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Endothelial nitric oxide synthase (eNOS)-related vessel relaxation is a highly coordinated process that regulates blood flow and pressure and is dependent on caveolae. Here, we investigated the role of caveolar plasma membrane stabilization by the dynamin-related ATPase EHD2 on eNOS-nitric oxide (NO)-dependent vessel relaxation. Loss of EHD2 in small arteries led to increased numbers of caveolae that were detached from the plasma membrane. Concomitantly, impaired relaxation of mesenteric arteries and reduced running wheel activity were observed in EHD2 knockout mice. EHD2 deletion or knockdown led to decreased production of nitric oxide (NO) although eNOS expression levels were not changed. Super-resolution imaging revealed that eNOS was redistributed from the plasma membrane to internalized detached caveolae in EHD2-lacking tissue or cells. Following an ATP stimulus, reduced cytosolic Ca2+ peaks were recorded in human umbilical vein endothelial cells (HUVECs) lacking EHD2. Our data suggest that EHD2-controlled caveolar dynamics orchestrates the activity and regulation of eNOS/NO and Ca2+ channel localization at the plasma membrane.

A fundamental computational problem in metabolic engineering is to find pathways between compounds. Pathfinding methods using atom tracking have been widely used to find biochemically relevant pathways. However, these methods require the user to define the atoms to be tracked. This may lead to failing to predict the pathways that do not conserve the user-defined atoms. In this work, we propose a pathfinding method called AGPathFinder to find biochemically relevant metabolic pathways between two given compounds. In AGPathFinder, we find alternative pathways by tracking the movement of atomic groups through metabolic networks and use combined information of reaction thermodynamics and compound similarity to guide the search towards more feasible pathways and better performance. The experimental results show that atomic group tracking enables our method to find pathways without the need of defining the atoms to be tracked, avoid hub metabolites, and obtain biochemically meaningful pathways. Our results also demonstrate that atomic group tracking, when incorporated with combined information of reaction thermodynamics and compound similarity, improves the quality of the found pathways. In most cases, the average compound inclusion accuracy and reaction inclusion accuracy for the top resulting pathways of our method are around 0.90 and 0.70, respectively, which are better than those of the existing methods. Additionally, AGPathFinder provides the information of thermodynamic feasibility and compound similarity for the resulting pathways.

The halotolerant chlorophyte Dunaliella salina can accumulate up to 10% of its dry weight as β-carotene in chloroplasts when subjected to adverse conditions, including nutrient deprivation. However, the mechanisms of carotenoid biosynthesis are poorly understood. Here, the physiological and molecular responses to the deprivation of nitrogen (-N), sulfur (-S), phosphorus (-P) and different combinations of those nutrients (-N-P, -N-S, -P-S and -N-P-S) were compared to gain insights into the underlying regulatory mechanisms of carotenoid biosynthesis. The results showed that both the growth and photosynthetic rates of cells were decreased during nutrient deprivation, accompanied by lipid globule accumulation and reduced chlorophyll levels. The SOD and CAT activities of the cells were altered during nutrient deprivation, but their responses were different. The total carotenoid contents of cells subjected to multiple nutrient deprivation were higher than those of cells subjected to single nutrient deprivation and non-stressed cells. The β-carotene contents of cells subjected to -N-P, -N-S and -N-P-S were higher than those of cells subjected to single nutrient deprivation. Cells subjected to sulfur deprivation accumulated more lutein than cells subjected to nitrogen and phosphorous deprivation. In contrast, no cumulative effects of nutrient deprivation on the transcription of genes in the carotenogenic pathway were observed because MEP and carotenogenic pathway genes were up-regulated during single nutrient deprivation but were downregulated during multiple nutrient deprivation. Therefore, we proposed that the carotenoid biosynthesis pathway of D. salina is regulated at both the transcriptional and posttranscriptional levels and that a complex crosstalk occurs at the physiological and molecular levels in response to the deprivation of different nutrients.

Metabolic pathway alignment has been widely used to find one-to-one and/or one-to-many reaction mappings to identify the alternative pathways that have similar functions through different sets of reactions, which has important applications in reconstructing phylogeny and understanding metabolic functions. The existing alignment methods exhaustively search reaction sets, which may become infeasible for large pathways. To address this problem, we present an effective alignment method for accurately extracting reaction mappings between two metabolic pathways. We show that connected relation between reactions can be formalized as binary relation of reactions in metabolic pathways, and the multiplications of zero-one matrices for binary relations of reactions can be accomplished in finite steps. By utilizing the multiplications of zero-one matrices for binary relation of reactions, we efficiently obtain reaction sets in a small number of steps without exhaustive search, and accurately uncover biologically relevant reaction mappings. Furthermore, we introduce a measure of topological similarity of nodes (reactions) by comparing the structural similarity of the k-neighborhood subgraphs of the nodes in aligning metabolic pathways. We employ this similarity metric to improve the accuracy of the alignments. The experimental results on the KEGG database show that when compared with other state-of-the-art methods, in most cases, our method obtains better performance in the node correctness and edge correctness, and the number of the edges of the largest common connected subgraph for one-to-one reaction mappings, and the number of correct one-to-many reaction mappings. Our method is scalable in finding more reaction mappings with better biological relevance in large metabolic pathways.

