Deuterium in marine organic biomarkers: toward a new tool for quantifying aquatic mixotrophy.

The traditional separation between primary producers (autotrophs) and consumers (heterotrophs) at the base of the marine food web is being increasingly replaced by the paradigm that mixoplankton, planktonic protists with the nutritional ability to use both phago(hetero)trophy and photo(auto)trophy to access energy are widespread globally. Thus, many 'phytoplankton' eat, while 50% of 'protozooplankton' also perform photosynthesis. Mixotrophy may enhance primary production, biomass transfer to higher trophic levels and the efficiency of the biological pump to sequester atmospheric CO2 into the deep ocean. Although this view is gaining traction, science lacks a tool to quantify the relative contributions of autotrophy and heterotrophy in planktonic protists. This hinders our understanding of their impacts on carbon cycling within marine pelagic ecosystems. It has been shown that the hydrogen (H) isotopic signature of lipids is uniquely sensitive to heterotrophy relative to autotrophy in plants and bacteria. Here, we explored whether it is also sensitive to the trophic status in protists. The new understanding of H isotope signature of lipid biomarkers suggests it offers great potential as a novel tool for quantifying the prevalence of mixotrophy in diverse marine microorganisms and thus for investigating the implications of the 'mixoplankton' paradigm.

A high-temperature water vapor equilibration method to determine non-exchangeable hydrogen isotope ratios of sugar, starch and cellulose.

The analysis of the non-exchangeable hydrogen isotope ratio (δ2 Hne ) in carbohydrates is mostly limited to the structural component cellulose, while simple high-throughput methods for δ2 Hne values of non-structural carbohydrates (NSC) such as sugar and starch do not yet exist. Here, we tested if the hot vapor equilibration method originally developed for cellulose is applicable for NSC, verified by comparison with the traditional nitration method. We set up a detailed analytical protocol and applied the method to plant extracts of leaves from species with different photosynthetic pathways (i.e., C3 , C4 and CAM). δ2 Hne of commercial sugars and starch from different classes and sources, ranging from -157.8 to +6.4‰, were reproducibly analysed with precision between 0.2‰ and 7.7‰. Mean δ2 Hne values of sugar are lowest in C3 (-92.0‰), intermediate in C4 (-32.5‰) and highest in CAM plants (6.0‰), with NSC being 2 H-depleted compared to cellulose and sugar being generally more 2 H-enriched than starch. Our results suggest that our method can be used in future studies to disentangle 2 H-fractionation processes, for improving mechanistic δ2 Hne models for leaf and tree-ring cellulose and for further development of δ2 Hne in plant carbohydrates as a potential proxy for climate, hydrology, plant metabolism and physiology.

Implementation of In vivo exposure therapy to decrease injury-related fear in females with a history of ACL-Reconstruction: A pilot study.

OBJECTIVE: The objective of this pilot study was to examine the preliminary feasibility and efficacy of in vivo exposure therapy (IVET) to decrease injury-related fear in females with history of ACLR. DESIGN: Pilot Study. SETTING: Sports Medicine Research Laboratory. PARTICIPANTS: 12 female participants with history of ACLR (≥ 1 year post-operative) were randomized into a 5-week IVET group (n = 6) or 5-week sham physical activity (PA) monitoring group (n = 6). MAIN OUTCOME MEASURES: The independent variables were Group and Time. The dependent variables were the Photographic Series of Sports Activities for ACLR (PHOSA-ACLR) and the Tampa Scale of Kinesiophobia-11 (TSK-11) scores. A Group x Time repeated measures two-way analysis of variance was completed for the PHOSA-ACLR and the TSK-11. Partial η2 effect sizes were used to examine clinically meaningful differences. RESULTS: High retention and adherence rates were observed in the intervention group. The PHOSA-ACLR exhibited a significant main effect for Time (F1,10 = 9.92, p = 0.01, partial η2 = 0.50), but not for Group. No statistically significant or clinically meaningful differences were observed for the TSK-11. CONCLUSION: Both groups exhibited decreased injury-related fear for specific functional tasks. Future research should further examine the efficacy of IVET and PA monitoring to decrease injury-related fear in patients after ACLR.

