Background. The amount of excess weight loss (%EWL) among obese patients after bariatric surgery varies greatly. However, reliable predictors have not been established yet. The present study evaluated the preoperative psychological burden, coping style, and motivation to lose weight as factors determining postoperative treatment success. Methods. The sample included 64 morbidly obese patients with a preoperative BMI of 51 ± 8 kg/m2 who had undergone laparoscopic sleeve gastrectomy (LSG). Well-established questionnaires were applied before surgery to assess the psychological burden in terms of “perceived stress” (PSQ-20), “depression” (PHQ-9), “anxiety” (GAD-7), and “mental impairment” (ISR) as well as coping style (Brief COPE) and motivation to lose weight. %EWL as an indicator for treatment success was assessed on average 20 months after surgery. Results. Based on the %EWL distribution, patients were classified into three %EWL groups: low (14–39%), moderate (40–59%), and high (60–115%). LSG patients with high %EWL reported significantly more “active coping” behavior prior to surgery than patients with moderate and low %EWL. Patients' preoperative psychological burden and motivation to lose weight were not associated with %EWL. Conclusion. An “active coping” style might be of predictive value for better weight loss outcomes in patients following LSG intervention. Currently, 13% of the world's adult population aged 18 years and older are obese, with a body mass index (BMI) ≥ 30 kg/m2 is recognized as the lowest limit of overweight, and therefore excess weight is calculated relative to a BMI of 25 kg/m2 [ versus failure, with an arbitrarily defined cut-off at 50% EWL, does not reflect the considerably wide individual variation in postoperative weight loss that has been described in previous studies [In previous studies of weight loss outcomes after bariatric surgery, the amount of excess weight loss as a percentage (%EWL) was commonly used as a marker of weight loss success EWL ≥ 50%) or weight loss failure (EWL < 50%) , 5. %EWL0% or wei25 kg/m2 , 7. Howe studies , 9. Appr studies , 11; ina studies . SurprisGiven the background of a multifactorial etiology of obesity, a multidisciplinary evaluation before bariatric surgery is recommended, and the assessment of psychosocial factors in addition to the medical examination has become highly relevant , 14. HowPsychological factors and mental disorders have been associated with weight loss results after bariatric surgery. Notably, up to 70% of obese patients considering bariatric surgery present with high rates of mental comorbidities, with depression, anxiety, and eating disorders being the most prevalent , 18. WitObese patients with comorbid mental disorders might have difficulties in adhering to the behavioral changes required to benefit from bariatric surgery . However versus patients seeking conservative treatment options [ versus resilient/high-functioning prebariatric women [Coping strategies, used when confronting difficult situations in daily life, and the motivation to undergo surgery might also be related to different weight loss outcomes; however, research is scarce. Recent studies have reported more avoidant and (delegated) active coping in patients seeking bariatric surgery options  or more ic women . AlthougTo date, most research on the role of prebariatric psychological variables in treatment success has focused on gastric bypass surgery. Laparoscopic sleeve gastrectomy (LSG) as a restrictive single-stage procedure is relatively new in the field of bariatric surgery but has already proven its efficacy in weight reduction with low surgical risks . To our Hence, assessing a broad range of preoperative patient characteristics might be useful to identify homogeneous subgroups of LSG patients with different needs to tailor interventions and optimize postoperative weight loss outcomes and well-being. Therefore, the main aim of the present study was to characterize patients with low, moderate, and high postoperative %EWL retrospectively using between-group comparisons to examine whether LSG patients with different %EWL levels after surgery differed preoperatively in (1) their psychological burden in terms of the levels of “perceived stress,” “depression,” “anxiety,” and “mental impairment,” (2) coping style , and (3) their motivation to lose weight. As a secondary aim, we wanted to study whether the %EWL groups differed on a range of preoperative patient characteristics, such as (a) weight and BMI, (b) sociodemographic status, (c) clinically diagnosed comorbidities including eating disorders, and (d) the use of psychotherapy. To do so, LSG patients were evaluated approximately 12 months before surgery and on average 20 months after surgery.As part of the presurgical psychosomatic evaluation for bariatric intervention at the Multidisciplinary Obesity Center of the University Hospital Charité-Universitätsmedizin Berlin in Germany, patients who planned to undergo bariatric surgery were assessed by an experienced clinical psychologist or physician specialized in psychosomatic medicine. A semistructured interview was performed for the psychosocial assessment and diagnosis of mental disorders including eating disorders according to the International Classification of Diseases (ICD-10) . Addition = 68) of the patients participated in the follow-up, and 29% (n = 28) of them declined participation or could not be reached. For the present study, we excluded patients with less than 1 year of follow-up after LSG surgery (n = 4). Moreover, bariatric surgery patients who underwent gastric banding or Roux-en-Y gastric bypass surgery were not included in our study because we sought to assess a homogeneous sample and to eliminate the effect of type of surgical procedure on %EWL .The present cohort consisted of 64 morbidly obese patients (46 women and 18 men) who had undergone LSG. The average points of assessment were 12 months prior to surgery and 20 months after surgery. The study was approved by the local Institutional Review Board. Written informed consent was obtained from all participants included in the study. via tablet PCs.Data collected preoperatively included sociodemographic information, clinically diagnosed comorbidities , and the use of past or current psychotherapy. For the psychometric assessment of psychological variables, the following validated (with the exception of the “motivation to lose weight” survey) self-rating questionnaires were employedStress was assessed using the Perceived Stress Questionnaire (PSQ) [α = 0.94.re (PSQ)  in its rre (PSQ) . The insDepression was assessed using the German version of the Patient Health Questionnaire Depression Scale (PHQ-9) [α = 0.87, and the level of agreement between the clinical diagnosis of depression according to ICD-10 and the PHQ-9 score (cut-off ≥ 10) was moderate (Cohen's k = 0.45); specificity was 84%, and sensitivity was 64%. (PHQ-9) , 38. PHQAnxiety was assessed using the German version of the Generalized Anxiety Disorder 7-item (GAD-7) scale [α = 0.88.7) scale , 40. GADMental impairment was assessed using the ICD-10 Symptom Rating (ISR) [α = 0.92.ng (ISR) . This 29Coping style was assessed using the German version of the Brief COPE questionnaire [α = 0.38–0.85.ionnaire , 43. PatMotivation to lose weight was assessed with specific ad hoc questions. A motivation survey was developed with 10 items asking how strongly patients were “self-motivated” to lose weight or motivated by their “social environment”  or “treatment environment” . Patients indicated their answers on a 5-point scale that ranged from 1 = not at all to 5 = very strong. The levels of internal consistencies were indicated by Cronbach's α = 0.79–0.84.2) were extracted from the medical database. On average, 20 months after surgery, weight status was either extracted from the medical database or self-reported, and weight loss was assessed. As a standard for evaluation, weight change is reported as the percentage of excess weight loss (%EWL), which is calculated using the following formula: (postoperative weight loss)/(preoperative excess weight at time of surgery) × 100. BMI ≥ 25 kg/m2 is recognized as the lowest limit of overweight, and therefore excess weight is calculated in relation to a BMI of 25 kg/m2 [Body weight (kg), height (cm), and BMI  with respect to group differences in preoperative (1) psychological burden, (2) coping style, and (3) motivation to lose weight (= dependent variables). An exploratory principal component analysis showed high correlations among “perceived stress” (PSQ-20), “depression” (PHQ-9), “anxiety” (GAD-7), and “mental impairment” (ISR) (r = 0.78–0.89), and strong loadings (>0.90) of each of these variables on a single factor explained 87% of the total variance in the data set. Therefore, in accordance with the constructs “coping style” and “motivation to lose weight,” the broader construct “psychological burden” was generated and used for further multivariate analyses to retain statistical power. For the combined dependent variable “psychological burden,” four variables were used: “perceived stress” (PSQ-20), “depression” (PHQ-9), “anxiety” (GAD-7), and “mental impairment” (ISR). For the combined dependent variable “coping style,” four variables