A better understanding of metabolic fluxes is important for manipulating microbial metabolism toward desired end products, or away from undesirable by-products. A mutant strain, Gluconacetobacter xylinus AX2-16, was obtained by combined chemical mutation of the parent strain (G. xylinus CGMCC 2955) using DEC (diethyl sulfate) and LiCl. The highest bacterial cellulose production for this mutant was obtained at about 11.75 g/L, which was an increase of 62% compared with that by the parent strain. In contrast, gluconic acid (the main byproduct) concentration was only 5.71 g/L for mutant strain, which was 55.7% lower than that of parent strain. Metabolic flux analysis indicated that 40.1% of the carbon source was transformed to bacterial cellulose in mutant strain, compared with 24.2% for parent strain. Only 32.7% and 4.0% of the carbon source were converted into gluconic acid and acetic acid in mutant strain, compared with 58.5% and 9.5% of that in parent strain. In addition, a higher flux of tricarboxylic acid (TCA) cycle was obtained in mutant strain (57.0%) compared with parent strain (17.0%). It was also indicated from the flux analysis that more ATP was produced in mutant strain from pentose phosphate pathway (PPP) and TCA cycle. The enzymatic activity of succinate dehydrogenase (SDH), which is one of the key enzymes in TCA cycle, was 1.65-fold higher in mutant strain than that in parent strain at the end of culture. It was further validated by the measurement of ATPase that 3.53-6.41 fold higher enzymatic activity was obtained from mutant strain compared with parent strain.

Background: The aim of this study was to investigate the interaction effects of pri-let-7a-1 rs10739971 with pepsinogen C (PGC) and excision repair cross complementing group 6 (ERCC6) gene polymorphisms and its association with the risks of gastric cancer and atrophic gastritis. We hoped to identify miRNA polymorphism or a combination of several polymorphisms that could serve as biomarkers for predicting the risk of gastric cancer and its precancerous diseases. Methods: Sequenom MassARRAY platform method was used to detect polymorphisms of pri-let-7a-1 rs10739971 G→A, PGC rs4711690 C→G, PGC rs6458238 G→A, PGC rs9471643 G→C, and ERCC6 rs1917799 in 471 gastric cancer patients, 645 atrophic gastritis patients and 717 controls. Results: An interaction effect of pri-let-7a-1 rs10739971 polymorphism with ERCC6 rs1917799 polymorphism was observed for the risk of gastric cancer (Pinteraction = 0.026); and interaction effects of pri-let-7a-1 rs10739971 polymorphism with PGC rs6458238 polymorphism (Pinteraction = 0.012) and PGC rs9471643 polymorphism (Pinteraction = 0.039) were observed for the risk of atrophic gastritis. Conclusion: The combination of pri-let-7a-1 rs10739971 polymorphism and ERCC6 and PGC polymorphisms could provide a greater prediction potential than a single polymorphism on its own. Large-scale studies and molecular mechanism research are needed to confirm our findings. [ABSTRACT FROM AUTHOR]

Glutathione S-transferase P1 (GSTP1) is a critical enzyme in the phase II detoxification pathway. One of the common functional polymorphisms of GSTP1 is A→G at nucleotide 313, which results in an amino acid substitution (Ile105Val) at the substrate binding site and reduced catalytic activity. We evaluated the interaction between GSTP1 Val allele and Helicobacter pylori infection, smoking and alcohol consumption, increasing the risk of gastric cancer among the Chinese population. Information on potential gastric cancer risk factors and blood specimens were collected from 618 incident gastric cancer cases and 1,830 non-cancer controls between March 2002 and December 2011 in Liaoning Province, China. GSTP1 Ile105Val was genotyped by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and polymerase chain reaction-restriction fragment length polymorphism. Serum levels of anti-H. pylori IgG were measured by ELISA. Odds ratio (OR) and 95% confidence interval (CI) were calculated using multivariate logistic regression, adjusted by sex and age. The risk of gastric cancer was significantly elevated in patients with the GSTP1 Val/Val genotype (adjusted OR = 3.324; 95% CI = 1.790-6.172). An elevated risk of gastric cancer was observed in patients with H. pylori infection, smoking, or alcohol consumption, and together with the GSTP1 Ile/Val +Val/Val genotype (OR = 3.696; 95% CI = 2.475-5.521; OR = 1.638; 95% CI = 1.044-2.571; OR = 1.641; 95% CI = 0.983-2.739, respectively) (p<0.05). The GSTP1 Val allele shows an interaction with smoking, alcohol consumption, and especially H. pylori infection for increasing the risk of gastric cancer. These findings could demonstrate new pathophysiological pathways for the development of gastric cancer. [ABSTRACT FROM AUTHOR]