Neuroplasticity in Corticolimbic Brain Regions in Patients After Anterior Cruciate Ligament Reconstruction.

CONTEXT: Fear has been cited as the primary barrier to return to sport (RTS) by athletes after anterior cruciate ligament reconstruction (ACLR). Understanding the neural factors that contribute to fear after ACLR may help us to identify interventions for this population. OBJECTIVE: To characterize the underlying neural substrate of injury-related fear in patients after ACLR versus healthy matched control individuals during a picture imagination task (PIT) consisting of sport-specific images and images of activities of daily living (ADL). DESIGN: Case-control study. SETTING: Research laboratory. PATIENTS OR OTHER PARTICIPANTS: A total of 24 right-hand-dominant participants (12 with left-sided ACLR and 12 control individuals) were enrolled. Participants underwent full-brain functional magnetic resonance imaging. MAIN OUTCOME MEASURE(S): Functional data were acquired using blood oxygen level-dependent (BOLD) echoplanar imaging. Independent t tests were conducted to identify between-groups differences in BOLD signal changes during all images of the PIT. Paired t tests were computed to examine differences in BOLD signal change between sport-specific and ADL images in the ACLR group. RESULTS: Increased activation in the inferior parietal lobule and the mediodorsal thalamus was observed during PIT in the ACLR group. An inability to suppress the default mode network in the ACLR group was noted. The ACLR group exhibited increased activation in the cerebellum and inferior occipital regions during the sport-specific images versus the ADL images, but no other regions of interest demonstrated differences. CONCLUSION: After ACLR, patients may be more predisposed to fear, anxiety, and pain during sport-specific activities and ADLs. Psychosocial interventions may be warranted after ACLR to reduce injury-related fear and mitigate potentially maladaptive neuroplasticity.

Examining the Relationship Between Neuroplasticity and Learned Helplessness After ACLR: Early Versus Late Recovery.

CONTEXT: Altered neural signaling is known to have a direct impact on psychological wellness. Therefore, disruptions in neural signaling after anterior cruciate ligament reconstruction may influence psychological dysfunction, in some cases manifesting as learned helplessness. Helplessness is a psychological paradigm that presents as altered neuromuscular control, reduced motivation, and psychological deficits. OBJECTIVES: The authors sought to evaluate the relationship between helplessness, neural activity, and quadriceps function at different time points after anterior cruciate ligament reconstruction. EVIDENCE ACQUISITION: Twenty-nine individuals with unilateral anterior cruciate ligament reconstruction were categorized into early group (<2 y, age: 19.13 [2.18] y; height: 1.77 [0.11] m; mass: 76.903 [11.87] kg) or late group (>2 y, age: 22 [23] y; height: 1.67 [0.07] m; mass: 65.66 [11.33] kg). Quadriceps function (activation and strength), spinal-reflexive and corticospinal excitability (active motor thresholds and motor evoked potentials), and helplessness were obtained. A principal component analysis was performed by group (early and late) to identify which factors of helplessness were most associated with neural activity and quadriceps function. Pearson product moment correlation analyses were performed by group to determine associations between individual components and main outcomes. EVIDENCE SYNTHESIS: In the early group, cognitive readiness was associated with quadriceps strength of the injured limb (r2 = .513, P = .004), and self-awareness/management was associated with motor threshold of the injured limb (r2 = .238, P = .05). In the late group, intrinsic helplessness was associated with motor output of injured limb (r2 = .653, P = .01). CONCLUSION: Helplessness is made up of several attributional constructs, which are altered at different phases of recovery. Helplessness constructs interact differently with neural activity and quadriceps function across time. These findings are preliminary and do not establish a causal link between neural alterations and learned helplessness. Future studies should serially evaluate both changes in neural activity and learned helplessness attributes throughout recovery.

Phenolic acid phenethylesters and their corresponding ketones: Inhibition of 5-lipoxygenase and stability in human blood and HepaRG cells.

