Potential unreliability of ALK variant allele frequency in the efficacy prediction of targeted therapy in NSCLC / 医学前沿

Anaplastic lymphoma kinase (ALK) is the most common fusion gene involved in non-small cell lung cancer (NSCLC), and remarkable response has been achieved with the use of ALK tyrosine kinase inhibitors (ALK-TKIs). However, the clinical efficacy is highly variable. Pre-existing intratumoral heterogeneity (ITH) has been proven to contribute to the poor treatment response and the resistance to targeted therapies. In this work, we investigated whether the variant allele frequencies (VAFs) of ALK fusions can help assess ITH and predict targeted therapy efficacy. Through the application of next-generation sequencing (NGS), 7.2% (326/4548) of patients were detected to be ALK positive. On the basis of the adjusted VAF (adjVAF, VAF normalization for tumor purity) of four different threshold values (adjVAF < 50%, 40%, 30%, or 20%), the association of ALK subclonality with crizotinib efficacy was assessed. Nonetheless, no statistical association was observed between median progression-free survival (PFS) and ALK subclonality assessed by adjVAF, and a poor correlation of adjVAF with PFS was found among the 85 patients who received first-line crizotinib. Results suggest that the ALK VAF determined by hybrid capture-based NGS is probably unreliable for ITH assessment and targeted therapy efficacy prediction in NSCLC.

Ceritinib as First-line Treatment for Advanced Lung Adenocarcinoma with COX7A2L-ALK Fusion: A Case Report and Literature Review / 中国肺癌杂志

Lung cancer is the most common in incidence and mortality worldwide. With the development of next generation sequencing (NGS) detection technology, more and more patients with rare anaplastic lymphoma kinase (ALK) fusion mutations were detected. A case of advanced lung adenocarcinoma with rare COX7A2L-ALK (C2:A20) fusion detected by NGS was reported in Peking Union Medical College Hospital, and all cases with rare ALK fusion mutations were searched from medical datebase from January 1, 2014 to March 31, 2021, to investigate the treatment of rare ALK fusion mutations with ALK inhibitors. The best response of the patient was assessed as partial response (PR) with Ceritinib treatment. By literature review, 22 cases of rare ALK fusion were reported in 19 articles. Combined with this case, 23 cases were analyzed. The objective response rate (ORR) was 82.6% (19/23) and disease control rate (DCR) was 95.7% (22/23) for rare ALK fusions patients treated with ALK inhibitors. Lung adenocarcinoma patients with rare ALK fusion could benefit from ALK inhibitors.
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Research status and prospect of immunotherapy in gastrointestinal stromal tumors / 中华胃肠外科杂志

Tyrosine kinase inhibitors (TKI) significantly reduce the risk of recurrence and metastasis and prolong survival in patients with gastrointestinal stromal tumors (GIST), but drug resistance is often inevitable. Immunotherapy has been proven effective in multiple solid tumors, but the efficacy in GIST is unclear. The efficacy of immunotherapy depends on the tumor microenvironment (TME). Tumor-infiltrating immune cells and immune checkpoints are important components of TME, which not only participate in the regulation of tumor immune response but are also the key target of immunotherapy. A comprehensive analysis of them can clarify the mechanism of tumor immune escape. This review found that there are abundant tumor-infiltrating immune cells in GIST, which play an important role in tumor immune surveillance and escape. Although early clinical studies have shown that patients with GIST have a good tolerance to immunotherapy, the curative effect is not satisfactory. Therefore, how to select the responders of immunotherapy and coordinate the relationship between immunotherapy and TKIs is the key issue to be explored. At the same time, the gradual deepening of basic research and large sample prospective clinical trials will certainly provide more strategies for the application of immunotherapy in GIST.

Effect of almonertinib on the proliferation, invasion, and migration in non-small cell lung cancer cells / 中南大学学报(医学版)

OBJECTIVES@#Lung cancer is one of the most common malignant tumors in the world, and its lethality ranks the first among many malignant tumors. For non-small cell lung cancer (NSCLC) patients, due to the high mortality rate, the overall 5-year survival rate is less than 15%. When NSCLC undergoes local invasion, the 5-year survival rate is only 20%, and it is even lower when distant metastasis occurs up to 4%. Almonertinib is an innovative drug independently researched and developed by China with independent intellectual property rights. As an epidermal growth factor receptor tyrosine kinase inhibitor, almonertinib is mainly used for locally advanced or metastatic NSCLC patients with epidermal growth factor receptor (EGFR) T790M mutation. This study aims to investigate the effects of almonertinib on the proliferation, invasion and migration of NSCLC cells in vitro.@*METHODS@#NSCLC cells H1975 and PC-9 were cultured in vitro. The effects of almonertinib on the proliferation, apoptosis, invasion, and migration of H1975 and PC-9 cells were detected by CCK-8 assay, apoptotic assay and Transwell assay. The expression of invasion and migration related proteins was detected by Western blotting.@*RESULTS@#The CCK-8 experiment showed that almonertinib inhibited the proliferation of H1975 and PC-9 cells in a time- and dose-dependent manner. The IC@*CONCLUSIONS@#Almonertinib can inhibit the proliferation, invasion, and migration of NSCLCH1975 and PC-9 cells in vitro and vivo, and promote the apoptosis of H1975 and PC-9 cells. The underlying mechanism may be related to the inhibition of tumor cell epithelial mesenchymal transformation and metalloproteinase expression.

