Anthropometric Characteristics, Body Composition and Nutritional Status of Younger Primary School Children in Montenegro: National Study / Características Antropométricas, Composición Corporal y Estado Nutricional de los Niños más Pequeños de la Escuela Primaria en Montenegro. Estudio Nacional

SUMMARY: This article has two aims: (a) first aim was to determine what is the most applicable and the simplest alternative for recommended BMI categories for underweight, overweight and obesity related to IOTF references, from the practical standpoint; (b) second aim was to determine the prevalence of the nutritional status in Montenegro on this representative sample of school children aged 9-13 years and compare them with peers from relevant and similar studies from both the local region and globally. A total sample of 1478 healthy children from Montenegro participated in this study divided into two sub-samples of 732 girls and 746 boys. According to the IOTF body mass index (BMI) reference values were used through ROC curve analysis to evaluate potential alternatives for estimation of the nutritional status of this sample of children. Only WHtR did not show significant age-related differences in the case of both genders. Considering the nutritional status of children from this study it has been found that boys have a considerably higher prevalence of being overweight (22.7 % vs. 16.4 %) and obese (7.5 % vs. 3.3 %) compared to girls. On the other hand, girls were more prevalent to be underweight (10.5 % vs. 7.5 %). WHtR seems like the best alternative for the estimation of obesity and being overweight due to simplicity and the equipment needed.

Este artículo tiene dos objetivos: (a) el primero fue determinar cuál es la alternativa más aplicable y más sencilla para las categorías de IMC recomendadas para bajo peso, sobrepeso y obesidad relacionadas con las referencias de la IOTF, desde el punto de vista práctico; (b) el segundo objetivo fue determinar la prevalencia del estado nutricional en Montenegro en esta muestra representativa de escolares de 9 a 13 años y compararlos con pares de estudios relevantes y similares tanto de la región local como a nivel mundial. En el estudio participaron 1478 niños sanos de Montenegro divididos en dos submuestras de 732 niñas y 746 niños. De acuerdo con el índice de masa corporal (IMC) de la IOTF, se utilizaron valores de referencia mediante análisis de curvas ROC para evaluar posibles alternativas para la estimación del estado nutricional de esta muestra en niños. Sólo el ICT no mostró diferencias significativas relacionadas con la edad en el caso de ambos sexos. Teniendo en consideración el estado nutricional de los niños, se determinó que los éstos tenían una prevalencia considerablemente mayor de sobrepeso (22,7 % frente a 16,4 %) y obesidad (7,5 % frente a 3,3 %) en comparación con las niñas. Por otro lado, las niñas tenían más prevalencia de bajo peso (10,5 % frente a 7,5 %). El WHtR parece la mejor alternativa para la estimación de la obesidad y el sobrepeso por su sencillez y equipamiento necesario.

Biochemical and molecular responses of the ascorbate-glutathione cycle in wheat seedlings exposed to different forms of selenium.

Biofortification aims to increase selenium (Se) concentration and bioavailability in edible parts of crops such as wheat (Triticum aestivum L.), resulting in increased concentration of Se in plants and/or soil. Higher Se concentrations can disturb protein structure and consequently influence glutathione (GSH) metabolism in plants which can affect antioxidative and other detoxification pathways. The aim of this study was to elucidate the impact of five different concentrations of selenate and selenite (0.4, 4, 20, 40 and 400 mg kg-1) on the ascorbate-glutathione cycle in wheat shoots and roots and to determine biochemical and molecular tissue-specific responses. Content of investigated metabolites, activities of detoxification enzymes and expression of their genes depended both on the chemical form and concentration of the applied Se, as well as on the type of plant tissue. The most pronounced changes in the expression level of genes involved in GSH metabolism were visible in wheat shoots at the highest concentrations of both forms of Se. Obtained results can serve as a basis for further research on Se toxicity and detoxification mechanisms in wheat. New insights into the Se impact on GSH metabolism could contribute to the further development of biofortification strategies.

Crosstalk of platelets with macrophages and fibroblasts aggravates inflammation, aortic wall stiffening, and osteopontin release in abdominal aortic aneurysm.