5-lipoxygenase (5-LO) catalyzes the biosynthesis of leukotrienes, potent lipid mediators involved in inflammatory diseases, and both 5-LO and the leukotrienes are validated therapeutic targets. Caffeic acid phenethyl ester (CAPE) is an effective inhibitor of 5-LO and leukotriene biosynthesis but is susceptible to hydrolysis by esterases. In this study a number of CAPE analogues were synthesized with modifications to the caffeoyl moiety and the replacement of the ester linkage with a ketone. Several new molecules showed better inhibition of leukotriene biosynthesis than CAPE in isolated human neutrophils and in whole blood with IC50 values in the nanomolar (290-520 nmol/L) and low micromolar (1.0-2.3 µmol/L) ranges, respectively. Sinapic acid and 2,5-dihydroxy derivatives were more stable than CAPE in whole blood, and ketone analogues were degraded more slowly in HepaRG hepatocyte cultures than esters. All compounds underwent modification consistent with glucuronidation in HepaRG cultures as determined using LC-MS/MS analysis, though the modified sinapoyl ketone (10) retained 50% of its inhibitory activity after up to one hour of incubation. This study has identified at least one CAPE analogue, compound 10, that shows favorable properties that warrant further in vivo investigation as an antiinflammatory compound.

Discovery of a novel 2,5-dihydroxycinnamic acid-based 5-lipoxygenase inhibitor that induces apoptosis and may impair autophagic flux in RCC4 renal cancer cells.

The inhibition of 5-lipoxygenase (5-LO), the key enzyme for the biosynthesis of leukotrienes (LTs), has generated increasing enthusiasm as anti-inflammatory and antitumor strategies in recent years. Based on our previous studies, we synthesized a series of dihydroxycinnamic acid-based analogs that might be 5-LO inhibitors. LTs biosynthesis inhibition in HEK293 cells and polymorphonuclear leukocytes (PMNL) was measured and antitumor activities were investigated in Renal Cell Carcinoma (RCC). Results showed that the 2,5-dihydroxycinnamic acid phenethyl ester (10b) was the best 5-LO inhibitor and was 7-fold more potent than Zileuton (1), the only clinically approved 5-LO inhibitor. 2,5-Dihydroxy substitution was more favorable to 5-LO inhibition since compound 10b is twice as active as CAPE (2) which is a 3,4-dihydroxylcinnamic acid ester. Meanwhile, 10b reduced the cell viability of renal cancer cells  and was more selective toward RCC4 and 786.0 cells which are deficient for the Von Hippel-Lindau (VHL) tumor suppressor gene. As to the underlying cell-death mechanisms, 10b induced apoptosis in VHL-deficient RCC4 cells. Also, increases in LC3B and p62 expression suggest a blockage of the autophagic flux in RCC in response to 10b.

Influence of starch deficiency on photosynthetic and post-photosynthetic carbon isotope fractionations.

Carbon isotope (13C) fractionations occurring during and after photosynthetic CO2 fixation shape the carbon isotope composition (δ13C) of plant material and respired CO2. However, responses of 13C fractionations to diel variation in starch metabolism in the leaf are not fully understood. Here we measured δ13C of organic matter (δ13COM), concentrations and δ13C of potential respiratory substrates, δ13C of dark-respired CO2 (δ13CR), and gas exchange in leaves of starch-deficient plastidial phosphoglucomutase (pgm) mutants and wild-type plants of four species (Arabidopsis thaliana, Mesembryanthemum crystallinum, Nicotiana sylvestris, and Pisum sativum). The strongest δ13C response to the pgm-induced starch deficiency was observed in N. sylvestris, with more negative δ13COM, δ13CR, and δ13C values for assimilates (i.e. sugars and starch) and organic acids (i.e. malate and citrate) in pgm mutants than in wild-type plants during a diel cycle. The genotype differences in δ13C values could be largely explained by differences in leaf gas exchange. In contrast, the PGM-knockout effect on post-photosynthetic 13C fractionations via the plastidic fructose-1,6-bisphosphate aldolase reaction or during respiration was small. Taken together, our results show that the δ13C variations in starch-deficient mutants are primarily explained by photosynthetic 13C fractionations and that the combination of knockout mutants and isotope analyses allows additional insights into plant metabolism.