Mining Polo-Box domain of Polo-like kinase 1 as a new therapeutic target for cancer / 生物工程学报

Polo-like kinase 1 (Plk1) is widely regarded as one of the most promising targets for cancer therapy due to its essential role in cell division and tumor cell survival. At present, most Plk1 inhibitors have been developed based on kinase domain, some of which are in clinical trial. However, inhibitors targeting kinase domain face off-target effect and drug resistance owing to the conserved nature and the frequent mutations in the ATP-binding pocket. In addition to a highly conserved kinase domain, Plk1 also contains a unique Polo-Box domain (PBD), which is essential for Plk1's subcellular localization and mitotic functions. Inhibitors targeting Plk1 PBD show stronger selectivity and less drug resistance for cancer therapy. Therefore, Plk1 PBD is an attractive target for the development of anti-cancer agents. In this review, we will summarize the up-to date drug discovery for targeting Plk1 PBD, including the molecular structure and cellular functions of Plk1 PBD. Small-molecule inhibitors targeting Plk1 PBD not only provide an opportunity to specifically inhibit Plk1 activity for cancer treatment, but also unveil novel biological basis regarding the molecular recognition of Plk1 and its substrates.

Research Advance on the Effect of Autophagy on the Survival of Chronic Myeloid Leukemia Cells--Review / 中国实验血液学杂志

Tyrosine kinase inhibitor (TKI) has significantly improved the treatment of chronic myeloid leukemia (CML), however the resistance often resulted in treatment failure. Currently, it is known that the survival of CML cells can be affected by regulating autophagy, oxidative stress and mitochondrial metabolism, among which autophagy is an evolution-conserved catabolism process, and closely related to the pathogenesis of CML, thus playing a dual role in regulating the biological characteristics of cells. On the one hand, autophagy can promote the apoptosis of CML cells, and also can induce the drug resistance of CML cells on the other hand. In this review, the effect of autophagy on CML cells was summarzed briefly, so as to provide a useful idea to explore the combination of TKI with the autophagy inhibitor or inducer to overcome the resistance of CML to TKI.

Astemizole, an inhibitor of ether-à-go-go-1 potassium channel, increases the activity of the tyrosine kinase inhibitor gefitinib in breast cancer cells

Abstract Background Expression and activity of the potassium channel ether-à-go-go-1 (EAG1) are strongly related to carcinogenesis and tumor progression, which can be exploited for therapeutic purposes. EAG1 activity may be reduced by preventing its phosphorylation with epidermal growth factor receptor (EGFR) kinase inhibitors and by astemizole, which blocks the channel pore and downregulates its gene expression. Objective We aimed to study the potential cooperative antiproliferative effect of the EGFR inhibitor gefitinib and the EAG1-blocker astemizole, in breast cancer cells. Materials and Methods The cells were characterized by immunocytochemistry. Inhibitory concentrations were determined by non-linear regression analysis using dose-response curves. The nature of the pharmacological effect was evaluated by the combination index equation while cell cycle analysis was studied by flow cytometry. Results Astemizole and gefitinib inhibited cell proliferation in a concentration-dependent manner, with inhibitory concentrations (IC 50) values of 1.72 µM and 0.51 µM, respectively. All combinations resulted in a synergistic antiproliferative effect. The combination of astemizole and gefitinib diminished the percentage of cells in G2/M and S phases, while increased accumulation in G0/G1 of the cell cycle. Conclusions Astemizole and gefitinib synergistically inhibited proliferation in breast cancer cells expressing both EGFR and EAG1. Our results suggest that the combined treatment increased cell death by targeting the oncogenic activity of EAG1.

Involvement of endothelial CK2 in the radiation induced perivascular resistant niche (PVRN) and the induction of radioresistance for non-small cell lung cancer (NSCLC) cells

BACKGROUND: Tumor microenvironment (TME) plays a vital role in determining the outcomes of radiotherapy. As an important component of TME, vascular endothelial cells are involved in the perivascular resistance niche (PVRN), which is formed by inflammation or cytokine production induced by ionizing radiation (IR). Protein kinase CK2 is a constitutively active serine/threonine kinase which plays a vital role in cell proliferation and inflammation. In this study, we investigated the potential role of CK2 in PVRN after IR exposure. RESULT: Specific CK2 inhibitors, Quinalizarin and CX-4945, were employed to effectively suppressed the kinase activity of CK2 in human umbilical vein endothelial cells (HUVECs) without affecting their viability. Results showing that conditioned medium from IR-exposed HUVECs increased cell viability of A549 and H460 cells, and the pretreatment of CK2 inhibitors slowed down such increment. The secretion of IL-8 and IL-6 in HUVECs was induced after exposure with IR, but significantly inhibited by the addition of CK2 inhibitors. Furthermore, IR exposure elevated the nuclear phosphorylated factor-κB (NF-κB) p65 expression in HUVECs, which was a master factor regulating cytokine production. But when pretreated with CK2 inhibitors, such elevation was significantly suppressed. CONCLUSION: This study indicated that protein kinase CK2 is involved in the key process of the IR induced perivascular resistant niche, namely cytokine production, by endothelial cells, which finally led to radioresistance of non-small cell lung cancer cells. Thus, the inhibition of CK2 may be a promising way to improve the outcomes of radiation in nonsmall cell lung cancer cells.

