ADAMTS-7 Modulates Atherosclerotic Plaque Formation by Degradation of TIMP-1.

BACKGROUND: The ADAMTS7 locus was genome-wide significantly associated with coronary artery disease. Lack of the ECM (extracellular matrix) protease ADAMTS-7 (A disintegrin and metalloproteinase-7) was shown to reduce atherosclerotic plaque formation. Here, we sought to identify molecular mechanisms and downstream targets of ADAMTS-7 mediating the risk of atherosclerosis. METHODS: Targets of ADAMTS-7 were identified by high-resolution mass spectrometry of atherosclerotic plaques from Apoe-/- and Apoe-/-Adamts7-/- mice. ECM proteins were identified using solubility profiling. Putative targets were validated using immunofluorescence, in vitro degradation assays, coimmunoprecipitation, and Förster resonance energy transfer-based protein-protein interaction assays. ADAMTS7 expression was measured in fibrous caps of human carotid artery plaques. RESULTS: In humans, ADAMTS7 expression was higher in caps of unstable as compared to stable carotid plaques. Compared to Apoe-/- mice, atherosclerotic aortas of Apoe-/- mice lacking Adamts-7 (Apoe-/-Adamts7-/-) contained higher protein levels of Timp-1 (tissue inhibitor of metalloprotease-1). In coimmunoprecipitation experiments, the catalytic domain of ADAMTS-7 bound to TIMP-1, which was degraded in the presence of ADAMTS-7 in vitro. ADAMTS-7 reduced the inhibitory capacity of TIMP-1 at its canonical target MMP-9 (matrix metalloprotease-9). As a downstream mechanism, we investigated collagen content in plaques of Apoe-/- and Apoe-/-Adamts7-/- mice after a Western diet. Picrosirius red staining of the aortic root revealed less collagen as a readout of higher MMP-9 activity in Apoe-/- as compared to Apoe-/- Adamts7-/- mice. To facilitate high-throughput screening for ADAMTS-7 inhibitors with the aim of decreasing TIMP-1 degradation, we designed a Förster resonance energy transfer-based assay targeting the ADAMTS-7 catalytic site. CONCLUSIONS: ADAMTS-7, which is induced in unstable atherosclerotic plaques, decreases TIMP-1 stability reducing its inhibitory effect on MMP-9, which is known to promote collagen degradation and is likewise associated with coronary artery disease. Disrupting the interaction of ADAMTS-7 and TIMP-1 might be a strategy to increase collagen content and plaque stability for the reduction of atherosclerosis-related events.

Fragmentomic analysis of circulating tumor DNA-targeted cancer panels.

BACKGROUND: The isolation of cell-free DNA (cfDNA) from the bloodstream can be used to detect and analyze somatic alterations in circulating tumor DNA (ctDNA), and multiple cfDNA-targeted sequencing panels are now commercially available for Food and Drug Administration (FDA)-approved biomarker indications to guide treatment. More recently, cfDNA fragmentation patterns have emerged as a tool to infer epigenomic and transcriptomic information. However, most of these analyses used whole-genome sequencing, which is insufficient to identify FDA-approved biomarker indications in a cost-effective manner. PATIENTS AND METHODS: We used machine learning models of fragmentation patterns at the first coding exon in standard targeted cancer gene cfDNA sequencing panels to distinguish between cancer and non-cancer patients, as well as the specific tumor type and subtype. We assessed this approach in two independent cohorts: a published cohort from GRAIL (breast, lung, and prostate cancers, non-cancer, n = 198) and an institutional cohort from the University of Wisconsin (UW; breast, lung, prostate, bladder cancers, n = 320). Each cohort was split 70%/30% into training and validation sets. RESULTS: In the UW cohort, training cross-validated accuracy was 82.1%, and accuracy in the independent validation cohort was 86.6% despite a median ctDNA fraction of only 0.06. In the GRAIL cohort, to assess how this approach performs in very low ctDNA fractions, training and independent validation were split based on ctDNA fraction. Training cross-validated accuracy was 80.6%, and accuracy in the independent validation cohort was 76.3%. In the validation cohort where the ctDNA fractions were all <0.05 and as low as 0.0003, the cancer versus non-cancer area under the curve was 0.99. CONCLUSIONS: To our knowledge, this is the first study to demonstrate that sequencing from targeted cfDNA panels can be utilized to analyze fragmentation patterns to classify cancer types, dramatically expanding the potential capabilities of existing clinically used panels at minimal additional cost.

