reintroduction of CAV1 in three of these cell lines impairs their clonogenic capacity and promotes features of muscular differentiation. In vitro, CAV1-expressing cells show high expression of Caveolin-3 (CAV3), a muscular differentiation marker. Blockade of MAPK signaling is also observed. In vivo, CAV1-expressing xenografts show growth delay, features of muscular differentiation and increased cell death. In summary, our results suggest that CAV1 could function as a potent tumor suppressor in ARMS tumors. Inhibition of CAV1 function therefore, could contribute to aberrant cell proliferation, leading to ARMS development.

EphA2-induced angiogenesis in ewing sarcoma cells works through bFGF production and is dependent on caveolin-1.

Angiogenesis is the result of the combined activity of the tumor microenvironment and signaling molecules. The angiogenic switch is represented as an imbalance between pro- and anti-angiogenic factors and is a rate-limiting step in the development of tumors. Eph receptor tyrosine kinases and their membrane-anchored ligands, known as ephrins, constitute the largest receptor tyrosine kinase (RTK) subfamily and are considered a major family of pro-angiogenic RTKs. Ewing sarcoma (EWS) is a highly aggressive bone and soft tissue tumor affecting children and young adults. As other solid tumors, EWS are reliant on a functional vascular network for the delivery of nutrients and oxygen and for the removal of waste. Based on the biological roles of EphA2 in promoting angiogenesis, we explored the functional role of this receptor and its relationship with caveolin-1 (CAV1) in EWS angiogenesis. We demonstrated that lack of CAV1 results in a significant reduction in micro vascular density (MVD) on 3 different in vivo models. In vitro, this phenomenon correlated with inactivation of EphA2 receptor, lack of AKT response and downregulation of bFGF. We also demonstrated that secreted bFGF from EWS cells acted as chemoattractant for endothelial cells. Furthermore, interaction between EphA2 and CAV1 was necessary for the right localization and signaling of the receptor to produce bFGF through AKT and promote migration of endothelial cells. Finally, introduction of a dominant-negative form of EphA2 into EWS cells mostly reproduced the effects occurred by CAV1 silencing, strongly suggesting that the axis EphA2-CAV1 participates in the promotion of endothelial cell migration toward the tumors favoring EWS angiogenesis.

The dual inhibitory effect of thiostrepton on FoxM1 and EWS/FLI1 provides a novel therapeutic option for Ewing's sarcoma.

The poor prognosis of Ewing's sarcoma (EWS), together with its high lethal recurrence rate and the side­effects of current treatments, call for novel targeted therapies with greater curative effectiveness and substantially reduced side­effects. The oncogenic chimeric protein EWS/FLI1 is the key malignancy driver in most EWSs, regulating numerous target genes, many of which influence cell cycle progression. It has often been argued that targeting proteins regulated directly or indirectly by EWS/FLI1 may provide improved therapeutic options for EWS. In this context, our study examined FoxM1, a key cell cycle regulating transcription factor, reported to be expressed in EWS and influenced by EWS/FLI1. Thiostrepton, a naturally occurring small molecule, has been shown to selectively inhibit FoxM1 expression in cancer cells. We demonstrate that in EWS, in addition to inhibiting FoxM1 expression, thiostrepton downregulates the expression of EWS/FLI1, both at the mRNA and protein levels, leading to cell cycle arrest and, ultimately, to apoptotic cell death. We also show that thiostrepton treatment reduces the tumorigenicity of EWS cells, significantly delaying the growth of nude mouse xenograft tumors. Results from this study demonstrate a novel action of thiostrepton as inhibitor of the expression of the EWS/FLI1 oncoprotein in vitro and in vivo, and that it shows greater efficacy against EWS than against other tumor types, as it is active on EWS cells and tumors at concentrations lower than those reported to have effective inhibitory activity on tumor cells derived from other cancers. Owing to the dual action of this small molecule, our findings suggest that thiostrepton may be particularly effective as a novel agent for the treatment of EWS patients.