The effect of hirudin on antagonisting thrombin induced apoptosis of human microvascular endothelial cells

Abstract Purpose: To investigate whether hirudin exerts its antithrombin action to decrease the ratio of Human Microvascular Endothelial Cells (HMVECs) apoptosis. Methods: Human microvascular endothelial cells (HMVECs) cultured in the third and fifth generations were used. HMVECs were divided into normal group, thrombin group (T group), natrual hirudin group (H group), thrombin + natrual hirudin group (T + H group), AG490 group, thrombin + AG490 group (T + AG490 group), natrual hirudin + AG490 group (H + AG490 group), thrombin + natural hirudin + AG490 (T + H + AG490 group).Apart from the normal group, the other groups were exposed to the relevant drugs for 24 hours.HMVEC apoptosis was assessed by flow cytometric and double Immunofluorescence of phosphorylation of JAK (P-JAK2) and TUNEL assay. Results: Compared with the normal group, in thrombin group the HMVECs apoptosis rate were significantly increased (P<0.05).The results indicated that the index of apoptosis and the apoptosis rate were improved in cultures treated by natural hirudin (T + H group), relative to cultures with thrombin only (T group). We found that the index of apoptosis and the apoptosis rate in the AG490 + thrombin group were higher than that in the hirudin + thrombin group (P<0.05). Double Immunofluorescence of p-JAK2 and TUNEL assays showed that cells were double positive for P-JAK2 uptake and TUNEL detection liquid binding. Conclusion: The natural hirudin and JAK2/STATs signal inhibitor AG490 could block the effects of thrombin. Natural hirudin could attenuate HMVECs apoptosis via antagonizing thrombin and it is suggested that this effect may occur by blocking the JAK2/STATs signaling pathway and this signaling pathways appears to be not the only pathway.

The Application Methods of Mitogen-activated Protein Kinase Signal Pathway Inhibitors SP600125 and SB203580 in Long-term in Vivo Experiments / Métodos de aplicación de los inhibidores SP600125 y SB203580 de la vía de señalización de la proteína quinasa activada por mitógeno en experimentos in vivo a largo plazo

ABSTRACT Objective: To explore the application methods of mitogen-activated protein kinase signal pathway inhibitors SP600125 and SB203580 in long-term in vivo experiments. Methods: A total of 55 healthy New Zealand rabbits were randomly divided into blank control group, model control group, SP low dose group, SP high dose group, SP blank group, SB low dose group, SB high dose group, SB blank group, dimethyl sulfoxide (DMSO) control group, DMSO blank group, and positive control group. Since the first day of the experiment, each group was administered the corresponding treatment for four weeks continuously. Then, the myocardial c-Jun N-terminal kinase (JNK) and the total protein of p38, protein phosphorylation and its gene expression levels were detected. Results: After intravenous treatment with adriamycin, the myocardial phosphorylate-JNK (p-JNK) and phosphorylate-p38 (p-p38) levels in all groups were increased to varying degrees, of which the model control group increased the most significantly (p < 0.05). Compared with the model control group, the myocardial p-JNK and p-p38 increased more slowly in the SP low dose group, SP high dose group, SB low dose group, SB high dose group and positive control group (p < 0.05), of which the increase in the SP high dose group and the SB high dose group was the slowest (p < 0.05). After four weeks, the total protein and messenger ribonucleic acid of the myocardial JNK and p38 in all groups had no statistically significant difference (p > 0.05). Conclusion: The continuous intravenous injection of SP600125 and SB203580 for four weeks significantly reduced the protein phosphorylation levels of JNK and p38, which provides a practical avenue for the long-term study in vivo.