AIMS: Abdominal aortic aneurysm (AAA) is a highly lethal disease with progressive dilatation of the abdominal aorta accompanied by degradation and remodelling of the vessel wall due to chronic inflammation. Platelets play an important role in cardiovascular diseases, but their role in AAA is poorly understood. METHODS AND RESULTS: The present study revealed that platelets play a crucial role in promoting AAA through modulation of inflammation and degradation of the extracellular matrix (ECM). They are responsible for the up-regulation of SPP1 (osteopontin, OPN) gene expression in macrophages and aortic tissue, which triggers inflammation and remodelling and also platelet adhesion and migration into the abdominal aortic wall and the intraluminal thrombus (ILT). Further, enhanced platelet activation and pro-coagulant activity result in elevated gene expression of various cytokines, Mmp9 and Col1a1 in macrophages and Il-6 and Mmp9 in fibroblasts. Enhanced platelet activation and pro-coagulant activity were also detected in AAA patients. Further, we detected platelets and OPN in the vessel wall and in the ILT of patients who underwent open repair of AAA. Platelet depletion in experimental murine AAA reduced inflammation and ECM remodelling, with reduced elastin fragmentation and aortic diameter expansion. Of note, OPN co-localized with platelets, suggesting a potential role of OPN for the recruitment of platelets into the ILT and the aortic wall. CONCLUSION: In conclusion, our data strongly support the potential relevance of anti-platelet therapy to reduce AAA progression and rupture in AAA patients.

Myosin-X recruits lamellipodin to filopodia tips.

Myosin-X (MYO10), a molecular motor localizing to filopodia, is thought to transport various cargo to filopodia tips, modulating filopodia function. However, only a few MYO10 cargoes have been described. Here, using GFP-Trap and BioID approaches combined with mass spectrometry, we identified lamellipodin (RAPH1) as a novel MYO10 cargo. We report that the FERM domain of MYO10 is required for RAPH1 localization and accumulation at filopodia tips. Previous studies have mapped the RAPH1 interaction domain for adhesome components to its talin-binding and Ras-association domains. Surprisingly, we find that the RAPH1 MYO10-binding site is not within these domains. Instead, it comprises a conserved helix located just after the RAPH1 pleckstrin homology domain with previously unknown functions. Functionally, RAPH1 supports MYO10 filopodia formation and stability but is not required to activate integrins at filopodia tips. Taken together, our data indicate a feed-forward mechanism whereby MYO10 filopodia are positively regulated by MYO10-mediated transport of RAPH1 to the filopodium tip.

Role of extracellular matrix architecture and signaling in melanoma therapeutic resistance.

The extracellular matrix (ECM) is critical for maintaining tissue homeostasis therefore its production, assembly and mechanical stiffness are highly regulated in normal tissues. However, in solid tumors, increased stiffness resulting from abnormal ECM structural changes is associated with disease progression, an increased risk of metastasis and poor survival. As a dynamic and key component of the tumor microenvironment, the ECM is becoming increasingly recognized as an important feature of tumors, as it has been shown to promote several hallmarks of cancer via biochemical and biomechanical signaling. In this regard, melanoma cells are highly sensitive to ECM composition, stiffness and fiber alignment because they interact directly with the ECM in the tumor microenvironment via cell surface receptors, secreted factors or enzymes. Importantly, seeing as the ECM is predominantly deposited and remodeled by myofibroblastic stromal fibroblasts, it is a key avenue facilitating their paracrine interactions with melanoma cells. This review gives an overview of melanoma and further describes the critical roles that ECM properties such as ECM remodeling, ECM-related proteins and stiffness play in cutaneous melanoma progression, tumor cell plasticity and therapeutic resistance. Finally, given the emerging importance of ECM dynamics in melanoma, future perspectives on therapeutic strategies to normalize the ECM in tumors are discussed.

Targeting Discoidin Domain Receptors DDR1 and DDR2 overcomes matrix-mediated tumor cell adaptation and tolerance to BRAF-targeted therapy in melanoma.