A propagation procedure for Cuphea aequipetala Cav. (Lythraceae) and antioxidant properties of wild and greenhouse-grown plants / Procedimiento para la propagación de Cuphea aequipetala Cav. (Lythraceae) y propiedades antioxidantes de plantas silvestres y crecidas en invernadero

Cuphea aequipetala Cav. (Lythraceae) is native to Mexico and is used in folk medicine to treat tumors. An efficient protocol for in vitro shoot proliferation and plant acclimatization of C. aequipetala was developed. Total phenolic compounds and flavonoids contents were determined in methanolic extracts of roots, stems, and leaves from wild and greenhouse-grown plants. Their antioxidant properties were compared using in vitro assays (scavenging of 2,2-Diphenyl-1-picrylhydrazyl (DPPH) and 2,2'-Azino-bis (3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) radicals, and reducing power in the phosphomolybdenum assay). This is the first report of a successful propagation procedure for C. aequipetala. These methods offer a viable approach for long-term in vitro conservation and proliferation of this species. C. aequipetala shoots maintained their proliferation capacity during long-term subculture (3 years). The propagated shoots can successfully acclimatize and grow to maturity, and they retain the ability to accumulate antioxidants.

Cuphea aequipetala Cav. (Lythraceae) es una planta nativa de México que se utiliza en la medicina tradicional para tratar tumores. En este trabajo se desarrolló un procedimiento para la proliferación de brotes y la aclimatización de plantas de C. aequipetala. Se determinó la concentración de compuestos fenólicos totales y de flavonoides en extractos metanólicos de raíces, tallos y hojas de plantas silvestres y crecidas en invernadero. Sus propiedades antioxidantes fueron comparadas utilizando ensayos in vitro (captura de radicales DPPH y ABTS y poder reductor por el ensayo de fosfomolibdeno). Este es el primer reporte exitoso sobre un procedimiento para la propagación de C. aequipetala. Este método ofrece una alternativa viable para la conservación a largo plazo y la proliferación de esta especie. Los brotes de C. aequipetala han mantenido su capacidad de multiplicación a largo plazo (tres años). Los brotes se convirtieron en plantas adultas aclimatadas, manteniendo su habilidad para acumular compuestos antioxidantes.

Brush border myosin Ia inactivation in gastric but not endometrial tumors.

Brush border Myosin Ia (MYO1A) has been shown to be frequently mutated in colorectal tumors with microsatellite instability (MSI) and to have tumor suppressor activity in intestinal tumors. Here, we investigated the frequency of frameshift mutations in the A8 microsatellite in exon 28 of MYO1A in MSI gastric and endometrial tumors and found a high mutation rate in gastric (22/47; 46.8%) but not endometrial (3/48; 6.2%) tumors. Using a regression model, we show that MYO1A mutations are likely to confer a growth advantage to gastric, but not endometrial tumors. The mutant MYO1A(7A) protein was shown to lose its membrane localization in gastric cancer cells and a cycloheximide-chase assay demonstrated that the mutant MYO1A(7A) protein has reduced stability compared to the wild type MYO1A. Frequent MYO1A promoter hypermethylation was also found in gastric tumors. Promoter methylation negatively correlates with MYO1A mRNA expression in a series of 58 non-MSI gastric primary tumors (Pearson's r = -0.46; p = 0.0003) but not in a cohort of 54 non-MSI endometrial tumors and treatment of gastric cancer cells showing high MYO1A promoter methylation with the demethylating agent 5-aza-2'-deoxycytidine, resulted in a significant increase of MYO1A mRNA levels. We found that normal gastric epithelial cells, but not normal endometrial cells, express high levels of MYO1A. Therefore, when considered together, our findings suggest that MYO1A has tumor suppressor activity in the normal gastric epithelium but not in the normal endometrium and inactivation of MYO1A either genetically or epigenetically may confer gastric epithelial cells a growth advantage.