RESUMEN Objetivo: Explorar los métodos de aplicación de los inhibidores SP600125 y SB203580 de la vía de señalización de la proteína quinasa activada por mitógeno en experimentos in vivo a largo plazo. Métodos: Un total de 55 conejos sanos de Nueva Zelandia fueron divididos aleatoriamente en los grupos siguientes: grupo de control en blanco, grupo de control modelo, grupo de dosis baja SP, grupo de dosis alta SP, grupo en blanco SP, grupo de dosis baja SB, grupo de dosis alta SB, grupo en blanco SB, grupo de control dimetilsulfóxido (DMSO), grupo en blanco DMSO, y grupo de control positivo. Desde el primer día del experimento, a cada grupo se le administró el tratamiento correspondiente por cuatro semanas continuas. Entonces, se detectaron la quinasa c-Jun N-terminal (JNK) miocárdica y la proteína p38 total, así como la fosforilación proteica y sus niveles de expresión génica. Resultados: Después del tratamiento intravenoso con adriamicina, los niveles de fosfo-JNK (p-JNK) y fosfo-p38 (p-p38) del miocardio aumentaron en todos los grupos en diversos grados, siendo el aumento del grupo de control modelo el más significativo (p < 0.05). En comparación con el grupo de control modelo, p-JNK y p-p38 miocárdicos aumentaron más lentamente en el grupo de dosis baja SP, el grupo de dosis alta SP, el grupo de dosis baja SB, el grupo de dosis alta SB, y el grupo de control positivo (p < 0.05). De estos, el aumento en el grupo de dosis alta SP y el grupo de dosis alta SB fue el más lento (p < 0.05). Después de cuatro semanas, la proteína total y el ácido ribonucleico mensajero de JNK y p38 miocárdicos en todos los grupos, no tuvieron diferencias significativas (p > 0.05). Conclusión: La inyección intravenosa continua de SP600125 y SB203580 durante cuatro semanas redujo significativamente los niveles de fosforilación proteica de JNK y p38, lo que proporciona una vía práctica para el estudio a largo plazo in vivo.

La inhibición de Rho quinasa post infarto mejora el remodelado y la función ventricular: mecanismos involucrados a nivel preclínico / Rho-kinase inhibition post myocardial infarction improves remodeling and systolic function: a preclinical study of intervening factors in a pre-clinical study

Resumen: Objetivo: Determinar algunos mecanismos moleculares por los cuales la activación de ROCK cardíaca post infarto del miocardio (IAM) participa en el remodelado y en deterioro de la función sistólica. Métodos: Determinación simultánea de niveles de proteínas blanco de ROCK cardíaca, de función sistólica in vivo del ventrículo izquierdo (VI) y de fibrosis e hipertrofia cardíaca en ratas con IAM en condiciones de inhibición de ROCK con fasudil. Resultados : Siete días post IAM la masa ventricular relativa aumentó significativamente en un 30% en el grupo MI y se redujo con fasudil. La disfunción sistólica VI mejoró significativamente con fasudil mientras que la activación de ROCK cardíaca se redujo a niveles del grupo control. El inhibidor de ROCK también redujo significativamente los niveles cardíacos elevados de las isoformas ROCK1 y ROCK2, de MHC-β y del colágeno miocárdico. En el grupo con IAM aumentaron significativamente los niveles de fosforilación de ERK 42 y ERK 44 (en 2 veces y en 63%, respectivamente), mientras que en el grupo IAM tratado con fasudil estos niveles fueron similares a los del grupo control. El IAM aumentó significativamente los niveles fosforilados del factor de transcripción GATA-4, que se normalizaron con el inhibidor de ROCK. Conclusiones: La disfunción sistólica post IAM se asoció fuertemente con la activación del ROCK cardíaca y con la fosforilación de proteínas río abajo de ROCK que promueven remodelado cardíaco como β-MHC y la vía ERK / GATA-4.

Abstracts: Objective: to determine some molecular mechanisms by which cardiac ROCK activation after myocardial infarction (MI) intervene in cardiac systolic function decline and remodeling. Methods: simultaneous measurement of different cardiac ROCK target proteins levels, in vivo left ventricular (LV) systolic function, myocardial fibrosis, and hypertrophy in rats with MI under ROCK inhibition with fasudil were performed. Results: seven days after MI the relative ventricular mass increased significantly by 30% in the MI groupand was reduced with fasudil. LV systolic dysfunction improved significantly with fasudil whereas at the same time cardiac ROCK activation was reduced to sham levels. The ROCK inhibitor also reduced increased cardiac levels of both ROCK1 and ROCK2 isoforms, β-MHC levels and myocardial collagen volume fraction decline. MI significantly increased phosphorylation levels of ERK 42 and ERK 44 by 2-fold and 63% respectively whereas in the fasudil-treated MI group these levels were similar to those in the sham group. MI significantly increased phosphorylated levels of the transcription factor GATA-4 which were normalyzed by the ROCK inhibitor. Conclusion: LV systolic dysfunction after MI was strongly associated to cardiac ROCK activation and subsequent phosphorylation of ROCK target proteins that promote ventricular remodeling, such as β-MHC and the ERK/GATA-4 pathway. ROCK inhibition with fasudil significantly improved systolic function, diminished myocardial fibrosis, and normalized β-MHC and ERK/GATA-4 phosphorylation levels.