Resistance to BRAF/MEK inhibitor therapy in BRAFV600 -mutated advanced melanoma remains a major obstacle that limits patient benefit. Microenvironment components including the extracellular matrix (ECM) can support tumor cell adaptation and tolerance to targeted therapy; however, the underlying mechanisms remain poorly understood. Here, we investigated the process of matrix-mediated drug resistance (MMDR) in response to BRAFV600 pathway inhibition in melanoma. We demonstrate that physical and structural cues from fibroblast-derived ECM abrogate anti-proliferative responses to BRAF/MEK inhibition. MMDR is mediated by drug-induced linear clustering of phosphorylated DDR1 and DDR2, two tyrosine kinase collagen receptors. Depletion and pharmacological targeting of DDR1 and DDR2 overcome ECM-mediated resistance to BRAF-targeted therapy. In xenografts, targeting DDR with imatinib enhances BRAF inhibitor efficacy, counteracts drug-induced collagen remodeling, and delays tumor relapse. Mechanistically, DDR-dependent MMDR fosters a targetable pro-survival NIK/IKKα/NF-κB2 pathway. These findings reveal a novel role for a collagen-rich matrix and DDR in tumor cell adaptation and resistance. They also provide important insights into environment-mediated drug resistance and a preclinical rationale for targeting DDR signaling in combination with targeted therapy in melanoma.

Chelidonium majus crude extract induces activation of peripheral blood mononuclear cells and enhances their cytotoxic effect toward HeLa cells.

The aim of the study was to examine the immunomodulatory effect of crude Chelidonium majus L ethanolic extract on ex vivo harvested peripheral blood mononuclear cells (PBMNCs). PBMNCs were isolated by density gradient centrifugation. The PBMNC cytotoxicity assay was performed with HeLa tumor cells as target cells. MTT assay was used to estimate the proliferation effect of extract and cytotoxic efficiency of treated PBMNCs. Flow cytometric analysis was used for immunophenotyping. Treatment induced moderate proliferative response, perturbation in PBMNC ratios, and the emergence of some unconventional subpopulations. The percentage ratio of double positive CD4+ and CD8+ T lymphocytes and monocytes, ratio of T and B lymphocytes expressing CD14, and percentage of NK cells expressing CD57 increased after treatment, indicating activation of PBMNC subpopulations. Cytotoxic activity against HeLa cells was enhanced. Activation of PBMNCs and enhancement of their cytotoxic effect toward HeLa cells indicate the immunostimulatory effect of Ch. majus ethanolic extract.

Myosin-X and talin modulate integrin activity at filopodia tips.

Filopodia assemble unique integrin-adhesion complexes to sense the extracellular matrix. However, the mechanisms of integrin regulation in filopodia are poorly defined. Here, we report that active integrins accumulate at the tip of myosin-X (MYO10)-positive filopodia, while inactive integrins are uniformly distributed. We identify talin and MYO10 as the principal integrin activators in filopodia. In addition, deletion of MYO10's FERM domain, or mutation of its ß1-integrin-binding residues, reveals MYO10 as facilitating integrin activation, but not transport, in filopodia. However, MYO10's isolated FERM domain alone cannot activate integrins, potentially because of binding to both integrin tails. Finally, because a chimera construct generated by swapping MYO10-FERM by talin-FERM enables integrin activation in filopodia, our data indicate that an integrin-binding FERM domain coupled to a myosin motor is a core requirement for integrin activation in filopodia. Therefore, we propose a two-step integrin activation model in filopodia: receptor tethering by MYO10 followed by talin-mediated integrin activation.

TBX3 Promotes Melanoma Migration by Transcriptional Activation of ID1, which Prevents Activation of E-Cadherin by MITF.

In melanoma, a phenotype switch from proliferation to invasion underpins metastasis, the major cause of melanoma-associated death. The transition from radial to vertical growth phase (invasive) melanoma is characterized by downregulation of both E-cadherin (CDH1) and MITF and upregulation of the key cancer-associated gene TBX3 and the phosphatidylinositol 3 kinase signaling pathway. Yet, whether and how these diverse events are linked remains poorly understood. Here, we show that TBX3 directly promotes expression of ID1, a dominant-negative regulator of basic helix-loop-helix transcription factors, and that ID1 decreases MITF binding and upregulation of CDH1. Significantly, we show that TBX3 activation of ID1 is necessary for TBX3 to enhance melanoma cell migration, and the mechanistic links between TBX3, ID1, MITF, and invasion revealed here are reflected in their expression in human melanomas. Our results reveal that melanoma migration is promoted through a TBX3-ID1-MITF-E-cadherin axis and that ID1-mediated repression of MITF activity may reinforce maintenance of an MITFLow phenotype associated with disease progression and therapy resistance.