Learned Helplessness After Anterior Cruciate Ligament Reconstruction: An Altered Neurocognitive State?

Traumatic knee injuries, such as anterior cruciate ligament (ACL) sprains, have detrimental effects on long-term health as they initiate a cycle of chronic pain, physical inactivity, and disability. Alterations in strength and neural activity are factors that contribute to rehabilitation failure after ACL reconstruction (ACLR); however, psychological deficits also hinder rehabilitative success. Neural impairments observed following injury and ACLR may be associated with psychological dysfunction, a phenomenon defined as learned helplessness (LH). The proposed framework establishes the link between depressed neural activity and psychological dysfunction after ACL injury using foundational evidence from neuroscience and psychology to support the integration of LH into recovery.

Identification of Peracetylated Quercetin as a Selective 12-Lipoxygenase Pathway Inhibitor in Human Platelets.

The inflammatory response is necessary for the host's defense against pathogens; however, uncontrolled or unregulated production of eicosanoids has been associated with several types of chronic inflammatory diseases. Thus, it is not surprising that enzymes implicated in the production of eicosanoids have been strategically targeted for potential therapeutic approaches. The 12(S)-hydroxyeicosatetraenoic acid [12(S)-HETE] lipid mediator is among inflammatory molecules that are abundantly produced in various diseases and is primarily biosynthesized via the 12(S)-lipoxygenase pathway. The effects of the abundance of 12(S)-HETE and its contribution to several chronic inflammatory diseases have been well studied over the last few years. While most developed compounds primarily target the 5-lipoxygenase (5-LO) or the cyclooxygenase (COX) pathways, very few compounds selectively inhibiting the 12-lipoxygenase (12-LO) pathway are known. In this study, we examined whether the distribution of hydroxyl groups among flavones could influence their potency as 12-LO inhibitors. Using human platelets, the human embryonic kidney 293 (HEK293) cell line expressing 5-LO, and human polymorphonuclear leukocytes (PMNLs) we investigated the effects of these compounds on several inflammatory pathways, namely, 12-LO, 5-LO, and COX. Using high-resolution respirometry and flow cytometry, we also evaluated some normal cell functions that could be modulated by our compounds. We identified a peracetylated quercetin (compound 6) that exerts potent inhibitory activity toward the platelet 12-LO pathway (IC50 = 1.53 µM) while having a lesser affinity toward the COX pathway. This study characterizes the peracetylated quercetin (compound 6) as a more selective platelet-type 12-LO inhibitor than baicalein, with no measurable nontargeted effects on the platelet's activation or overall cell's oxygen consumption.

Eccentric cross-exercise after anterior cruciate ligament reconstruction: Novel case series to enhance neuroplasticity.

OBJECTIVES: Substantial changes in neural function are historically present after anterior cruciate ligament reconstruction (ACLR), and are not rectified with traditional rehabilitation. Cross-exercise is a potential means to enhance neural excitability and improve recovery after ACLR. Hence our purpose, was to detail changes in brain activation, neural excitability and patient-reported outcomes in a cohort that completed an 8-week quadriceps-focused eccentric cross-exercise training program immediately following ACLR. DESIGN: Case series. SETTING: University. PARTICPANTS: Five patients participated in an 8-week (24-session) eccentric cross-exercise intervention after ACLR. MAIN OUTCOME MEASURES: Brain activation, neural activity and patient-reported outcomes were evaluated within 2 weeks post-ACLR and again at 10-weeks post-ACLR after the intervention. Each cross-exercise session consisted of 4 sets of 10 isokinetic eccentric contractions at 60 deg/sec with the noninvolved limb. RESULTS: Following the intervention, patients demonstrated a facilitated spinal reflexive and muscle activity response from the motor cortex during a time when these measures are known to be depressed. Patients also demonstrated a reduce dependence on frontal cortex activity to generate quadriceps contractions. Further patients reported significant reductions in pain and symptoms and greater knee function. CONCLUSIONS: Eccentric cross-exercise after ACLR helps to facilitate positive adaptations in neural function and patient reported outcomes.