Angiotensin III increases monocyte chemoattractant protein-1 expression in cultured human proximal tubular epithelial cells

BACKGROUND/AIMS: We investigated whether angiotensin III (Ang III) is involved in monocyte recruitment through regulation of the chemokine monocyte chemoattractant protein-1 (MCP-1) in cultured human proximal tubular epithelial cells (HK-2 cells). METHODS: We measured MCP-1 levels in HK-2 cells that had been treated with various concentrations of Ang III and Ang II type-1 (AT1) receptor antagonists at various time points. The phosphorylation states of p38, c-Jun N-terminal kinases (JNK), and extracellular-signal-regulated kinases were measured in Ang III-treated cells to explore the mitogen-activated protein kinase (MAPK) pathway. MCP-1 levels in HK-2 cell-conditioned media were measured after pre-treatment with the transcription factor inhibitors curcumin or pyrrolidine dithiocarbamate. RESULTS: Ang III increased MCP-1 protein production in dose- and time-dependent manners in HK-2 cells, which was inhibited by the AT1 receptor blocker losartan. p38 MAPK activity increased significantly in HK-2 cells exposed to Ang III for 30 minutes, and was sustained at higher levels after 60 minutes (p < 0.05). Total phosphorylated JNK protein levels tended to increase 20 minutes after stimulation with Ang III. Pre-treatment with a p38 inhibitor, a JNK inhibitor, or curcumin significantly inhibited Ang III-induced MCP-1 production. CONCLUSIONS: Ang III increases MCP-1 synthesis via stimulation of intracellular p38 and JNK MAPK signaling activity and subsequent activated protein-1 transcriptional activity in HK-2 cells.

Use of deferasirox, an iron chelator, to overcome imatinib resistance of chronic myeloid leukemia cells

BACKGROUND/AIMS: The treatment of chronic myeloid leukemia (CML) has achieved impressive success since the development of the Bcr-Abl tyrosine kinase inhibitor, imatinib mesylate. Nevertheless, resistance to imatinib has been observed, and a substantial number of patients need alternative treatment strategies. METHODS: We have evaluated the effects of deferasirox, an orally active iron chelator, and imatinib on K562 and KU812 human CML cell lines. Imatinib-resistant CML cell lines were created by exposing cells to gradually increasing concentrations of imatinib. RESULTS: Co-treatment of cells with deferasirox and imatinib induced a synergistic dose-dependent inhibition of proliferation of both CML cell lines. Cell cycle analysis showed an accumulation of cells in the subG1 phase. Western blot analysis of apoptotic proteins showed that co-treatment with deferasirox and imatinib induced an increased expression of apoptotic proteins. These tendencies were clearly identified in imatinib-resistant CML cell lines. The results also showed that co-treatment with deferasirox and imatinib reduced the expression of BcrAbl, phosphorylated Bcr-Abl, nuclear factor-kappaB (NF-kappaB) and beta-catenin. CONCLUSIONS: We observed synergistic effects of deferasirox and imatinib on both imatinib-resistant and imatinib-sensitive cell lines. These effects were due to induction of apoptosis and cell cycle arrest by down-regulated expression of NF-kappaB and beta-catenin levels. Based on these results, we suggest that a combination treatment of deferasirox and imatinib could be considered as an alternative treatment option for imatinib-resistant CML.

Anti-Proliferative Effects of Rutin on OLETF Rat Vascular Smooth Muscle Cells Stimulated by Glucose Variability

PURPOSE: Proliferation of vascular smooth muscle cells (VSMCs) plays a crucial role in atherosclerosis. Rutin is a major representative of the flavonol subclass of flavonoids and has various pharmacological activities. Currently, data are lacking regarding its effects on VSMC proliferation induced by intermittent hyperglycemia. Here, we demonstrate the effects of rutin on VSMC proliferation and migration according to fluctuating glucose levels. MATERIALS AND METHODS: Primary cultures of male Otsuka Long-Evans Tokushima Fatty (OLETF) rat VSMCs were obtained from enzymatically dissociated rat thoracic aortas. VSMCs were incubated for 72 h with alternating normal (5.5 mmol/L) and high (25.0 mmol/L) glucose media every 12 h. Proliferation and migration of VSMCs, the proliferative molecular pathway [including p44/42 mitogen-activated protein kinases (MAPK), mitogen-activated protein kinase kinase 1/2 (MEK1/2), p38 MAPK, phosphoinositide 3-kinase (PI3K), c-Jun N-terminal protein kinase (JNK), nuclear factor kappa B (NF-kappaB), and Akt], the migratory pathway (big MAPK 1, BMK1), reactive oxygen species (ROS), and apoptotic pathway were analyzed. RESULTS: We found enhanced proliferation and migration of VSMCs when cells were incubated in intermittent high glucose conditions, compared to normal glucose. These effects were lowered upon rutin treatment. Intermittent treatment with high glucose for 72 h increased the expression of phospho-p44/42 MAPK (extracellular signal regulated kinase 1/2, ERK1/2), phospho-MEK1/2, phospho-PI3K, phospho-NF-kappaB, phospho-BMK1, and ROS, compared to treatment with normal glucose. These effects were suppressed by rutin. Phospho-p38 MAPK, phospho-Akt, JNK, and apoptotic pathways [B-cell lymphoma (Bcl)-xL, Bcl-2, phospho-Bad, and caspase-3] were not affected by fluctuations in glucose levels. CONCLUSION: Fluctuating glucose levels increased proliferation and migration of OLETF rat VSMCs via MAPK (ERK1/2), BMK1, PI3K, and NF-kappaB pathways. These effects were inhibited by the antioxidant rutin.