of Brief COPE were used: “avoidant coping,” “seeking support,” “positive reframing,” and “active coping.” For the combined dependent variable “motivation to lose weight,” three variables were used: “self-motivation,” “social environment,” and “treatment environment.” For additional exploratory analyses, univariate ANOVAs were performed with Bonferroni-adjusted post hoc comparisons using Tukey's HSD test to control for Type I error. Chi-square tests were conducted for the nominal dependent variables, with post hoc comparisons using standardized residuals and a critical value of ±1.96 indicating significant group differences. The magnitude of group differences was further analyzed by means of effect sizes; for metrical data, we used Cohen's d , and for nominal data, we used Cohen's w . For correlative analyses, Pearson's product-moment correlation coefficient r was used. Preliminary analyses included describing the variables, screening for missing values and outliers, and testing for the normality, linearity, homogeneity of variance/variance-covariance matrices, and multicollinearity of the dependent variables, with no serious violations noted.All analyses were conducted using SPSS version 22 , and the alpha level of statistical significance was set at n = 21, EWL range: 14–39%), moderate , and high . Postoperative %EWL differed significantly between the three %EWL groups (p < 0.001). The results showed no significant group difference in the follow-up time interval (p > 0.05) . In relation to the observed %EWL distribution in our study, patients were classified into three %EWL groups of nearly equal size based on tertiles: low ( > 0.05) .p < 0.001). Significant preoperative group differences between patients of moderate and high %EWL were detected: patients with high %EWL had a lower preoperative weight (p = 0.018), excess weight (p = 0.012), and BMI (p = 0.015). Preoperative weight and BMI differences between the low-%EWL and moderate-%EWL groups and between the low-%EWL and high-%EWL groups did not reach statistical significance (p > 0.05). In the high-%EWL group, 38% (n = 8) of the patients attained a postoperative nonobese BMI of <30 kg/m2.Weight and BMI characteristics for the low-%EWL, moderate-%EWL, and high-%EWL groups before and after bariatric surgery are presented in p > 0.05). A small but statistically significant group difference in education levels was found: on average, the high-%EWL group had 2.2 more years of education than the low-%EWL group (p = 0.010). There were no statistically significant differences in education levels between the low-%EWL and moderate-%EWL groups or between the moderate-%EWL and high-%EWL groups (p > 0.05).Preoperative sociodemographic characteristics for the groups with low, moderate, and high %EWL are presented in p > 0.05). With regard to mental comorbidities, the groups with low and high %EWL presented with particularly high rates of presurgically diagnosed mental disorders ; however, the results showed no statistically significant group differences in the prevalence of mental disorders (p > 0.05) (p > 0.05) (n = 25) of the patients reported past or current psychotherapeutic treatment. The results showed a statistically significant group difference in the use of psychotherapy: slightly more patients in the low-%EWL group underwent psychotherapeutic treatment before LSG surgery compared with those in the moderate-%EWL group (p = 0.013). For the high-%EWL group, no statistically significant difference in the use of psychotherapy could be identified (p > 0.05)  . All thr > 0.05) . Altoget > 0.05) .F = 0.9, p = 0.535; Pillai's trace = 0.1; d = 0.5). Furthermore, the three %EWL groups showed no statistically significant difference in their overall preoperative motivation to lose weight in terms of “self-motivation” or motivation from the “social environment” or “treatment environment”  = 1.0, p = 0.428; Pillai's trace = 0.1; d = 0.4).Preoperative psychological characteristics for the groups with low, moderate, and high %EWL are presented in F = 1.6, p = 0.145; Pillai's trace = 0.2; d = 0.7). To further investigate our main research question (2), exploratory post hoc analyses were conducted. When the results for the different coping styles  were considered separately in Bonferroni-adjusted comparisons, a marked difference could be identified: patients with high postoperative %EWL reported slightly but significantly higher scores for “active coping” behavior prior to surgery compared with patients with low %EWL (p = 0.019) and moderate %EWL (p = 0.022). The low-%EWL and moderate-%EWL groups did not differ in this regard (p > 0.05) .In terms of the overall preoperative coping style (Brief COPE), the three %EWL groups showed no statistically significant difference ( > 0.05) . The posN = 28) and patients who were reassessed after LSG (N = 64) in preoperative variables such as age, sex, BMI, years of education, prevalence of metabolic syndrome symptoms, mental disorders including eating disorders, or psychotherapy use (p > 0.05). Furthermore, the two groups showed no statistically significant differences in their overall preoperative psychological burden  = 1.9, p > 0.05; Pillai's trace = 0.08; d = 0.6), coping style  = 1.0, p > 0.05; Pillai's trace = 0.04; d = 0.4), or motivation to lose weight  = 2.1, p > 0.05; Pillai's trace = 0.07; d = 0.5) (data not shown).The results showed no statistically significant differences between patients who dropped out of the study  tend to stagnate earlier in their weight loss process and begin to regain weight as early as 12 months after surgery [Our results showed that LSG patients with high %EWL had a lower weight and BMI prior to surgery compared with patients with moderate %EWL, but no significant weight and BMI differences compared with patients with low %EWL were found. This initially counterintuitive finding is in line with previous studies of gastric bypass surgery reporting that patients who were preoperatively less obese have a higher postoperative %EWL , 44, 45. surgery . AlthougThe preoperative psychological burden and motivation to lose weight were not associated with the amount of %EWL in patients following LSG intervention. However, more self-reported “active coping” behavior  seemed to be beneficial for treatment success in terms of more favorable weight loss outcomes. In summary, a more “active coping” style might be predictive of higher postoperative weight loss outcomes. However, the results provided no clear evidence of a clinically important role of preoperative mental health and the motivation to lose weight in the weight-related treatment success of LSG.Previous studies have identified psychological predictors of weight loss outcomes after bariatric surgery; for example, some have observed less weight loss in patients with mood and anxiety disorders , 27. HowHowever, because bariatric surgery leads to a profound change in patients' gastrointestinal and whole-body physiology, it is important to also consider potential physiological mechanisms . In factThe strength of our study is that it was conducted in a naturalistic clinical setting. In addition, whereas the majority of previous research in this area focused on Roux-en-Y gastric bypass surgery, we had access to a homogeneous clinical sample of patients undergoing LSG. To the best of our knowledge, this was the first study that systematically assessed the relationship between postoperative %EWL on average 20 months after surgery and the preoperative psychological burden , coping style, and motivation to undergo surgery in a sample of LSG patients. The size of our sample was comparable to that of other small-scale studies in the field of bariatric surgery. Moreover, the attrition rate in our study was rather small, and our attrition analyses showed no significant differences in preoperative variables between patients who provided data at follow-up compared with those who did not. However, from a statistical perspective, the sample size is still small, allowing for only a few statistical tests in addition to a large set of exploratory analyses. The patients' postoperative weight was self-reported, which might undermine the validity of the %EWL classification scheme. However, self-reporting of weight is relatively common in obesity studies, and there is evidence that objectively measured and self-reported weights are not significantly different in bariatric surgery patients . FinallyAlthough an “active coping” style seemed to be of value in predicting more favorable postoperative weight loss outcomes, neither biomedical markers nor an extensive set of other psychological constructs assessed before surgery enabled a clinically relevant prediction of the weight-related treatment success of LSG. Our study highlights the need for further research on the psychological correlates of postbariatric health outcomes. Nevertheless, despite our finding of a considerably large degree of variation in postoperative weight loss in our sample of LSG patients, all three %EWL groups showed significant weight loss and decreases in body weight and BMI, including the low-%EWL group. Therefore, the bariatric intervention proved to be highly beneficial for the vast majority of patients, thus promoting positive health outcomes.