Plasticity in Compensatory Growth to Artificial Defoliation and Light Availability in Four Neotropical Understory and Forest Edge Herb Species.

Defoliation from falling branches is a major factor in the survival of understory herbs in tropical rainforests. Experimental studies of defoliation under three levels of light environment compared responses to partial and total defoliation in four species of tropical rainforest understory herbs. We predicted that elevated levels of light availability would help compensate for damage to through compensatory growth in both understory and forest edge species and that forest edge species would more effectively compensate under high light conditions than shade-tolerant species from the forest understory All species showed a high tolerance to defoliation under high and intermediate light conditions. Under low-light conditions survival differed dramatically with minimal mortality in forest-edge species compared to high mortality in completely defoliated understory species. Defoliation, and light × defoliation interactions, impacted multiple growth traits in understory species. In contrast, forest-edge species showed no effect of defoliation except on total biomass, and only one light × defoliation interaction was observed. Our results indicate that differences in biomass allocation, leaf ecophysiology, and other growth parameters between forest understory and edge species may be structuring post-damage response in understory and forest edge herbs.

Changes in gastrocnemii activation at mid-to-late stance markedly affects the intact and anterior cruciate ligament deficient knee biomechanics and stability in gait.

INTRODUCTION: We aimed to quantify the sensitivity in biomechanical response and stability of the intact and anterior cruciate ligament deficient (ACL-D) joints at mid-to-late stance periods of gait to the alterations in activation of gastrocnemii (Gas) muscles. METHODS: A validated kinematics-driven musculoskeletal finite-element model of the lower extremity is used to compute knee joint response and stability under reported kinetics-kinematics of healthy subjects. Activation in Gas is altered under prescribed gait data at the mid-to-late stance of gait and associated changes in remaining muscle forces/contact forces/areas/ACL force and joint stability are computed in both intact and ACL-D joints. RESULTS: In the intact joint, the anterior-tibial-translation (ATT) as well as ACL and joint contact forces follow variations in Gas forces. Both the stability and ATT of an ACL-D joint are restored to the near-intact levels when the activity in Gas is reduced. Knee joint instability, excessive ATT as well as larger peak articular contact stresses with a posterior shift in contact areas are estimated under greater Gas forces. CONCLUSIONS: ACL-D joint is unstable with ATT > 10 mm under larger activities in Gas. Gas is an ACL-antagonist while hamstrings and soleus are ACL-agonists. The near-intact joint stability and ATT of an ACL-D joint can be restored at a lower activation in Gas; or in other words, when activation in ACL-antagonist muscles drops compared with that in ACL-agonist muscles. Results could help analyze the gait of ACL-D copers and non-copers and provide better understanding towards improved preventive, diagnostic, and treatment approaches.

Knee flexion angle and muscle activations control the stability of an anterior cruciate ligament deficient joint in gait.

Anterior cruciate ligament (ACL) is a primary structure and a commonly injured ligament of the knee joint. Some patients with ACL deficiency (ACLD) experience joint instability and require a reconstructive surgery to return to daily routines, some can adapt by limiting their activities while others, called copers, can return to high-level activities with no instability. We investigated the effects of alterations in the knee flexion angle (KFA) and muscle force activations on the stability and biomechanics of ACLD joints at 25, 50, and 75% periods of gait stance. ACLD joint stability is controlled by variations in both KFA and knee muscle forces. For the latter, a parameter called activity index is defined as the ratio of forces in ACL antagonists (quadriceps and gastrocnemii) to those in ACL agonists (hamstrings). Under a greater KFA (2-6° beyond the mean of reported values in healthy subjects), an ACLD joint regains its pre-injury stability levels. The ACLD joint stability also markedly improves at smaller quadriceps and larger hamstrings forces (activity indices of 2.0-3.6 at 25%) at the first half of stance and smaller gastrocnemii and larger hamstrings forces (activity indices of 0.1-1.1 at 50% and 0.1-1.2 at 75%) at the second half of stance. Activity index and KFA are both crucial when assessing the dynamic stability of an ACLD joint. These results are helpful in our understanding of the biomechanics and stability of ACLD joints towards improved prevention and treatment strategies.

Functional traits of broad-leaved monocot herbs in the understory and forest edges of a Costa Rican rainforest.