Angiotensin II Modulates p130Cas of Podocytes by the Suppression of AMP-Activated Protein Kinase

Angiotensin II (Ang II) induces the pathological process of vascular structures, including renal glomeruli by hemodynamic and nonhemodynamic direct effects. In kidneys, Ang II plays an important role in the development of proteinuria by the modification of podocyte molecules. We have previously found that Ang II suppressed podocyte AMP-activated protein kinase (AMPK) via Ang II type 1 receptor and MAPK signaling pathway. In the present study, we investigated the roles of AMPK on the changes of p130Cas of podocyte by Ang II. We cultured mouse podocytes and treated them with various concentrations of Ang II and AMPK-modulating agents and analyzed the changes of p130Cas by confocal imaging and western blotting. In immunofluorescence study, Ang II decreased the intensity of p130Cas and changed its localization from peripheral cytoplasm into peri-nuclear areas in a concentrated pattern in podocytes. Ang II also reduced the amount of p130Cas in time and dose-sensitive manners. AMPK activators, metformin and AICAR, restored the suppressed and mal-localized p130Cas significantly, whereas, compound C, an AMPK inhibitor, further aggravated the changes of p130Cas. Losartan, an Ang II type 1 receptor antagonist, recovered the abnormal changes of p130Cas suppressed by Ang II. These results suggest that Ang II induces the relocalization and suppression of podocyte p130Cas by the suppression of AMPK via Ang II type 1 receptor, which would contribute to Ang II-induced podocyte injury.

Hemodynamic responses during and after multiple sets of stretching exercises performed with and without the Valsalva maneuver

OBJECTIVE: This study investigated the acute hemodynamic responses to multiple sets of passive stretching exercises performed with and without the Valsalva maneuver. METHODS: Fifteen healthy men aged 21 to 29 years with poor flexibility performed stretching protocols comprising 10 sets of maximal passive unilateral hip flexion, sustained for 30 seconds with equal intervals between sets. Protocols without and with the Valsalva maneuver were applied in a random counterbalanced order, separated by 48-hour intervals. Hemodynamic responses were measured by photoplethysmography pre-exercise, during the stretching sets, and post-exercise. RESULTS: The effects of stretching sets on systolic and diastolic blood pressure were cumulative until the fourth set in protocols performed with and without the Valsalva maneuver. The heart rate and rate pressure product increased in both protocols, but no additive effect was observed due to the number of sets. Hemodynamic responses were always higher when stretching was performed with the Valsalva maneuver, causing an additional elevation in the rate pressure product. CONCLUSIONS: Multiple sets of unilateral hip flexion stretching significantly increased blood pressure, heart rate, and rate pressure product values. A cumulative effect of the number of sets occurred only for systolic and diastolic blood pressure, at least in the initial sets of the stretching protocols. The performance of the Valsalva maneuver intensified all hemodynamic responses, which resulted in significant increases in cardiac work during stretching exercises. .

The perceptions of medical researchers on qualitative methodologies / Percepción de los investigadores médicos en metodologías cualitativas / Percepção de pesquisadores médicos sobre metodologias qualitativas

We aimed to verify doctor's perception of the qualitative research method, via a qualitative study of interviews with questions on the academic profile of doctors and on the methodology. We interviewed 42 professionals, of which 18 had experience with the qualitative method and 24 with the quantitative method. The results showed that knowledge on the qualitative method was virtually nil among "quantitative researchers", who did not value qualitative research, although some of those realized that it would be important to be more accepting in clinical practice. Others only considered the method as subsidiary to quantitative. The majority considered qualitative methods as lacking academic structure, taking too long to conduct empirical studies, and being difficult to publish. All of them criticized the misuse of the method, and the "quantitatives" pointed out the problem of being unable to reproduce. We concluded that widening the use of the qualitative method by doctors requires investment from the beginning of the academic career and participation in qualitative research projects.

El objetivo es verificar la percepción de médicos sobre el método de investigación cualitativa. Se trata de un estudio cualitativo por medio de entrevistas con preguntas sobre el perfil de los médicos y sobre el método. Entrevistamos a 42 profesionales, 18 con experiencia en el método cualitativo y 24 con el cuantitativo. Los resultados mostraron que el conocimiento sobre lo cualitativo es casi nulo entre los "cuantitativistas", que no valoran la investigación cualitativa, aunque algunos se dan cuenta de que sería importante tener un enfoque más amplio en la práctica clínica. Otros la ven como subsidiaria a lo cuantitativo. Sus dificultades para utilizar ese abordaje son: falta de formación, cantidad de tiempo que exigen y problemas de publicación. Todos han criticado el mal uso del método. Los "cuantitativistas" han destacado como fragilidad, la no reproductibilidad. Llegamos a la conclusión de que para ampliar el uso de los abordajes cualitativos entre los médicos es importante invertir en su formación desde el inicio del curso y la participación en proyectos de investigación cualitativa.