Sinapic acid phenethyl ester as a potent selective 5-lipoxygenase inhibitor: Synthesis and structure-activity relationship.

Given the hepatotoxicity and an unfavorable pharmacokinetic profile of zileuton (Zyflo® ), currently the only approved and clinically used 5-Lipoxygenase (5-LO) inhibitor, the search for potent and safe 5-LO inhibitors is highly demanded. The action of several phenolic acid phenethyl esters as potential 5-Lipoxygenase (5-LO) inhibitors has been investigated. For this purpose, a series of 14 phenethyl esters was synthesized and their impact on 5-LO inhibition was evaluated. The effects of position and number of hydroxyl and methoxy groups on the phenolic acid were investigated. The shortening of the linker between the carbonyl and the catechol moiety as well as the presence of the α,ß-unsaturated carbonyl group was also explored. The sinapic acid phenethyl ester (10), which can be named SAPE (10) by analogy to caffeic acid phenethyl ester (CAPE), inhibited 5-LO in a concentration-dependent manner and outperformed both zileuton (1) and CAPE (2). With an IC50 of 0.3 µm, SAPE (10) was threefold more potent than CAPE (2) and 10-fold more potent than zileuton (1), the only 5-LO inhibitor approved for clinical use. Unlike CAPE (2), SAPE (10) had no effect on 12-lipoxygenase (12-LO) and less effect on cyclooxygenase 1 (COX-1) which makes it a more selective 5-LO inhibitor.

Quadriceps Function, Knee Pain, and Self-Reported Outcomes in Patients With Anterior Cruciate Ligament Reconstruction.

CONTEXT: Interactions among muscle strength, pain, and self-reported outcomes in patients with anterior cruciate ligament reconstruction (ACLR) are not well understood. Clarifying these interactions is of clinical importance because improving physical and psychological function is thought to optimize outcomes after ACLR. OBJECTIVE: To examine the relationships among neuromuscular quadriceps function, pain, self-reported knee function, readiness to return to activity, and emotional response to injury both before and after ACLR. DESIGN: Descriptive laboratory study. PATIENTS OR OTHER PARTICIPANTS: Twenty patients (11 females and 9 males; age = 20.9 ± 4.4 years, height = 172.4 ± 7.5 cm, weight = 76.2 ± 11.8 kg) who were scheduled to undergo unilateral ACLR. MAIN OUTCOME MEASURE(S): Quadriceps strength, voluntary activation, and pain were measured at presurgery and return to activity, quantified using maximal voluntary isometric contractions (MVICs), central activation ratio, and the Knee Injury and Osteoarthritis Outcome Score pain subscale, respectively. Self-reported knee function, readiness to return to activity, and emotional responses to injury were evaluated at return to activity using the International Knee Documentation Committee questionnaire (IKDC), ACL Return to Sport After Injury scale (ACL-RSI), and Psychological Response to Sport Injury Inventory (PRSII), respectively. Pearson product moment correlations and linear regressions were performed using raw values and percentage change scores. RESULTS: Presurgical levels of pain significantly predicted 31% of the variance in the ACL-RSI and 29% in the PRSII scores at return to activity. The MVIC and pain collected at return to activity significantly predicted 74% of the variance in the IKDC, whereas only MVIC significantly predicted 36% of the variance in the ACL-RSI and 39% in the PRSII scores. Greater increases in MVIC from presurgery to return to activity significantly predicted 49% of the variance in the ACL-RSI and 59% of the variance in the IKDC scores. CONCLUSION: Decreased quadriceps strength and higher levels of pain were associated with psychological responses in patients with ACLR. A comprehensive approach using traditional rehabilitation that includes attention to psychological barriers may be an effective strategy to improve outcomes in ACLR patients.

2 H-fractionations during the biosynthesis of carbohydrates and lipids imprint a metabolic signal on the δ2 H values of plant organic compounds.