Effect of ß-cyclodextrin encapsulation on cytotoxic activity of acetylshikonin against HCT-116 and MDA-MB-231 cancer cell lines.

Acetylshikonin (AcSh), as a red colored pigment found in roots of the plants from family Boraginaceae, showed excellent cytotoxic activity. Due to its hydrophobic nature, and thus poor bioavailability, the aim of this study was to prepare acetylshikonin/ß-cyclodextrin (AcSh/ß-CD) inclusion complex by using coprecipitation method, characterize obtained system by using UV/VIS, IR and 1H NMR spectroscopy, and determine cytotoxic activity. Phase solubility test indicated formation of AL-type binary system (substrate/ligand ratio was 1:1 M/M), with stability constant Ks of 306.01 M-1. Formation of noncovalent bonds between inner layer of the hole of ß-CD and AcSh was observed using spectroscopic methods. Notable changes in chemical shifts of two protons (-0.020 ppm) from naphthoquinone moiety (C6-H and C7-H), as well as protons from hydroxyl groups (-0.013 and -0.009, respectively) attached to C5 and C8 carbons from naphthoquinone part indicate that the molecule of AcSh enters the ß-CD cavity from the aromatic side. Cytotoxic activity against HCT-116 and MDA-MB-231 cell lines was measured by MTT test and clonogenic assay. Mechanisms of action of free AcSh and inclusion complex were assessed by flow cytometry. In comparison to free AcSh, AcSh/ß-CD showed stronger short-term effect on HCT-116 cells and superior long-term effect on both cell lines. Inclusion complex induced more pronounced cell cycle arrest and autophagy inhibition, and induced increase in accumulation of intracellular ROS more effectively than free AcSh. In conclusion, AcSh/ß-CD binary system showed better performances regarding cytotoxic activity against tested tumor cell lines.

Exploring Bacterial Carboxylate Reductases for the Reduction of Bifunctional Carboxylic Acids.

Carboxylic acid reductases (CARs) selectively reduce carboxylic acids to aldehydes using ATP and NADPH as cofactors under mild conditions. Although CARs attracts significant interest, only a few enzymes have been characterized to date, whereas the vast majority of CARs have yet to be examined. Herein the authors report that 12 bacterial CARs reduces a broad range of bifunctional carboxylic acids containing oxo-, hydroxy-, amino-, or second carboxyl groups with several enzymes showing activity toward 4-hydroxybutanoic (4-HB) and adipic acids. These CARs exhibits significant reductase activity against substrates whose second functional group is separated from the carboxylate by at least three carbons with both carboxylate groups being reduced in dicarboxylic acids. Purified CARs supplemented with cofactor regenerating systems (for ATP and NADPH), an inorganic pyrophosphatase, and an aldo-keto reductase catalyzes a high conversion (50-76%) of 4-HB to 1,4-butanediol (1,4-BDO) and adipic acid to 1,6-hexanediol (1,6-HDO). Likewise, Escherichia coli strains expressing eight different CARs efficiently reduces 4-HB to 1,4-BDO with 50-95% conversion, whereas adipic acid is reduced to a mixture of 6-hydroxyhexanoic acid (6-HHA) and 1,6-HDO. Thus, our results illustrate the broad biochemical diversity of bacterial CARs and their compatibility with other enzymes for applications in biocatalysis.

Prospective randomized controlled trial comparing standard analgesia with combined intra-operative cystic plate and port-site local anesthesia for post-operative pain management in elective laparoscopic cholecystectomy.