Objetivamos verificar a percepção de médicos sobre o método qualitativo de pesquisa. Estudo qualitativo por meio de entrevistas com questões sobre o perfil acadêmico do médico e perguntas abertas a respeito do método. Entrevistamos 42 profissionais, sendo 18 com experiência no método qualitativo e 24 com o quantitativo. Os resultados evidenciaram que o conhecimento sobre o qualitativo é quase nulo entre os pesquisadores "quantitativistas", os quais não valorizam a pesquisa qualitativa, embora alguns percebam que seria importante ter uma postura mais compreensiva na prática clínica. Outros só a veem como subsidiária ao quantitativo. As principais dificuldades da maioria são: falta de formação, tempo longo despendido nos estudos empíricos e dificuldade de publicação. Todos os entrevistados criticaram o mau uso do método, e os "quantitativistas" ressaltaram, como problema, sua não reprodutibilidade. Concluímos que ampliar o uso do método qualitativo por médicos exige investimento na formação desde o início da graduação e participação em projetos de pesquisa qualitativa.

Children's eating behavior: comparison between normal and overweight children from a school in Pelotas, Rio Grande do Sul, Brazil / Comportamento alimentar infantil: comparação entre crianças sem e com excesso de peso em uma escola do município de Pelotas, RS

OBJECTIVE: To evaluate differences in children's eating behavior in relation to their nutritional status, gender and age. METHODS: Male and female children aged six to ten years were included. They were recruited from a private school in the city of Pelotas, Rio Grande do Sul, southern Brazil, in 2012. Children´s Eating Behaviour Questionnaire (CEBQ) subscales were used to assess eating behaviors: Food Responsiveness (FR), Enjoyment of Food (EF), Desire to Drink (DD), Emotional Overeating (EOE), Emotional Undereating (EUE), Satiety Responsiveness (SR), Food Fussiness (FF) and Slowness in Eating (SE). Age-adjusted body mass index (BMI) z-scores were calculated according to the WHO recommendations to assess nutritional status. RESULTS: The study sample comprised 335 children aged 87.9±10.4 months and 49.3% had normal weight (n=163), 26% were overweight (n=86), 15% were obese (n=50) and 9.7% were severely obese (n=32). Children with excess weight showed higher scores at the CEBQ subscales associated with "food approach" (FR, EF, DD, EOE, p<0.001) and lower scores on two "food avoidance" subscales (SR and SE, p<0.001 and p=0.003, respectively) compared to normal weight children. Differences in the eating behavior related to gender and age were not found. CONCLUSIONS: "Food approach" subscales were positively associated to excess weight in children, but no associations with gender and age were found. .

OBJETIVO: Avaliar diferenças no comportamento alimentar infantil em função do estado nutricional, do sexo e da idade. MÉTODOS: O estudo incluiu crianças na faixa de seis a dez anos, de ambos os sexos, de uma escola privada em Pelotas (RS), em 2012. Para avaliar o comportamento alimentar usaram-se as subescalas do questionário Children's Eating Behaviour Questionnaire (CEBQ): resposta à comida (FR), prazer de comer (EF), desejo de beber (DD), sobreingestão emocional (EOE), subingestão emocional (EUE), resposta à saciedade (SR), seletividade (FF) e ingestão lenta (SE). Avaliou-se o estado nutricional por meio do escore-z do IMC/idade. RESULTADOS: Participaram 335 crianças de 87,9±10,4 meses. Apresentaram eutrofia 49,3% (n=163), sobrepeso 26% (n=86), obesidade 15% (n=50) e obesidade grave 9,7% (n=32). Crianças com excesso de peso tiveram maior pontuação nas subescalas de "interesse pela comida" (FR, EF, DD, EOE, p<0,001) e menor pontuação nas subescalas de "desinteresse pela comida" (SR e SE, p<0,001 e p=0,003, respectivamente), se comparadas com as crianças com peso adequado. Não foram observadas diferenças no comportamento alimentar segundo sexo e idade. CONCLUSÕES: Observou-se que comportamentos alimentares que refletem "interesse pela comida" estão associados positivamente ao excesso de peso, mas não foi encontrada associação com o sexo e a idade da criança. .

Risk factors associated with structural postural changes in the spinal column of children and adolescents / Fatores de risco associados a alterações posturais estruturais da coluna vertebral em crianças e adolescentes

OBJECTIVE: To investigate the association between behavioral risk factors, specifically postural habits, with the presence of structural changes in the spinal column of children and adolescents. METHODS: 59 students were evaluated through the self-reporting Back Pain and Body Posture Evaluation Instrument and spinal panoramic radiographic examination. Spine curvatures were classified based on Cobb angles, as normal or altered in the saggital plane and as normal or scoliotic in the frontal plane. Data were analyzed using SPSS 18.0, based on descriptive statistics and chi-square association test (a=0,05). RESULTS: The prevalence of postural changes was 79.7% (n=47), of which 47.5% (n=28) showed frontal plane changes and 61% (n=36) sagital plane changes. Significant association was found between the presence of thoracic kyphosis and female gender, practice of physical exercises only once or twice a week, sleep time greater than 10 hours, inadequate postures when sitting on a seat and sitting down to write, and how school supplies are carried. Lumbar lordosis was associated with the inadequate way of carrying the school backpack (asymmetric); and scoliosis was associated wuth the practice of competitive sports and sleep time greater than 10 hours. CONCLUSIONS: Lifestyle may be associated with postural changes. It is important to develop health policies in order to reduce the prevalence of postural changes, by decreasing the associated risk factors. .