Hippocampal gamma rhythms increase during mnemonic operations  and may affect memory encoding by coordinating activity of neurons that code related information . Here, a hippocampal-dependent, object–place association task  was used in rats to investigate how slow and fast gamma rhythms in the hippocampus relate to encoding of memories for novel object–place associations. In novel object tasks, the degree of hippocampal dependence has been reported to vary depending on the type of novelty . Therefore, gamma activity was examined during three novelty conditions: a novel object presented in a location where a familiar object had been (NO), a familiar object presented in a location where no object had been (NL), and a novel object presented in a location where no object had been (NO+NL). The strongest and most consistent effects were observed for fast gamma rhythms during the NO+NL condition. Fast gamma power, CA3–CA1 phase synchrony, and phase-locking of place cell spikes increased during exploration of novel, compared to familiar, object–place associations. Additionally, place cell spiking during exploration of novel object–place pairings was increased when fast gamma rhythms were present. These results suggest that fast gamma rhythms promote encoding of memories for novel object–place associations. This study provides the first evidence that links fast gamma rhythms in the hippocampus to encoding of novel object–place associations in a behavioral task. The results also relate these effects to firing patterns in place cells that resemble stimulation patterns that are routinely used to induce long-term potentiation, the presumed synaptic substrate of memory formation.Gamma oscillations (∼25–100 Hz) are prominent in the entorhinal-hippocampal network and have been shown to appear during a variety of memory tasks in rats, monkeys, and humans . Gamma rhythms occur as two distinct variants that are thought to route different streams of information entering hippocampal subfield CA1 . Slow gamma (∼25–55 Hz) may facilitate transmission of inputs to CA1 from CA3, a hippocampal subfield thought to be important for memory retrieval . Fast gamma (∼60–100 Hz) may promote inputs from the medial entorhinal cortex (MEC) that transmit ongoing spatial information . Functional correlates of these gamma subtypes have been reported for CA1 place cells in the form of different spatial coding modes . The firing properties exhibited in each case were hypothesized to reflect cellular mechanisms of memory retrieval during slow gamma and memory encoding during fast gamma. However, if these neuronal coding modes are involved in memory function, then effects should also be evident during behaviors in which these mnemonic processes are explicitly demonstrated. In the present study, memory encoding and retrieval were examined at the behavioral level using an object–place association task. Slow and fast gamma activities were measured during periods of exploration of novel and familiar object–place pairings. Memory encoding presumably occurs during exploration of novel object–place pairings, and memory retrieval presumably occurs during exploration of familiar object–place pairings.In standard novel object exploration tasks, rats are presented with a novel object and a familiar object in the same environment and are allowed to freely explore each item. Rats have been shown to spend more time exploring novel objects compared with familiar objects , providing behavioral evidence that rats recognize one object as novel and the other as familiar. The ability to discriminate novel and familiar objects is impaired in rats with hippocampal lesions, but this deficit is variable and appears to depend on the specific type of novelty involved . When novelty involves only the identity of the object, some studies report no deficits in rats with hippocampal lesions , whereas other studies report variable deficits depending on the size of the lesion  or the length of delay between familiarization and novelty exposure . In contrast, when novelty involves changes in the location of an object, deficits are more reliably observed following hippocampal lesions .Due to the reported variability of hippocampal involvement in novel object exploration tasks, gamma activity was examined during three types of novelty: novel object identity (NO), novel object location (NL), and novel object identity in a novel object location NO+NL; B. When Guide for the Care and Use of Laboratory Animals under an IACUC-approved protocol, in accordance with the Society for Neuroscience’s Policies on the Use of Animals in Neuroscience Research.Ten male Long–Evans rats weighing approximately 350–500 g were used in the study. Rats were housed on a reverse light/dark cycle (lights off from 8:00 A.M. to 8:00 P.M.) and tested during the dark phase. After drive implantation, rats were housed individually in cages (40 × 40 × 40 cm) constructed from clear acrylic and containing enrichment materials . Rats recovered from surgery for at least 1 week prior to the start of behavioral testing. All experiments were conducted according to the guidelines of the United States National Institutes of Health Recording drives contained 14  or 26  independently movable tetrodes. Tetrodes were constructed from 17 μm polyimide-coated platinum-iridium (90–10%) wire . Electrode tips in tetrodes targeted toward cell body layers were plated with platinum to reduce single channel impedances to ∼150–300 kΩ at 1 kHz.Recording drives were surgically implanted above the right hippocampus on the day of surgery. Stereotaxic coordinates were as follows (in mm): 3.8 AP, 3.0 ML, 1.0 DV in nine rats and 5.0 AP, 5.0 ML, 1.0 DV in one rat. In the latter rat, only those tetrodes that were histologically verified to be in dorsal hippocampus were used . Bone screws were placed in the skull, and the screws and the base of the drive were covered with dental cement to affix the drive to the skull. Two screws in the skull were connected to the recording drive ground.Over the course of a few weeks after drive implantation, tetrodes were slowly lowered toward their target locations. In six of the rats implanted with hyperdrives, six tetrodes were targeted toward the CA1 cell body layer and six toward the CA3 cell body layer. In the other four rats, all of the 12 or 24 recording tetrodes were targeted toward the CA1 cell body layer. In each rat, one tetrode was targeted toward the apical dendritic layers of CA1. Another tetrode was used as a reference for differential recording and was placed at the level of the corpus callosum or higher; the reference tetrode was recorded against ground to make sure that it was placed in a quiet location. All recording locations were verified histologically after experiments were finished (see Histology). Representative examples of final recording locations are shown in Data were collected using the Neuralynx data acquisition system. The headstage output of recording drives was conducted via lightweight tether cables through a multichannel slip-ring commutator to a data acquisition system that processed the signals through individual 24 bit AD converters . Unit activity was bandpass filtered from 600 to 6000 Hz, and spike waveforms were time-stamped and recorded at 32 kHz for 1 ms. Local field potentials (LFPs) were recorded continuously in the 0.1–500 Hz band at a sampling rate of 2000 Hz. Notch filters were not used. Continuously sampled LFPs were recorded differentially against a common reference electrode placed in an electrically quiet region (see Surgery and tetrode placement). Light-emitting diodes on the headstages were used to track rats’ movements at a 30 Hz sampling rate.On each day of the experiment, animals were allowed to freely explore an open-field environment (60 × 60 cm box) for three 10 min behavioral sessions, alternated with 10 min rest sessions