Hydrogen (H) isotope ratio (δ2 H) analyses of plant organic compounds have been applied to assess ecohydrological processes in the environment despite a large part of the δ2 H variability observed in plant compounds not being fully elucidated. We present a conceptual biochemical model based on empirical H isotope data that we generated in two complementary experiments that clarifies a large part of the unexplained variability in the δ2 H values of plant organic compounds. The experiments demonstrate that information recorded in the δ2 H values of plant organic compounds goes beyond hydrological signals and can also contain important information on the carbon and energy metabolism of plants. Our model explains where 2 H-fractionations occur in the biosynthesis of plant organic compounds and how these 2 H-fractionations are tightly coupled to a plant's carbon and energy metabolism. Our model also provides a mechanistic basis to introduce H isotopes in plant organic compounds as a new metabolic proxy for the carbon and energy metabolism of plants and ecosystems. Such a new metabolic proxy has the potential to be applied in a broad range of disciplines, including plant and ecosystem physiology, biogeochemistry and palaeoecology.

New Hydroxycinnamic Acid Esters as Novel 5-Lipoxygenase Inhibitors That Affect Leukotriene Biosynthesis.

Leukotrienes are inflammatory mediators that actively participate in the inflammatory response and host defense against pathogens. However, leukotrienes also participate in chronic inflammatory diseases. 5-lipoxygenase is a key enzyme in the biosynthesis of leukotrienes and is thus a validated therapeutic target. As of today, zileuton remains the only clinically approved 5-lipoxygenase inhibitor; however, its use has been limited due to severe side effects in some patients. Hence, the search for a better 5-lipoxygenase inhibitor continues. In this study, we investigated structural analogues of caffeic acid phenethyl ester, a naturally-occurring 5-lipoxygenase inhibitor, in an attempt to enhance the inhibitory activity against 5-lipoxygenase and determine structure-activity relationships. These compounds were investigated for their ability to attenuate the biosynthesis of leukotrienes. Compounds 13 and 19, phenpropyl and diphenylethyl esters, exhibited significantly enhanced inhibitory activity when compared to the reference molecules caffeic acid phenethyl ester and zileuton.

Substituted Caffeic and Ferulic Acid Phenethyl Esters: Synthesis, Leukotrienes Biosynthesis Inhibition, and Cytotoxic Activity.

Glioblastoma multiforme (GBM) is an aggressive brain tumor that correlates with short patient survival and for which therapeutic options are limited. Polyphenolic compounds, including caffeic acid phenethyl ester (CAPE, 1a), have been investigated for their anticancer properties in several types of cancer. To further explore these properties in brain cancer cells, a series of caffeic and ferulic acid esters bearing additional oxygens moieties (OH or OCH3) were designed and synthesized. (CAPE, 1a), but not ferulic acid phenethyl ester (FAPE, 1b), displayed substantial cytotoxicity against two glioma cell lines. Some but not all selected compounds derived from both (CAPE, 1a) and (FAPE, 1b) also displayed cytotoxicity. All CAPE-derived compounds were able to significantly inhibit 5-lipoxygenase (5-LO), however FAPE-derived compounds were largely ineffective 5-LO inhibitors. Molecular docking revealed new hydrogen bonds and π-π interactions between the enzyme and some of the investigated compounds. Overall, this work highlights the relevance of exploring polyphenolic compounds in cancer models and provides additional leads in the development of novel therapeutic strategies in gliomas.

Structure-activity relationship of caffeic acid phenethyl ester analogs as new 5-lipoxygenase inhibitors.

Leukotrienes (LTs) are a class of lipid mediators implicated in numerous inflammatory disorders. Caffeic acid phenethyl ester (CAPE) possesses potent anti-LTs activity through the inhibition of 5-lipoxygenase (5-LO), the key enzyme in the biosynthesis of LTs. In this study, we describe the design and synthesis of CAPE analogs as radical scavengers and 5-LO inhibitors. Caffeic esters bearing propargyl and allyl linkers between the caffeoyl and aryl moieties (4a-i and 5a-i, respectively) were synthesized by Sonogashira and Heck cross-coupling reactions to probe the effects of flexibility and aryl substitution on 5-LO inhibition. Caffeoyl alcohol and ethers (6, 7a-b) as well as caffeoyl aldehyde and ketones (8a-e) were synthesized to elucidate the importance of the ester linkage for inhibitory activity. All tested compounds proved to be good radical scavengers (IC50 of 10-30 µm). After preliminary anti-LTs activity screening in HEK293 cell models, 5-LO inhibition potential of selected compounds was determined in human polymorphonuclear leukocytes (PMNL). Most screened compounds outperformed CAPE 3 in concentration-dependent assays on PMNL, with ester dimers 4i and 5i along with caffeoyl ethers 7a-b being roughly eight-, seven-, and 16-fold more potent than Zileuton, with IC50 values of 0.36, 0.43, and 0.18 µm, respectively.