Broad-leaved monocot herbs are widespread and dominant components of the shaded understories of wet neotropical forests. These understory habitats are characterized by light limitation and a constant threat of falling branches. Many shaded understory herb species have close relatives that occupy forest edges and gaps, where light availability is higher and defoliation threat is lower, creating an opportunity for comparative analysis of functional traits in order to better understand the evolutionary adaptations associated with this habitat transition. We documented ecological, morphological and ecophysiological traits of multiple herb species in six monocot families from each of these two habitats in the wet tropical rainforest at the La Selva Biological Station, Costa Rica. We found that a mixture of phylogenetic canalization and ecological selection for specific habitats helped explain patterns of functional traits. Understory herbs were significantly shorter and had smaller leaves than forest edge species. Although the mean number of leaves per plant and specific leaf area did not differ between the two groups, the larger leaf size of forest edge species gave them more than three times the mean plant leaf area. Measures of leaf water content and nitrogen content varied within both groups and mean values were not significantly different. Despite the high leaf nitrogen contents, the maximum photosynthetic rates of understory herbs were quite low. Measures of δ 13C as an analog of water use efficiency found significantly lower (more negative) values in understory herbs compared to forest edge species. Clonality was strongly developed in several species but did not show strong phylogenetic patterns. This study highlights many functional traits that differ between broad-leaved monocot species characteristic of understory and forest edge habitats, as well as traits that vary primarily by phylogenetic relatedness. Overall, plant functional traits do not provide a simple explanation for the relative differences in abundance for individual understory and forest edge species with some occurring in great abundance while others are relatively rare.

Reduce proliferation of human bone marrow cells from acute myeloblastic leukemia with minimally differentiation by blocking lncRNA PVT1.

PURPOSE: Acute myeloblastic leukemia with minimally differentiation (AML-M0) is a subtype of acute leukemia with poor prognosis. The recent studies have shown that long non-coding RNAs (lncRNAs) play an important role in different cellular processes, such as cell cycle control and proliferation. Plasmacytoma variant translocation 1 (PVT1) is one of those lncRNAs that is significantly upregulated in AML. LncRNAs could be downregulated or blocked by locked nucleic acids (LNA) which are oligonucleotide strands. METHODS: In this study, lncRNA PVT1 was blocked by antisense LNA GapmeRs in human bone marrow cancerous blast cells. Cells were transfected with PVT1 antisense LNA GapmeRs at 24, 48, and 72 h post-transfection. Quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) was accomplished to evaluate the PVT1 and c-Myc expression. Cell viability was evaluated by MTT assay, and apoptosis and necrosis were assessed by Annexin V/propidium iodide staining assay. RESULTS: The results of this study indicated that the downregulation of PVT1 in blast cells could induce apoptosis, and necrosis and reduce cell viability. The expression of c-Myc was downregulated by blockage of PVT1 and it shows that the expression of these two genes are correlated. CONCLUSION: The findings declare that inhibition of PVT1 could be a new target in the treatment of AML-M0 and help to approach more to treatments with fewer side effects.

Retraction Note: Blockage of miR-92a-3p with locked nucleic acid induces apoptosis and prevents cell proliferation in human acute megakaryoblastic leukemia.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Uncertainty in and around biophysical modelling: insights from interdisciplinary research on agricultural digitalization.

Agricultural digitalization is providing growing amounts of real-time digital data. Biophysical simulation models can help interpret these data. However, these models are subject to complex uncertainties, which has prompted calls for interdisciplinary research to better understand and communicate modelling uncertainties and their impact on decision-making. This article develops two corresponding insights from an interdisciplinary project in a New Zealand agricultural research organization. First, we expand on a recent Royal Society Open Science journal article (van der Bles et al. 2019 Royal Society Open Science 6, 181870 (doi:10.1098/rsos.181870)) and suggest a threefold conceptual framework to describe direct, indirect and contextual uncertainties associated with biophysical models. Second, we reflect on the process of developing this framework to highlight challenges to successful collaboration and the importance of a deeper engagement with interdisciplinarity. This includes resolving often unequal disciplinary standings and the need for early collaborative problem framing. We propose that both insights are complementary and informative to researchers and practitioners in the field of modelling uncertainty as well as to those interested in interdisciplinary environmental research generally. The article concludes by outlining limitations of interdisciplinary research and a shift towards transdisciplinarity that also includes non-scientists. Such a shift is crucial to holistically address uncertainties associated with biophysical modelling and to realize the full potential of agricultural digitalization.