in a towel-lined flower pot B. The oThe total time during which a rat’s head was within 15 cm of the center of each object during the first 3 min of the 10 min novelty session was determined and used to calculate the discrimination index (DI) between novel and familiar object–place associations [ie, (novel time)/(novel time + familiar time); For LFP recording analyses and place cell phase-locking analyses, measures were computed only within time windows when animals were actively exploring an object see–5, 7. Ac–A stricter criterion for definition of object exploration was also used for a subset of analyses . Then, the absolute power for each frequency was Z-scored across time for the LFP recording from each tetrode, in order to allow for comparisons across different frequencies that would otherwise be difficult due to the 1/f decay of power in physiological signals. Running speed was calculated (see Estimation of running speed) and averaged within each 200 ms time window corresponding to the LFP segments. To produce power estimates across running speed bins, Z-scored absolute power at each frequency was averaged across all time windows that fell within a given speed bin and smoothed with a Gaussian kernel centered on that bin. Speed and frequency were plotted on a log-log scale for gamma frequencies . For each object exploration window, a single theta, slow gamma, and fast gamma phase synchrony measure was found by averaging phase synchrony estimates across time, across frequencies within each respective frequency range , and lastly across all CA1 and CA3 recording pairs associated with the object exploration window.n = 98 CA1 cells and 33 CA3 cells) with a place field peak firing position located less than 20 cm away from the center of either object were identified and included in this study. Only the spike times occurring in locations within 15 cm of the center of either object during object exploration time windows (described in Detection of object exploration periods) were included in this analysis. The time-varying phases for theta, slow gamma, and fast gamma were determined using the Hilbert transform of the bandpass filtered signal for each respective frequency range. The theta, slow gamma, and fast gamma spike phase distributions for each cell were then determined by identifying the theta, slow gamma, and fast gamma phases, respectively, at the EEG time point closest to each spike time. Phase-locking was quantified using the mean vector length of the resulting phase distributions  in which the firing rate was greater than 0.4 × the peak firing rate across the whole session. For each place field, a peak firing position was determined . A total of 131 place cells . Generalized linear mixed models were used to test for effects of novelty condition  and data type (objects vs no objects) on the discrimination index behavioral measure C, with nditions D. Genernditions –6, 7. Pama power  and phasma power  measuresma power . t Testsl spikes . Sign te periods . Paired ta power D,E. t Tl spikes G. For tl spikes F, a repFor verification of tetrode locations, brains were cut coronally into 30 μm sections and stained with cresyl violet A. All tF = 4.6, p = 0.04a; main effect of novelty condition, F = 5.3, p = 0.03a; main effect of data type, F = 9.4, p = 0.003a, generalized linear mixed models, n = 10 rats in F, NO+NL, and NO conditions, and n = 8 rats in NL condition; superscript letters following p values correspond to statistics presented in F = 4.2, p = 0.02a, repeated-measures ANOVA); however, no effect of novelty condition was found on DI values in control No objects sessions  = 1.8, p = 0.2a, repeated-measures ANOVA). Periods of exploration of novel object–place pairings in Session 2 of NO+NL, NL, and NO were compared to exploration of familiar object–place pairings in Session 2 of familiarization and re-familiarization days (F). Rats explored novel objects in new locations more than they explored familiar objects during familiarization and re-familiarization days . Rats also explored novel objects in new locations more than they explored novel objects in familiar locations  and more than they explored the same locations in sessions in which no objects were presented  = 4.8, p = 0.001a, paired t test). Significant novelty effects on behavior were not obtained for the NL condition or NO condition, however . The lack of significant novelty effects on behavior for the NL and NO conditions was not explained by lower levels of familiar exploration during Session 1 in the NL and NO conditions compared to the NO+NL condition  = 1.2, p = 0.3b; repeated-measures ANOVA). It is thus possible that the NO and NL conditions were insufficiently novel for piquing rats’ curiosity, especially in the NO condition given that novel and familiar objects were constructed from the same materials .Continuously sampled LFP recordings and place cell spike trains were obtained from strata pyramidale of hippocampal subfields CA1 and CA3 of six rats and CA1 of an additional four rats A perforF = 2.6, p = 0.1c, generalized linear mixed models), and thus measures were averaged across running speeds in subsequent analyses. A significant interaction of hippocampal subregion, novelty condition, object–place pairing type, and gamma type  = 4.0, p = 0.05c, generalized linear mixed models, n = 10 rats in F, NO+NL, and NO conditions and n = 8 rats in the NL condition), and significant main effects of object–place pairing type  = 7.5, p = 0.007c) and gamma type  = 4.0, p = 0.05c), were obtained. Recordings from CA1  = 12.0, p = 0.001d, generalized linear mixed models, n = 10 rats in F, NO+NL, and NO conditions, and n = 8 rats in NL condition). For the NO+NL condition, CA1 power in the fast but not slow gamma range increased across a broad range of running speeds when animals explored novel objects in novel locations . This effect was not observed for exploration of the familiar object in the NO+NL session . This indicates that fast gamma power increased selectively during exploration of the novel object in the NO+NL session. Corresponding effects were not observed for slow gamma . Accordingly, there was a significant interaction between gamma type and object type  on gamma power increases during the NO+NL session relative to F sessions  = 7.0, p = 0.01d, generalized linear mixed models), indicating that slow and fast gamma power changed differently during exploration of novel object–place pairings. Relative to fast gamma power in F sessions, fast gamma power during novel object exploration in NO+NL increased significantly more than fast gamma power during familiar object exploration in NO+NL . Analogous results were not observed for slow gamma . These results indicate that fast, but not slow, gamma was enhanced during novel, but not familiar, object exploration in the NO+NL session. The same pattern of results was observed when a stricter criterion was used to define object exploration .Whether CA1 gamma power during object exploration changed between novel and familiar sessions depended on which novelty condition was assessed, whether the object–place pairing was familiar or novel, and which type of gamma was measured . However, gamma power changes during exploration of the novel object–place pairing in NL were not significantly different than gamma power changes during exploration of the familiar object–place pairing in NL  = 6.1, p = 0.02d; fast gamma, p = 0.2d; slow gamma, p = 0.2d; generalized linear mixed models, n = 8 rats). Moreover, significant behavioral effects were not observed in the NL condition  in NL relative to fast gamma power during familiar object exploration in F . Also, there was no significant object type × gamma type interaction and no significant main effects on power changes during novel object exploration compared to familiar object exploration in the NO session  = 0.05, p = 