BACKGROUND: Various mechanisms, including somatic and visceral nociceptive stimulation, have been suggested as a cause for pain after laparoscopic cholecystectomy (LC). We therefore conducted a prospective randomized controlled trial (PRCT) to evaluate whether somatovisceral pain blockade reduces pain after LC. HYPOTHESIS: Analgesic efficacy of multimodal analgesia is superior to standard analgesia for patients undergoing elective LC for symptomatic cholelithiasis. Specifically, topical cystic plate and port-site injection with 0.25 % bupivacaine significantly reduces pain after LC. DESIGN: This study was designed as single-blinded PRCT. SETTING: This study was conducted in an academic medical center. PATIENTS AND METHODS: Between February and May 2010 we randomly assigned 63 patients with symptomatic cholelithiasis in a 1:1 ratio to non-opioid/opioid analgesic combinations (Control Group, n = 32) and non-opioid/opioid analgesic combinations plus topical 0.25 % bupivacaine onto the cystic plate and local 0.25 % bupivacaine port-site injection, post-LC (Study Group, n = 31). Primary endpoint was patient-reported pain 1, 4, 6, 12, 24 h and 1 week post-LC using the Visual Analog Scale (VAS 0-10). RESULTS: Study groups were comparable clinicopathologically. There were no adverse events. A statistically significant reduction in mean pain score was apparent in Study Group patients in comparison with Control Group (mean VAS 4.83 ± 2.33 vs. 6.80 ± 1.87; p < 0.001) at all early (1-6 h) post-operative time points following LC. CONCLUSION: This PRCT shows significantly improved pain control with somatovisceral pain blockade over non-opioid/opioid analgesic combinations following LC for symptomatic cholelithiasis. For centers not utilizing adjunctive local anesthetic for LC, this topical use of bupivacaine may improve patient comfort during recovery. This trial was registered on www.ClinicalTrials.gov NCT# 01972620.

Biochemical and structural studies of conserved Maf proteins revealed nucleotide pyrophosphatases with a preference for modified nucleotides.

Maf (for multicopy associated filamentation) proteins represent a large family of conserved proteins implicated in cell division arrest but whose biochemical activity remains unknown. Here, we show that the prokaryotic and eukaryotic Maf proteins exhibit nucleotide pyrophosphatase activity against 5-methyl-UTP, pseudo-UTP, 5-methyl-CTP, and 7-methyl-GTP, which represent the most abundant modified bases in all organisms, as well as against canonical nucleotides dTTP, UTP, and CTP. Overexpression of the Maf protein YhdE in E. coli cells increased intracellular levels of dTMP and UMP, confirming that dTTP and UTP are the in vivo substrates of this protein. Crystal structures and site-directed mutagenesis of Maf proteins revealed the determinants of their activity and substrate specificity. Thus, pyrophosphatase activity of Maf proteins toward canonical and modified nucleotides might provide the molecular mechanism for a dual role of these proteins in cell division arrest and house cleaning.

[Role of surgical resection in treatment of pancreatic adenocarcinoma].

INTRODUCTION: Pancreatic adenocarcinoma is the fifth leading cause of death from malignant diseases. The total five-year rate is bellow 5%, but in patients who underwent pancreatic resection, the five-year rate may be up to 20%. Surgical resection is still the only therapeutic option that offers the possibility of cure. In recent decades, the perioperative mortality rate has been significantly reduced in the institutions performing a number of these operations per year and has become less than 5%. Postoperative morbidity remains high. MATERIAL AND METHODS: The results of surgical resection in the treatment of pancreatic adenocarcinoma have been analyzed. A retrospective study included the patients operated at the Department for Abdominal, Endocrine and Transplantation surgery, Clinical Center of Vojvodina. RESULTS: In the period from February 1st 1998 to February 1st 2007 a total of 67 patients with pancreatic adenocarcinoma underwent resection. The average age of patients was 58.81 +/- 1.42 years. There were 44 (65.7%) male and 23 (34.3%) female patients. The most common locations of cancer were the head, then the body and the tail of the pancreas and they were found in 57 (85.1%) cases, 7 (10,4%) cases and 3 (4,47%) cases, respectively. The postoperative mortality appeared in 3 (4.47%) cases and postoperative morbidity in 21 (31.3%) cases. The average survival was 22.89 +/- 3.87 months, the median being 9.0 +/- 2.18 months. The five-year survival rate was 13.5%. CONCLUSION: For patients with pancreatic cancer, surgical resection still remains the only chance of cure. These procedures are performed with acceptable postoperative mortality and morbidity rate. The percentage of cured patients is still unsatisfactorily low.