Sensitivity of the knee joint response, muscle forces and stability to variations in gait kinematics-kinetics.

Sensitivity analysis of the knee joint response to variations in gait kinematics-kinetics as reported in the literature is crucial for improved understanding and more effective prevention-treatment strategies. Using our validated finite element-musculoskeletal (FE-MS) model of lower extremity, we investigate the sensitivity of knee anterior cruciate ligament (ACL), muscle, and contact forces plus stability to the reported scatter in asymptomatic gait data. Three highly loaded stance instants (25, 50 and 75%) with five levels (mean, ±0.5SD and ±SD) for each of six knee joint angles-moments are used employing Taguchi approach (25 experiments) and regression equations. ACL force drops significantly at larger flexion angles (all periods) and smaller internal moment (at 75% only) but increases with the flexion moment. Tibiofemoral (TF) medial-lateral contact force partitioning is found, contrary to the common claim, most sensitive to changes in the adduction angle and not in the adduction moment. Total TF contact force increases significantly at greater moments (but not angles), especially in the sagittal plane. Forces in lateral hamstrings are significantly influenced by changes in adduction angles-moments. Larger flexion moments (at 25 and 50%) significantly increase forces in quadriceps and on patellofemoral (PF) contact. Sagittal moment, adduction moment (at 75%) and flexion angle (at 25%) contribute most to the joint stability. A strong inverse correlation exists between the joint stability and the total TF compression force. These findings can be exploited to adapt and modify intact, injured and reconstructed knee joint responses during gait.

Invasive annuals respond more negatively to drought than native species.

In his foundational list of 'ideal weed' characteristics, Baker (1965) proposed that weedy plants maximize reproductive output under high resource availability. Since then, the idea that invasive plant species are more responsive to fluctuating resources compared with native or noninvasive species has gained considerable traction, although few studies extend this hypothesis to include reproductive output. We revisit Baker's hypothesis in the context of invasion and drought in California grasslands, exploring whether invasives show greater growth and reproductive responses to water availability compared with the native wildflowers they displace. In an outdoor potted study, we grew eight native and eight invasive species of annuals commonly found in southern California grasslands to reproductive maturity under both well-watered and drought conditions. While drought negatively impacted plant performance overall, invasives showed more negative responses for growth and reproductive traits. Invasives also grew larger than native species, especially under well-watered conditions, and produced seed with higher rates of germination. Invasives may be more negatively impacted by drought compared with natives, but they are also able to capitalize on high resource conditions and greatly increase reproductive output. Such opportunistic responses exhibited by invasives might explain previously observed fluctuations in their abundance under variable precipitation.

Can we condition native plants to increase drought tolerance and improve restoration success?

A common method in ecological restoration is the transplanting of nursery-grown seedlings to the field and, with proper resources, this technique can be highly successful. However, stressors such as drought may negatively impact plant performance and restoration success, especially in dryland ecosystems. Furthermore, increasing environmental change may hamper the ability of practitioners to restore native vegetation. A growing body of research suggests that exposing plants to a stressor may improve tolerance to subsequent stress events later in life. We sought to understand if such a phenomenon could be exploited in order to improve plant drought-tolerance and aid native plant restoration in southern California. In a multi-phase experiment, we first exposed seedlings of native perennials to episodic drought and then later compared the response of these plants to a second drought event to that of well-watered controls. We also transplanted replicates of both treatments to a restoration site in the field to test whether exposure to drought as a seedling could improve plant performance. Plant species responded to drought differently, with species exhibiting the full range of positive, neutral, and negative responses to temporal variability in water stress. However, some species appeared to benefit from drought preconditioning, exhibiting greater growth and increased water-use efficiency compared to well-watered plants. This suggests that simple applications of stress treatments could improve plant growth and stress tolerance, but the success of this method is likely very species specific. Restoration practitioners should consider conducting pilot studies with target plant species to better understand if this technique could assist in achieving restoration goals.

Clinical and laboratory evaluation of cured and non-cured patients with cutaneous leishmaniasis treated by Glucantime.