0.8d; main effect of object type: F = 1.4, p = 0.2d; main effect of gamma type: F = 3.2, p = 0.08d; generalized linear mixed models, n = 10 rats). It should be noted that significant behavioral effects were not observed in the NO condition . Accordingly, CA3 gamma power during object exploration was not found to significantly change between novel and familiar conditions, regardless of novelty condition, object–place pairing type, and gamma type  = 1.1, p = 0.3e; novelty condition: F = 0.3, p = 0.6e; object–place pairing type, F = 1.0, p = 0.3e; gamma type: F = 0.5, p = 0.5e; generalized linear mixed models). Also, unlike for CA1, CA3 fast gamma power in the NO+NL session did not increase more, relative to CA3 fast gamma power in F sessions, during exploration of novel object–place pairings compared to familiar object place pairings . However, it is possible that CA3 effects were not detected due to the lower number of CA3 recordings compared with CA1 recordings .CA3 has been proposed to be critical for associative memory . Thus, encoding or retrieval of associations between objects and locations may involve CA3. However, no significant slow nor fast gamma power changes were found in CA3 in any of the novel conditions during exploration of novel object–place pairings relative to exploration of familiar object–place pairings in familiar conditions  = 11.0, p = 0.002f, generalized linear mixed models, n = 6 rats in F, NO+NL, and NO conditions, and n = 4 rats in NL condition). In the NO+NL session, the increase in fast gamma coupling during novel object exploration relative to familiar object exploration was significantly greater than the corresponding change in slow gamma coupling (t(5) = 4.3, p = 0.008f, paired t test, n = 6 rats). In the NL and NO sessions, no significant differences were observed between slow and fast gamma for phase synchrony measures (t(3) = 0.4, p = 0.7f, n = 4 rats; NO: t(5) = 1.7, p = 0.1f, n = 6 rats; paired t test). These results raise the possibility that enhanced fast gamma coupling between CA3 and CA1 facilitates encoding of memories of associations between novel objects and locations in which objects have not appeared previously.It may also be possible, though, that analogous effects in CA3 were not detected because of the nature of LFP signals in CA3. The curve of the cell body layer in CA3 may cause currents to flow in different directions. This may prevent currents from summing together nicely to generate a large and easily detectable LFP. Thus, slow and fast gamma coupling between CA1 and CA3 was also examined by estimating phase synchrony, which measures the consistency of phase differences between two signals and thus is potentially less affected by low amplitude LFPs . CA3–CA1n = 98 CA1 cells and 33 CA3 cells; F = 4.0, p = 0.05g, generalized linear mixed models). In the NO+NL condition, a significant interaction was found between gamma type and place cell type on the mean vector length of gamma phase distributions  = 4.8, p = 0.03g, generalized linear mixed models, n = 16 cells with fields close to novel Object C and n = 41 cells with fields close to familiar Object A), indicating that phase-locking to fast gamma was more strongly affected by the presence of novelty than was phase-locking to slow gamma. Accordingly, spikes of cells near the novel object were significantly more phase-locked to fast gamma than were spikes of cells near the familiar object . Analogous phase-locking effects were not observed for slow gamma , nor were significant effects obtained across the other novelty conditions  = 4.1, p = 0.05g for gamma type × place cell type interaction, p = 0.2g, post hoc for fast gamma, p = 0.4g, post hoc for slow gamma; NO: F = 0.4, p = 0.5g for gamma type × place cell type interaction, p = 0.05g, post hoc for fast gamma, p = 0.2g, post hoc for slow gamma; generalized linear mixed models). These findings suggest that fast gamma rhythms may coordinate ensembles of place cells that signal object novelty and code spatial information for locations where objects were not previously found.It is possible that enhanced fast gamma coupling in the hippocampal network during exploration of novel object–place associations coordinates ensembles of place cells that encode information about the location and the objects. If so, then place cell spiking should be more strongly modulated by fast gamma rhythms during exploration of novel object–place associations. To investigate this possibility, phase-locking of CA3 and CA1 place cell spikes to slow and fast gamma in CA1 was assessed in the subset of place cells that coded locations close to novel or familiar object–place pairings  = 4.6, p = 0.04h, generalized linear mixed models, n = 33 place cells with fields close to novel objects and n = 44 place cells with fields close to familiar objects). In the NO+NL condition, the in-field firing rates of CA1 place cells with fields near novel Object C, but not familiar Object A, were significantly higher during fast gamma periods than during slow gamma periods  = 4.5, p = 0.04h, generalized linear mixed models; cells with field near Object C, p = 0.04h; cells with field near Object A, p = 0.2h, sign test; n = 18 cells with fields near familiar Object A and n = 12 cells with fields near novel Object C). This result was not explained by effects of spiking on local fast gamma power because comparable findings were observed when place cell rate maps were constructed for slow and fast gamma episodes detected using non-local tetrodes . Similar findings were observed for object–place associations in the NL condition  = 33.5, p < 0.001h, generalized linear mixed models; place cells with field near Object A′, p = 0.008h; cells with field near Object A, p = 0.09h, sign test; n = 13 cells with fields close to A and n = 8 cells with fields close to A′) but not in the NO condition  = 1.3, p = 0.3h; main effect of cell type: F = 0.02, p = 0.9h; main effect of gamma type: F = 0.005, p = 0.9h; generalized linear mixed models; n = 13 cells with fields close to A and n = 13 cells with fields close to B).The phase-locking of place cell spikes to fast gamma rhythms during encoding of novel object–place pairings may also be associated with differences in firing rates. CA1 place cell in-field firing rates were significantly different, depending on which type of gamma was present and whether a cell’s field was located near a novel or familiar object–place pairing (j or theta phase synchronyk between CA3 and CA1 were observed during exploration of novel-object place associations (j). Still, the effects of novelty on theta power were rather variable , raising the possibility that these effects simply reflect changes in theta power. However, no significant changes in theta powerciations A-F. Sigciations j. Stillvariable D,E, andF = 3.1, p = 0.05l, two-way ANOVA). In the NO+NL condition, spikes of place cells with fields near the novel object were significantly more phase-locked to theta than were spikes of cells with fields near the familiar object (t(55) = 2.2, p = 0.03l, Student’s t test). Analogous phase-locking effects were not observed across the other novelty conditions (NL: t(50) = 0.8, p = 0.4l; NO: t(66) = 1.0, p = 0.3l, Student’s t test). Taken together with the fast gamma results reported above  interaction effect on mean vector length of theta phase distributions of place cell spikes , the only significant effects that were observed were increases in fast gamma power during exploration of the novel object–place pairing and place cell spiking near the novel object–place pairing during fast gamma periods. Thus, it is possible that fast gamma also enhanced encoding of novel object–location pairings when novelty only entailed a change in object location. However, this type of novelty was likely not as striking as novelty involving both object identity and location changes, considering