OBJETIVO: Verificar se existe associação de fatores de risco comportamentais, especificamente hábitos posturais, com a presença de alteração postural estrutural na coluna vertebral de crianças e adolescentes. MÉTODOS: Foram avaliados 59 crianças e adolescentes, que responderam ao questionário auto-aplicável Back Pain and Body Posture Evaluation Instrument e fizeram o exame radiográfico panorâmico da coluna vertebral. De acordo com o ângulo de Cobb, as curvaturas sagitais da coluna vertebral foram classificadas como normais ou alteradas e, no plano frontal, como escolioses ou normais. Os dados foram analisados no SPSS 18.0, a partir de estatística descritiva e do teste de associação qui-quadrado (a=0,05). RESULTADOS: A prevalência de alterações posturais foi de 79,7% (n=47), 47,5% (n=28) apresentavam alteração no plano frontal e 61% (n=36) no sagital. Foi encontrada associação entre cifose torácica e sexo feminino, prática de exercício físico apenas uma ou duas vezes na semana, tempo de sono superior a 10 horas, posturas inadequadas para sentar no banco e sentar para escrever e o meio de transporte do material escolar. Para lordose lombar, observou-se associação com o transporte da mochila escolar de modo inadequado (assimétrico). Houve associação significativa entre a presença de escoliose com a prática de esporte competitivo e o tempo de sono superior a 10 horas. CONCLUSÕES: Hábitos de vida podem estar associados a alterações posturais, é importante o desenvolvimento políticas de saúde a fim de reduzir a prevalência de alterações posturais por meio da redução dos fatores de risco associados. .

Neuropsychomotor developmental delay: conceptual map, term definitions, uses and limitations / Atraso do desenvolvimento neuropsicomotor: mapa conceitual, definições, usos e limitações do termo

OBJECTIVE: To retrieve the origin of the term neuropsychomotor developmental delay" (NPMD), its conceptual evolution over time, and to build a conceptual map based on literature review. DATA SOURCE: A literature search was performed in the SciELO Brazil, Web of Science, Science Direct, OneFile (GALE), Pubmed (Medline), Whiley Online, and Springer databases, from January of 1940 to January of 2013, using the following keywords: NPMD delay, NPMD retardation, developmental delay, and global developmental delay. A total of 71 articles were selected, which were used to build the conceptual map of the term. DATA SYNTHESIS: Of the 71 references, 55 were international and 16 national. The terms developmental delay and global developmental delay were the most frequently used in the international literature and, in Brazil, delayed NPMD was the most often used. The term developmental delay emerged in the mid 1940s, gaining momentum in the 1990s. In Brazil, the term delayed NPMD started to be used in the 1980s, and has been frequently cited and published in the literature. Delayed development was a characteristic of 13 morbidities described in 23 references. Regarding the type of use, 19 references were found, with seven forms of use. Among the references, 34 had definitions of the term, and 16 different concepts were identified. CONCLUSIONS: Developmental delay is addressed in the international and national literature under different names, various applications, and heterogeneous concepts. Internationally, ways to improve communication between professionals have been indicated, with standardized definition of the term and use in very specific situations up to the fifth year of life, which was not found in Brazilian publications. .

OBJETIVO: Resgatar a origem do termo atraso do desenvolvimento neuropsicomotor (DNPM), sua evolução conceitual ao longo do tempo e construir mapa conceitual do termo com base em busca bibliográfica. FONTES DE DADOS: Foi realizada busca nas bases de dados eletrônicas do Portal da Capes, que incluem Scielo Brazil, Web of Science, Science Direct, OneFile (GALE), Pubmed (Medline), Whiley Online e Springer, referente a Janeiro/1940-Janeiro/2013. Palavras-chave: atraso e retardo do DNPM, developmental delay e global developmental delay. Foram selecionados 71 artigos e construído o mapa conceitual do termo. SÍNTESE DE DADOS: Das 71 referências, 55 eram internacionais e 16 nacionais. Os termos mais encontrados foram global developmental delay e developmental delay na literatura internacional e retardo e atraso do DNPM no Brasil. Internacionalmente, o termo surgiu em meados da década de 40 ganhando força nos anos 90. No Brasil, o termo começou a ser usado na década de 80 e vem sendo frequentemente citado na literatura. O atraso é citado em 23 trabalhos como característica presente em 13 tipos de condições clínicas. Com relação ao uso, foram encontrados 19 estudos, com sete situações de uso. Dentre os artigos revisados, 34 deles apresentaram definições, sendo identificados 16 conceitos diferentes. CONCLUSÕES: O atraso do desenvolvimento é abordado na literatura internacional e nacional sob diversos nomes, diferentes aplicações e conceitos heterogêneos. Internacionalmente, apontam-se caminhos para melhorar a comunicação entre profissionais, com definição padronizada do termo e uso em situações específicas até o quinto ano de vida, o que não foi encontrado nas publicações nacionais. .