BACKGROUND & OBJECTIVES: Insufficient treatment of cutaneous leishmaniasis (CL) by conventional drugs is a major barrier in control strategies. This study was aimed to evaluate Glucantime efficacy and the susceptibility of Glucantime unresponsive and responsive CL isolates in the field and laboratory. METHODS: Chi-square test (x[2]) was used to determine the significance of difference between proportions in Glucantime-treated patients. The inhibitory activity of various concentrations of Glucantime against Leishmenia tropica stages was evaluated by a colorimetric cell viability MTT and macrophage assays. Mixed model, t-test and ANOVA were performed to determine the significance of difference between various concentrations of Glucantime unresponsive or responsive isolates and untreated control group and p <0.05 was defined as significant level. Altogether, 89.8% of the patients were cured by Glucantime, whilst 10.2% remained non-cured. RESULTS: The overall Glucantime efficacy in different age groups and genders was similar. The IC50 values of promastigotes and amastigotes for Glucanime unresponsive isolates were 2.1 and 2.6 times higher than the equivalent rates obtained for responsive cases, respectively. The overall mean number of amastigotes within macrophages in unresponsive isolates was significantly higher (32.68 ± 1.24) than that in responsive ones (18.68 ± 1.52, p <0.001). Glucantime unresponsive and responsive field isolates of anthroponotic CL (ACL) caused by L. tropica strongly correlated to in vitro assays. INTERPRETATION & CONCLUSION: Monitoring of Glucantime unresponsiveness by the health surveillance system is extremely important, where anthroponotic transmission occurs in humans. Hence, physicians should be aware of such clinical unresponsive presentations with ACL for antimonial therapeutic failure to improve management of disease in endemic regions.

Computation of the role of kinetics, kinematics, posterior tibial slope and muscle cocontraction on the stability of ACL-deficient knee joint at heel strike - Towards identification of copers from non-copers.

Rupture of anterior cruciate ligament (ACL) undermines normal activity and function of the knee joint and places it at higher risk of re-injury and degeneration. ACL reconstruction surgery neither necessarily ensures return to pre-injury activities nor alleviates risk of long-term degeneration. Here in this computational investigation of a lower-extremity hybrid model at heel strike (HS) of gait, we search for factors that influence the stability of the joint and hence the distinct performances between post-ACL injury copers and non-copers. Due to the very unstable state of the joint under the mean gait input data, joint rotations-moments, posterior tibial slope (PTS), and cocontraction were altered within the reported data in the literature and the effects on the joint stability (anterior tibial translation (ATT) and critical muscle stiffness coefficient (qcr)) were investigated. Results indicate that, in presence of both a small extension moment (0.1 or 0.2 Nm/kg) and a flexion rotation (∼5-8°), ACL-deficient (ACL-D) knee joint stability substantially improves to levels computed in the pre-injury intact joint. In addition, low cocontraction levels of 1-3% (in hamstrings and quads only and not in gastrocnemii) and reduced PTS (by 5°) further improve ACL-D joint stability. Therefore for a stable joint with ATT < 3 mm and qcr < 25 similar to those in the intact knee at HS, higher flexion angles (>5°) and a small extension moment (∼0.1-0.2 Nm/kg) (i.e., higher activity in hamstrings than quads) are required. A lower posterior tibial slope (by 5°) and a small minimum cocontraction level (1-3%) in hamstrings and quads (but not in gastrocnemii) are also beneficial. These results identify mechanisms likely in play at HS in gait of copers when compared to non-copers.

The effect of grapeseed oil on performance, rumen fermentation, antioxidant status and subcutaneous adipose fatty acid profile in lambs.

This experiment was carried out to examine the effect of grapeseed oil (GSO) on performance, rumen fermentation, antioxidant status and subcutaneous adipose fatty acid (FA) profile in lambs. Eighteen Baluchi lambs, 196 ± 14 days of age and 39.8 ± 1.7 kg body weight, were randomly assigned to three experimental diets: (i) diet without GSO (control), (ii) diet containing 2% of GSO (GSO2) and (iii) diet containing 4% of GSO (GSO4) for 42 days. Results showed that the experimental diets had no significant effect on dry matter intake and performance (p > 0.05). The supplemented diets with GSO had no effect on pH and NH3 -N of rumen fluid (p > 0.05), but GSO4 increased (p = 0.003) the concentration of propionic acid and reduced (p = 0.002) the concentration of acetate acid compared to the control. Inclusion 4% of GSO to the diet increased total antioxidant activity and decreased malondialdehyde in serum and muscle (p < 0.001). The level of blood glutathione peroxidase in diets containing GSO was higher than control diet (p = 0.02), but diets had no effect on superoxide dismutase in blood and muscle tissues (p > 0.05). The concentration of vaccenic acid, rumenic acid and linoleic acid and thus polyunsaturated fatty acid in subcutaneous fat was affected by experimental diets (p < 0.001), as control had the lowest amount of these FAs and GSO4 had the highest amount of them. Therefore, it can be concluded that the use of GSO up to 4% improves the antioxidant status and adipose fatty acid profile in lambs without effects on performance.