that significant behavioral effects of novelty were not observed in the NL condition C. It isNo fast or slow gamma-related effects were observed when a familiar object was replaced by a novel object in the same location (NO). It is possible that the saliency of the novel experience is relatively low in this type of paradigm, in which only the object identity changes, compared to a paradigm in which novel objects are presented in changed locations . In accord with this assumption, animals did not explore novel objects significantly more than familiar objects in the NO condition in the present study C. In thWith regard to fast gamma, the effects observed during the NO+NL condition are consistent with the notion that fast gamma plays a role in encoding of novel experiences. Previous studies have suggested that fast gamma is important for transmitting positional information from MEC to the hippocampus. Place cells in area CA1 preferentially code “place-based” representations of space during fast gamma , and ensembles of CA1 place cells more closely represent an animal’s location in real-time during fast gamma . Such communication about current spatial experience during fast gamma may complement a broader role of fast gamma in encoding memories of novel experiences.The hypothesis that fast gamma rhythms are important for novelty encoding is also supported by results from earlier studies. Enhanced fast gamma power has been observed in area CA1 in rats during exploration of a novel maze . A study in monkeys revealed an increase in coherence between hippocampal spikes and fast gamma rhythms during successful encoding of novel images . In humans, an increase in higher frequency, but not lower frequency, gamma was observed in the hippocampus during successful encoding of words in a free recall task . The present results may similarly indicate formation of a novel associative memory during enhanced fast gamma activity, when transmission of novel sensory information to the hippocampus from MEC is likely to be strongest.We did not find evidence for increases in Beta2 oscillations . It is unclear why increased slow gamma phase-locking occurred in a novel environment in the study by Kitanishi et al. (2015), whereas increased fast gamma phase-locking occurred during presentation of novel object–place pairings in a familiar environment in the present study. Additional investigations are required to determine why slow gamma plays a role in encoding completely novel environments but not novel object–place pairings in a familiar setting.A surprising finding in the present study was that fast gamma coupling between CA3 and CA1 was stronger than slow gamma coupling during exploration of novel object–place pairs A. This The present study also found that place cell spiking was higher during fast gamma than during slow gamma when rats explored novel object–place pairings . This efNovelty exploration was also associated with increases in fast gamma phase-locking of CA3 and CA1 place cell spikes . Fast ga


Triticum aestivum L.), one of the world’s most important food crops, is a strictly autogamous (self-pollinating) species with exclusively perfect flowers. Male sterility induced by chemical hybridizing agents has increasingly attracted attention as a tool for hybrid seed production in wheat; however, the molecular mechanisms of male sterility induced by the agent SQ-1 remain poorly understood due to limited whole transcriptome data. Therefore, a comparative analysis of wheat anther transcriptomes for male fertile wheat and SQ-1–induced male sterile wheat was carried out using next-generation sequencing technology. In all, 42,634,123 sequence reads were generated and were assembled into 82,356 high-quality unigenes with an average length of 724 bp. Of these, 1,088 unigenes were significantly differentially expressed in the fertile and sterile wheat anthers, including 643 up-regulated unigenes and 445 down-regulated unigenes. The differentially expressed unigenes with functional annotations were mapped onto 60 pathways using the Kyoto Encyclopedia of Genes and Genomes database. They were mainly involved in coding for the components of ribosomes, photosynthesis, respiration, purine and pyrimidine metabolism, amino acid metabolism, glutathione metabolism, RNA transport and signal transduction, reactive oxygen species metabolism, mRNA surveillance pathways, protein processing in the endoplasmic reticulum, protein export, and ubiquitin-mediated proteolysis. This study is the first to provide a systematic overview comparing wheat anther transcriptomes of male fertile wheat with those of SQ-1–induced male sterile wheat and is a valuable source of data for future research in SQ-1–induced wheat male sterility.Wheat ( Triticum aestivum L.) is one of the most important cultivated crops in the global food system; it supplies the needed food protein and calories for over 35% of the world’s population and is the principal food in more than 40 countries [Wheat (ountries –4. Aboutountries . The mosountries .Heterosis (or hybrid vigor) is a universal phenomenon in biology, and hybrid offspring often display increased yield, higher total biomass, and improved abiotic and biotic stress resistance compared with their homozygous parents –12. HybrSQ-1, an ideal chemical hybridization agent, is a systemic product that is efficiently transported in wheat from leaves to flowers . It is ede novo transcriptome sequencing for non-model organisms [Arabidopsis thaliana [Brassica rapa [In recent years, next-generation sequencing (NGS) techniques  have rapidly developed and have provided efficient, powerful, and inexpensive tools for advanced research in many fields, including genome re-sequencing, small RNA sequencing, Degradome sequencing, and rganisms –38. RNA thaliana , Brassicica rapa , chili pica rapa , sweet oica rapa , radish ica rapa , rice [4ica rapa , and corica rapa .The aim of this study was to seek out the key genes and important metabolic pathways associated with wheat PMS. It is the first study to characterize the complete transcriptome of wheat PMS using high-throughput Illumina HiSeq 2500 sequencing technology, and it provides new insights into wheat male sterility induced by SQ-1.-1 SQ-1 was evenly sprayed on the leaves of wheat at the 8.5 stage of the Feekes scale.SQ-1 was obtained from the Key Laboratory of Crop Heterosis of Shaanxi Province and used to produce hybrid wheat seed. It is a pyridazinecarboxylic acid compound that is readily soluble in water. Stamen sterility  was induced without affecting pistil fertility when 5.0 kg ha-1 SQ-1 was evenly sprayed on the leaves of the wheat in the treatment group, and the same amount of water was sprayed on the leaves of the wheat in the control group. At the uninucleate pollen stage of wheat, anthers were separately collected from wheat fertility spikes and SQ-1–induced male sterile spikes, frozen immediately in liquid nitrogen, and stored at -80°C until use.The wheat cultivar Xi-nong 1376 was sown in the experimental field of Northwest A & F University, Yangling, Shaanxi, China  on October 7, 2013. The experimental plot contained about 9,000 plants grown in 180 rows (1.5 m long each) at a density of 25 cm space between rows and 3 cm between plants with a row, it was divided into two groups (treatment group and control group) and each group contained 90 rows. When the average growth stage of the wheat reached the 8.5 stage of the Feekes scale, 5.0 kg haEach frozen anther was ground in a mortar with liquid nitrogen, after which total RNA was extracted using Trizol Reagent  according to the standard protocol. The quality of total RNA was checked by electrophoresis in a 1.5% agarose gel and the concentration of total RNA was determined by NanoDrop . The RNA integrity value was further verified using an Agilent 2100 Bioanalyzer . The two cDNA libraries of