Effect of O6-Substituted Guanine Analogs on O6-methylguanine DNA-methyltransferase Expression and Glioblastoma Cells Viability.

BACKGROUND: Glioblastoma multiforme (GBM) is often associated with a poor survival prognostic for patients. The main reason seems to be the acquired or inherent resistance to the chemotherapeutic agent used to treat the tumor, temozolomide (TMZ). To this day, the most recognized pathway of resistance is the DNA Direct Repair pathway by the means of the protein O6- methylguanine DNA-methyltransferase (MGMT). OBJECTIVES: To design and synthesize a series of MGMT inhibitors that can sensitize GBM cells to TMZ. METHODS: Twenty-five O6-alkyl, O6-aryl and O6-substituted-aryl guanine analogs including nine novel compounds were synthesized, characterized, analyzed by molecular docking and tested on the T98G GBM cells viability. RESULTS: Following molecular modeling with MGMT, the newly designed compounds 19, 22, and 24 emerged as the most promising MGMT ligands and displayed modest cytotoxicity. Guanine analog (19), bearing a p-nitrobenzyl moiety, reduced considerably the O6-methylguanine DNAmethyltransferase expression level. When combined with TMZ (1), which is used as first line treatment for brain tumors, compounds 19, 22, and 24 decreased T98G cells proliferation by 32%, 68% and 50%, respectively. TMZ (1) displayed negligible effect on the proliferation of these cells further supporting the notion that this cell model is resistant to this alkylating agent. CONCLUSION: Overall, these results notably highlight a group of MGMT inhibitors that warrants further exploration in the development of therapeutic options to circumvent TMZ resistance in brain tumors.

Athletic Trainers' Skills in Identifying and Managing Athletes Experiencing Psychological Distress.

CONTEXT: Athletic trainers (ATs) commonly use psychological skills during sport rehabilitation. However, little is known about their ability to accurately implement these skills. OBJECTIVE: To assess ATs' skills in identifying psychological symptoms, selecting appropriate strategies, and making referral decisions for athletes experiencing various degrees of psychological distress. DESIGN: Cross-sectional study. SETTING: Participants were recruited using the National Athletic Trainers' Association professional member database. PATIENTS OR OTHER PARTICIPANTS: Of the 2998 ATs who were selected randomly, 494 (16.5%) partially completed the questionnaire and 326 (10.9%) completed the entire survey (mean age = 34.7 ± 10.8 years, mean years of experience = 11.3 ± 9.9). MAIN OUTCOME MEASURE(S): Using the Web-based questionnaire created for this study, we collected ATs' demographic information and assessed their perceptions about responsibilities as ATs, psychosocial competencies, training in sport psychology, and referral behaviors. Additionally, respondents were asked to identify symptoms, match psychological strategies (eg, goal setting, imagery, progressive muscle relaxation), and make referral decisions for athletes in 3 case vignettes. RESULTS: The ATs demonstrated high accuracy in identifying symptoms and making referral decisions but struggled in selecting appropriate psychosocial strategies for athletes. Stepwise regression analyses revealed that ATs who had had specific coursework in sport psychology were able to more accurately identify symptoms (t = 3.01, P < .01), and those ATs with more experience reported lower accuracy scores for their intended course of action (t  =  -2.25, P < .05). CONCLUSIONS: Our analogue research design provided new insights into ATs' knowledge and use of sport psychology in practice. The results highlighted the importance of coursework focusing on applied areas of sport psychology in the training of ATs.