Objective: The aim of the current study was to investigate which is the most suitable classification for colorectal cancer, log odds of positive lymph nodes (LODDS) classification or the classifications based on the number of positive lymph nodes (pN) and positive lymph node ratio(LNR) in a Chinese single institutional population. Design: Clinicopathologic and prognostic data of 1297 patients with colorectal cancer were retrospectively studied. The log-rank statistics, Cox's proportional hazards model, the Nagelkerke R2 index and a Harrell's C statistic were used. Results: Univariate and three-step multivariate analyses identified that LNR was a significant prognostic factor and LNR classification was superior to both the pN and LODDS classifications. Moreover, the results of the Nagelkerke R2 index (0.130) and a Harrell's C statistic (0.707) of LNR showed that LNR and LODDS classifications were similar and LNR was a little better than the other two classifications. Furthermore, for patients in each LNR classification, prognosis was homologous between those in different pN or LODDS classifications. However, for patients in pN1a, pN1b, LODDS2 and LODDS3 classifications, significant differences in survival were observed among patients in different LNR classifications. Conclusions: For patients with colorectal cancer, the LNR classification is more suitable than pN and LODDS classifications for prognostic assessment in a Chinese single institutional population. [ABSTRACT FROM AUTHOR]

Methods for constructing trees using DNA sequences, known as molecular phylogenetics, have been applied to analyses of phylogenetic origin, evolutionary relatedness and taxonomic classification. Combining data sequenced in this study and downloaded from GenBank, we sampled 112 (chloroplast data) / 122 (ITS data) specimens belonging to 49 (chloroplast data) / 46 (ITS data) poplar species or hybrids from six (chloroplast data) / five sections (ITS data). Maximum parsimony and Bayesian inference were used to analyze phylogenetic relationships within the genus Populus based on eight chloroplast combinations and ITS regions. The results suggested that Bayesian inference might be more suitable for the phylogenetic reconstruction of Populus. All Populus species could be divided into two clades: clade 1, including subclades 1 and 2, and clade 2, including subclades 3 and 4. Species within clade 1, involving five sections except for Leuce, clustered coinciding with their two specific main geographical distribution areas: China (subclade 1) and North America (subclade 2). Clustering in subclade 3, section Leuce was confirmed to be of monophyletic origin and independent evolution. Its two subsections, namely Albidae and Trepidae, could be separated by chloroplast data but had frequent gene flow based on ITS data. Phylogeny analysis based on chloroplast data demonstrated once more that section Aigeiros was paraphyletic and further showed that the P. deltoides lineage is restricted in subclade 2 and that P. nigra lineage, located in subclade 3, originated from a hybrid of which an Albidae ancestor species was the material parent. Similarly, section Tacamahaca was found to be paraphyletic and had two lineages: a clade 1 lineage, such as P. cathayana, and a clade 2 lineage, such as P. simonii. Section Leucoides was paraphyletic and closely linked to section Tacamahaca. Their section boundaries were not conclusively delimitated by sequencing information.

ObjectiveTo investigate the biomechanical effects of the lumbar posterior complex on the adjacent segments after posterior lumbar interbody fusion (PLIF) surgeries.MethodsA finite element model of the L1-S1 segment was modified to simulate PLIF with total laminectomy (PLIF-LAM) and PLIF with hemilaminectomy (PLIF-HEMI) procedures. The models were subjected to a 400N follower load with a 7.5-N.m moment of flexion, extension, torsion, and lateral bending. The range of motion (ROM), intradiscal pressure (IDP), and ligament force were compared.ResultsIn Flexion, the ROM, IDP and ligament force of posterior longitudinal ligament, intertransverse ligament, and capsular ligament remarkably increased at the proximal adjacent segment in the PLIF-LAM model, and slightly increased in the PLIF-HEMI model. There was almost no difference for the ROM, IDP and ligament force at L5-S1 level between the two PLIF models although the ligament forces of ligamenta flava remarkably increased compared with the intact lumbar spine (INT) model. For the other loading conditions, these two models almost showed no difference in ROM, IDP and ligament force on the adjacent discs.ConclusionsPreserved posterior complex acts as the posterior tension band during PLIF surgery and results in less ROM, IDP and ligament forces on the proximal adjacent segment in flexion. Preserving the posterior complex during decompression can be effective on preventing adjacent segment degeneration (ASD) following PLIF surgeries.