fertile wheat and SQ-1–induced male sterile wheat were prepared according to the manufacturer’s instructions for mRNA-Seq sample preparation . The cDNA library products were sequenced by Illumina paired-end sequencing technology with read lengths of 100 bp, and they were sequenced on the Illumina HiSeq 2500 instrument by Biomarker Technologies Co. Ltd. . The dataset was submitted to the NIH Short Read Archive (accession number: SRP051670).Before assembly, the raw paired-end reads were filtered to obtain high-quality clean reads. Low-quality sequences were removed, including sequences with ambiguous bases (denoted with an “N” in the sequence trace) and reads with more than 20% low-quality bases . After purity filtering was completed, the high-quality reads were assembled by Trinity (release 20131110) with default parameters to construct unique consensus sequences .http://telethon.bio.unipd.it/bioinfo/IDEG6/). The false discovery rate (FDR) method was introduced to determine the threshold p-value at FDR ˂ 0.01, and the absolute value of log2Ratio ≥ 1 was used as the threshold to determine the significance of the differential expression of unigenes.The gene expression level was measured by the values of RPKM (reads per kb per million reads) . Unigenehttp://www.uniprot.org/) [http://www.ncbi.nlm.nih.gov/COG/) [http://www.genome.jp/kegg/) [http://www.geneontology.org/) [N represents the total number of unigenes with GO and KEGG pathway annotation; n represents the number of differentially abundant unigenes in N; M represents the number of unigenes that were annotated to certain GO or KO terms; and m represents the number of differentially abundant unigenes in M. The initial p-values were then adjusted using a Bonferroni Correction and a corrected p-value of 0.05 was adopted as a threshold.In order to perform functional annotation, the assembled unigenes were submitted to public databases and compared with the NCBI non-redundant protein database (NR) , the Swiot.org/) , the Cluov/COG/) , and thep/kegg/)  using Blgy.org/) . All difA total of 42,634,123 paired-end sequence reads  remained, with the Q30 percentage over 92%. The high-quality reads were aligned to assembled unigenes, with more than 80% of the high-quality reads mapping to a unique or to multiple unigene locations . TrinityWe used GetORF software from the EMBOSS (v. 6.0.1) analysis package to perform open reading frame (ORF) predictions. A total of 81,838 ORFs were detected from 82,356 unigenes, with an average length of 402 bp and an N50 length of 846 bp. The majority of ORFs (68.31%) contained 0–300 bp, whereas 17,566 ORFs (21.46%) contained 300–1000 bp and 8,369 ORFs (10.23%) exceeded 1,000 bp . The remA sequence similarity search was conducted against several public databases: the NR database, the Swiss-Prot database, the COG database, the GO database, and the KEGG database, using Blast with an E-value of less than 1e-5. A total of 967 differentially expressed unigenes (88.88%) exhibited gene annotation .To identify differentially expressed unigenes between fertile wheat and SQ-1–induced male sterile wheat, putative differentially expressed unigenes were identified on the basis of RPKM values that were calculated from the read counts mapped onto the reference transcriptome. A total of 1,088 unigenes were differentially expressed between fertile wheat and SQ-1–induced male sterile wheat according to a comparison of expression levels with Fold Change (FC) ≥ 2 and FDR ˂ 0.01 . Using tGO is an international classification system for standardized gene functions that may be obtained from the NR annotation information. The GO terms consist of the following three broad categories: cellular components, molecular functions, and biological processes. GO assignment was used to assign 637 differentially expressed unigenes to 57 subcategories. Among them, 394 differentially expressed unigenes were related to cellular components, 487 differentially expressed unigenes were grouped under molecular functions, and 492 differentially expressed unigenes were involved in biological processes. Within the cellular component category, the majority of differentially expressed unigenes were enriched in the subcategories of “cell part,” “cell,” and “organelle.” Within the biological process category, the great majority were related to “metabolic process,” “cellular process,” and “response to stimulus.” Within the molecular function category, the largest proportion of differentially expressed unigenes were involved in “binding” and “catalytic activity,” and a relatively large number were related to “transporter activity” , which mAll the differentially expressed unigenes were aligned to a search against the COG database for functional prediction and classification, and 333 unigenes were assigned to 25 COG functional categories. Among these categories, the cluster of “general function prediction only” was the largest group (22.22%), followed by “translation, ribosomal structure and biogenesis” (13.51%), “carbohydrate transport and metabolism” (12.31%), “posttranslational modification, protein turnover and chaperones” (12.01%), and “amino acid transport and metabolism” (11.41%). The smallest groups were “nucleotide transport and metabolism” (1.20%), “cytoskeleton” (0.90%), and “RNA processing and modification” (0.30%) .Functional classification and pathway assignment were performed using BlastX with an E-value of less than 1e-5. A total of 156 differentially expressed unigenes were annotated, and 126 were mapped onto 60 pathways in the KEGG database. The pathways containing the most differentially expressed unigenes were involved in “ribosome” (20 unigenes), followed by “photosynthesis” (8 unigenes), “fatty acid biosynthesis” (7 unigenes), “nitrogen metabolism” (7 unigenes), “arginine and proline metabolism” (6 unigenes), “biosynthesis of unsaturated fatty acids” (6 unigenes), “carbon fixation in photosynthetic organisms” (6 unigenes), “glutathione metabolism” (6 unigenes), “glycolysis/gluconeogenesis” (6 unigenes), “RNA transport” (6 unigenes), “starch and sucrose metabolism” (5 unigenes), “galactose metabolism” (4 unigenes), “peroxisome” (4 unigenes), “oxidative phosphorylation” (3 unigenes), and “ubiquitin-mediated proteolysis” (3 unigenes) .In all, 72 down-regulated unigenes were assigned to 40 pathways. The pathways containing the largest number of down-regulated unigenes were involved in “ribosome” (18), a large and complex molecular machine that serves as the primary site of biological protein synthesis. RNA polymerase may be closely involved in fertility, and DNA-directed RNA polymerase II subunit RPB1 was down-regulated in SQ-1–induced male sterile wheat. In addition, 3 down-regulated unigenes were involved in “purine metabolism” and 2 down-regulated unigenes were related to “pyrimidine metabolism.”We identified 17 down-regulated unigenes related to photosynthesis, including “photosynthetic electron transport chains” (8), “carbon fixation in photosynthetic organisms” (6), and “photosynthesis-antenna proteins” (3). Previous studies showed that SQ-1–induced wheat male sterility was closely linked with plant respiration, and the short supply of energy ultimately led to wheat male sterility . Our res1O2), superoxide radical (O2–), and hydrogen peroxide (H2O2), which can lead to the significant destruction of cells [ROS are chemically reactive molecules containing oxygen, including hydroxyl radical (OH), lipid peroxide (ROO–), singlet oxygen , or anthropic action through xenobiotics, such as herbicides . Increasabortion . In our The proteasomal degradation pathway is essential for many cellular processes, including the regulation of gene expression, the cell cycle, and responses to oxidative stress, and it also participates in plant pollen abortion. From the early stage of mononucleate pollen to the initial stage of trinucleate pollen, the