Capacity-oriented approaches to developing childhood obesity interventions: a systematic review.

Capacity-oriented approaches to health interventions seek to empower the target population or community to manage the health issue themselves using resources they can control. Positive deviance, resilience and asset-based approaches are three such methods of developing and implementing health interventions. This study aimed to review the efficacy of interventions explicitly applying these methods in addressing childhood obesity using adiposity as the primary outcome, measured by standardized body mass index. The search strategy was developed and implemented across four electronic databases. Of the 181 records identified and screened, 11 studies were identified as using a capacity-oriented approach overall. Asset-based approaches (n = 8 studies) consisted of 47 880 participants, positive deviance (n = 2 studies) consisted of 781 participants, and resilience-based interventions (n = 1 study) consisted of 35 participants. The asset-based approaches were mixed, with three of the eight studies showing a significant reduction in adiposity, while the other five did not find a difference. The positive deviance and resilience-based studies showed signs of efficacy in reducing adiposity. There was significant design heterogeneity across studies, and varied interpretations and definitions of the approaches were used. Further work should attempt to achieve some consensus on the use of these approaches to facilitate comparison and advance the science of capacity-oriented interventions for childhood obesity.

Computational stability of human knee joint at early stance in Gait: Effects of muscle coactivity and anterior cruciate ligament deficiency.

As one of the most complex and vulnerable structures of body, the human knee joint should maintain dynamic equilibrium and stability in occupational and recreational activities. The evaluation of its stability and factors affecting it is vital in performance evaluation/enhancement, injury prevention and treatment managements. Knee stability often manifests itself by pain, hypermobility and giving-way sensations and is usually assessed by the passive joint laxity tests. Mechanical stability of both the human knee joint and the lower extremity at early stance periods of gait (0% and 5%) were quantified here for the first time using a hybrid musculoskeletal model of the lower extremity. The roles of muscle coactivity, simulated by setting minimum muscle activation at 0-10% levels and ACL deficiency, simulated by reducing ACL resistance by up to 85%, on the stability margin as well as joint biomechanics (contact/muscle/ligament forces) were investigated. Dynamic stability was analyzed using both linear buckling and perturbation approaches at the final deformed configurations in gait. The knee joint was much more stable at 0% stance than at 5% due to smaller ground reaction and contact forces. Muscle coactivity, when at lower intensities (<3% of its maximum active force), increased dynamic stability margin. Greater minimum activation levels, however, acted asan ineffective strategy to enhance stability. Coactivation also substantially increased muscle forces, joint loads and ACL force and hence the risk of further injury and degeneration. A deficiency in ACL decreases total ACL force (by 31% at 85% reduced stiffness) and the stability margin of the knee joint at the heel strike. It also markedly diminishes forces in lateral hamstrings (by up to 39%) and contact forces on the lateral plateau (by up to 17%). Current work emphasizes the need for quantification of the lower extremity stability margin in gait.

Molecular modeling and molecular dynamics simulation study of archaeal leucyl-tRNA synthetase in complex with different mischarged tRNA in editing conformation.