The phosphoinositide 3-kinase (PI3K)/protein kinase B (PKB, AKT)/mammalian target of rapamycin (mTOR) signaling pathway plays a critical role in angiogenesis and cell growth, proliferation, metabolism, migration, differentiation, and apoptosis. Genetic diversity in key factors of this pathway may influence protein function and signal transduction, contributing to disease initiation and progression. Studies suggest that MTOR rs1064261 and AKT rs1130233 polymorphisms are associated with risk and/or prognosis of multiple cancer types. However, this relationship with gastric cancer (GC) remains unclear. The aim of this study was to investigate the role of MTOR and AKT polymorphisms in the risk and prognosis of GC.The Sequenom MassARRAY platform was used to genotype 1842 individuals for MTOR rs1064261 T→C and AKT rs1130233 G→A polymorphisms. ELISA was used to detect Helicobacter pylori antibodies in serum. Immunohistochemical analysis was used to detect total and phosphorylated MTOR and AKT proteins.The MTOR rs1064261 (TC+CC) genotype and the AKT rs1130233 (GA+AA) genotype were associated with increased risk of GC in men (P = 0.049, P = 0.030). In H. pylori-negative individuals, the AKT rs1130233 GA and (GA+AA) genotypes were related to increased risk of atrophic gastritis (AG; P = 0.012, P = 0.024). Notably, the AKT rs1130233 (GA+AA) genotype demonstrated significant interactions with H. pylori in disease progression from healthy controls (CON) to AG (P = 0.013) and from AG to GC (P = 0.049). Additionally, for individuals with the AKT rs1130233 variant, those in the H. pylori-positive group had higher levels of phosphorylated AKT (p-AKT) expression. The AKT rs1130233 genotype was found to be associated with clinicopathological parameters including lymph node metastasis and alcohol drinking (P

OBJECTIVE: CD4(+) latency-associated peptide (LAP)(+) regulatory T cells (Tregs) are a newly discovered T cell subset in humans and the role of these cells in patients with acute coronary syndrome (ACS) has not been explored. We designed to investigate whether circulating frequency and function of CD4(+)LAP(+) Tregs are defective in ACS. METHODS: One hundred eleven ACS patients (acute myocardial infarction and unstable angina) and 117 control patients were enrolled in the study. The control patients consisted of chronic stable angina (CSA) and chest pain syndrome (CPS). The frequencies of circulating CD4(+)LAP(+) Tregs and the expression of the transmembrane protein glycoprotein-A repetitions predominant (GARP) on CD4(+) T cells were determined by flow cytometry. The function of CD4(+)LAP(+) Tregs was detected using thymidine uptake. Serum interleukin-10 (IL-10) and transforming growth factor-β protein (TGF-β) levels were detected using ELISA and expression of GARP mRNA in peripheral blood mononuclear cells (PBMCs) was measured by real time-polymerase chain reaction. RESULTS: We found ACS patients had a significantly lower frequency of circulating CD4(+)LAP(+) Tregs, and the function of these cells was reduced compared to controls. The expression of GARP in CD4(+) T cells and the serum levels of TGF-β in ACS patients were lower than those of control patients. The serum levels of IL-10 were similar between the two cohorts. CONCLUSIONS: A novel regulatory T cell subset, defined as CD4(+)LAP(+) T cells is defective in ACS patients.

BACKGROUND: The aim of this study was to investigate the interaction effects of pri-let-7a-1 rs10739971 with pepsinogen C (PGC) and excision repair cross complementing group 6 (ERCC6) gene polymorphisms and its association with the risks of gastric cancer and atrophic gastritis. We hoped to identify miRNA polymorphism or a combination of several polymorphisms that could serve as biomarkers for predicting the risk of gastric cancer and its precancerous diseases. METHODS: Sequenom MassARRAY platform method was used to detect polymorphisms of pri-let-7a-1 rs10739971 G → A, PGC rs4711690 C → G, PGC rs6458238 G → A, PGC rs9471643 G → C, and ERCC6 rs1917799 in 471 gastric cancer patients, 645 atrophic gastritis patients and 717 controls. RESULTS: An interaction effect of pri-let-7a-1 rs10739971 polymorphism with ERCC6 rs1917799 polymorphism was observed for the risk of gastric cancer (P interaction = 0.026); and interaction effects of pri-let-7a-1 rs10739971 polymorphism with PGC rs6458238 polymorphism (P interaction = 0.012) and PGC rs9471643 polymorphism (P interaction = 0.039) were observed for the risk of atrophic gastritis. CONCLUSION: The combination of pri-let-7a-1 rs10739971 polymorphism and ERCC6 and PGC polymorphisms could provide a greater prediction potential than a single polymorphism on its own. Large-scale studies and molecular mechanism research are needed to confirm our findings.