Aminoacyl-tRNA synthetases (aaRSs) play important roles in maintaining the accuracy of protein synthesis. Some aaRSs accomplish this via editing mechanisms, among which leucyl-tRNA synthetase (LeuRS) edits non-cognate amino acid norvaline mainly by post-transfer editing. However, the molecular basis for this pathway for eukaryotic and archaeal LeuRS remain unclear. In this study, a complex of archaeal P. horikoshii LeuRS (PhLeuRS) with misacylated tRNALeu was modeled wherever tRNA's acceptor stem was oriented directly into the editing site. To understand the distinctive features of organization we reconstructed a complex of PhLeuRS with tRNA and visualize post-transfer editing interactions mode by performing molecular dynamics (MD) simulation studies. To study molecular basis for substrate selectivity by PhLeuRS's editing site we utilized MD simulation of the entire LeuRS complexes using a diverse charged form of tRNAs, namely norvalyl-tRNALeu and isoleucyl-tRNALeu. In general, the editing site organization of LeuRS from P.horikoshii has much in common with bacterial LeuRS. The MD simulation results revealed that the post-transfer editing substrate norvalyl-A76, binds more strongly than isoleucyl-A76. Moreover, the branched side chain of isoleucine prevents water molecules from being closer and hence the hydrolysis reaction slows significantly. To investigate a possible mechanism of the post-transfer editing reaction, by PhLeuRS we have determined that two water molecules (the attacking and assisting water molecules) are localized near the carbonyl group of the amino acid to be cleaved off. These water molecules approach the substrate from the opposite side to that observed for Thermus thermophilus LeuRS (TtLeuRS). Based on the results obtained, it was suggested that the post-transfer editing mechanism of PhLeuRS differs from that of prokaryotic TtLeuRS.

Apoptosis-inducing and antiproliferative effect by inhibition of miR-182-5p through the regulation of CASP9 expression in human breast cancer.

Recent advances in molecular medicine and gene therapy have offered new effective achievements in the treatment of cancers. One of the molecular research lines for the diagnosis and treatment of cancer is the use of microRNAs (miRNAs), which are single-stranded noncoding RNAs. miRNAs are involved in the post-transcriptional regulation of gene expression and have a role in the growth, differentiation, cell death and cancer development. One of the miRNAs that showed upregulation in breast cancer is miR-182-5p. Oncogenic features of miR-182-5p in some cancers were confirmed. In the present study, blockage of miR-182-5p was performed in human breast cancer cell line (MCF-7) using locked nucleic acid (LNA)-anti-miR. MTT (3-[4,5 dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide) assay and annexin/propidium iodide staining at different time points after LNA-anti-miR-182-5p transfection were accomplished. Our results showed that miR-182-5p inhibition can reduce the viability of MCF-7 cells because of apoptosis induction, probably through the upregulation of CASP9. A western blot analysis revealed that the expression of CASP9 protein is associated with miR-182-5p inhibition. The CASP9 protein acts as an initiator caspase of apoptosis in the mitochondrial cell death pathway. Our results can be used in translational medicine for future investigation in breast cancer and approach treatment based on antisense therapy.

The Effect of increasing levels of date palm (Phoenix dactylifera L.) seed on the performance, ruminal fermentation, antioxidant status and milk fatty acid profile of Saanen dairy goats.

This study was conducted to examine the effect of date palm (Phoenix dactylifera L.) seed (DPS) on the performance, ruminal fermentation, antioxidant status and milk fatty acid (FA) profile of dairy goats. Eight multiparous Saanen dairy goats, averaging 97 ± 7 days in milk (DIM) and 2150 ± 130 g of milk production, were used in a 4 × 4 replicated Latin square design. Experimental diets contained 0% (control), 6% (DPS6), 12% (DPS12) and 18% (DPS18) of DPS. Dry matter intake (DMI), milk production, and the composition and digestibility of the dry matter (DM), organic matter (OM), neutral detergent fibre (NDF) and crude protein (CP) were not affected by the diets. Adding DPS to the diet increased linearly total antioxidant capacity (TAC) in milk and blood (p < 0.05). No significant difference was found in the malondialdehyde (MDA) content in milk and blood. Superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) activities in the blood, ruminal pH, NH3 -N, ruminal total volatile fatty acid (VFA), acetate, butyrate, isovalerate and valeric acid concentration had no significant effect among the diets. The propionate acid concentration decreased linearly as DPS was added to the diet, and DPS18 had minimum propionate acid concentration (p < 0.05). The inclusion of DPS18 to diets increased linearly the proportion of C18:0 (compared to control) as well as cis-18:1, trans-18:1 (compared to control and DPS6) and decreased C14:1 (compared to control) in milk fat (p < 0.05). The concentration of conjugated linoleic acid (CLA) in DPS18 was higher than other diets (p = 0.04). The results of this study implied that replacing DPS with a part of dairy goats' ration had no negative effects on the performance of the animals and could also improve the antioxidant activity and increase the concentration of CLA in their milk.