expression level of ubiquitin-related modifier 1 gene was significantly higher in PMS wheat anthers than in fertile anthers, and the F-box protein gene was significantly up-regulated in PMS wheat anthers from the late stage of mononucleate pollen to the initial stage of binonucleate pollen. These genes are the promoters for pollen abortion of wheat PMS induced by the chemical hybridization agent SQ-1 .Differential proteomic analysis of polyubiquitinated proteins associated with wheat male sterility identified 127 differentially expressed polyubiquitinated proteins, including heat shock protein 70, ATPase subunit, ubiquitin-related enzyme, glycosyltransferase, and 20S proteasome subunit. Proteins in PMS wheat anthers whose expression was up-regulated included those in the categories of protein metabolism, carbohydrate and energy metabolism, amino acid metabolism, and plant secondary metabolism. However, PMS wheat anthers expressed much lower levels of photosynthesis-related proteins. Alteration of polyubiquitinated proteins is associated with wheat male sterility .Our results showed that most of the up-regulated unigenes were closely associated with the categories of protein degradation and glutathione metabolism. The down-regulated unigenes were related to the categories of “photosynthesis” (17 unigenes) and “ribosome” (18 unigenes) . The dowThis is the first study to obtain a large-scale SQ-1–induced male sterile wheat anther transcriptome dataset using high-throughput Illumina sequencing technology. Using functional annotation and enrichment analysis, we analyzed differentially abundant unigenes between fertile wheat and SQ-1–induced male sterile wheat and established a simple mode of SQ-1–induced male sterility in wheat .After SQ-1 was sprayed on the leaves of wheat, it was transported from the leaves to the flowers. Its effects included down-regulated expression of numerous unigenes involved in “nucleotide metabolism,” “ribosome,” “aerobic respiration,” “photosynthesis,” and “ROS metabolism.” In addition, a large number of unigenes were up-regulated, including those that were closely associated with “anaerobic respiration,” “glutathione metabolism,” “abnormal RNA degradation,” and “protein degradation.” We identified a great number of important genes related to wheat male sterility and provided a framework for further mechanism studies on SQ-1–induced male sterility in wheat.


Socioeconomic inequalities as social determinants of health are important issues in public health and health promotion. However, the association between socioeconomic status and eating behaviors has been investigated poorly in Japanese adults. To fill this gap, the present study examines the association of eating behaviors with household income and education.The sample comprised 3,137 Japanese adults  aged 30 to 59 years who responded to an Internet-based cross-sectional survey in 2014. Data on the following eating behaviors were collected via self-report: “taking care of one’s diet for health,” “eating vegetables,” “frequency of eating breakfast,” “frequency of family breakfasts,” “frequency of family dinners,” “using the information on nutrition labels,” and “conversations with family or friends during meals.” Self-reported data on socioeconomic status (household income and education) and demographic variables  were also collected. The associations between eating behaviors and household income or education were tested using binomial logistic regression analysis with eating behaviors as dependent variables and household income and education as independent variables. A trend P -value was calculated for three categories of household income  and education .Higher household income and education were significantly associated with higher rates of eating vegetables, using the information on nutrition labels, and conversation with family or friends during meals in Japanese men and women. Higher household incomes were significantly associated with lower rates of frequency of family breakfasts in Japanese men and lower rates of frequency of family dinners in Japanese men and women.Higher socioeconomic status as indicated by household income or education was associated with eating more vegetables and conversation with family or friends during meals in Japanese men and women. Socioeconomic status should be considered in health promotion and diet improvement. Health disparities are important issues in public health and health promotion. Socioeconomic disparities in health have been widely reported in many Western countries. In several previous studies, individual socioeconomic status (SES), as measured by household income, education, or occupation, has been shown to be closely related to lifestyle, mortality, and morbidity , 2. In JThe World Health Organization (WHO)  identifiDietary behavior is affected by knowledge, skills, attitudes, food preparation factors and environment . NutritiWe hypothesized that people with higher SES would have healthier eating habits, which would in turn influence their general health, because higher SES permits adequate intake of healthy food and therefore good health. We used seven items chosen from the Japan National Health and Nutrition Survey (NHNS)  and prevAn Internet-based cross-sectional survey was conducted in February 2014 by a Japanese online research service company that holds data on approximately 160,300 adult registrants, including their sociodemographic attributes. This allows the company to target particular demographic groups as necessary. We aimed to have a sample of approximately 3,000 adults aged 30 to 59 years, with 500 men and 500 women from three age groups . We targeted adults aged 30 to 59 because we felt that both promotion of healthy eating and reduction in health disparities were particularly important in this group. The set sample size and attributes were stratified by distribution of the Japanese average age, the 2013 Population Census of Japan for sex , and theHousehold income and educational level were used to measure SES. Educational level was grouped into three categories: junior high/high school, 2-year college, and 4-year college/graduate school. We were unable to accurately ascertain individual-level equivalent incomes because the choices were for levels of household income: less than 3,000,000 JPY, 3,000,000–5,000,000 JPY, 5,000,000–7,000,000 JPY, 7,000,000–10,000,000 JPY, 10,000,000–15,000,000 JPY, and over 15,000,000 JPY. We therefore analyzed income at category level. Household income was classified into three equally distributed categories: less than 3,000,000 JPY, 3,000,000–7,000,000 JPY, and over 7,000,000 JPY.Seven items were used to measure eating behavior: taking care of one’s diet for health, eating vegetables , 21, freThe next questions were preceded by the phrase, “The following questions are about your normal lifestyle”. Respondents were asked, “How often do you usually eat breakfast?”, with five possible responses: (1) Every day; (2) 4 or 5 days/week; (3) 2 or 3 days/week; (4) 1 day/week; and (5) Never. Participants who answered (1) to this question were defined as those who eat breakfast regularly. Respondents were asked, “How often each week do you usually eat breakfast with all the members of your family?” and “How often each week do you usually eat dinner with all the members of your family?” The following five response choices were provided: (1) Every day; (2) 4 or 5 days/week; (3) 2 or 3 days/week; (4) 1 day/week; and (5) Never. People who answered (1) were defined as those who regularly eat meals with their families each question.The final questions were preceded by, “The following questions are about your normal eating habits”. Respondents were asked, “Do you use the information on nutrition labels or calorie information on store displays and menus?” Possible responses were: (1) Very often; (2) Often; (3) Not much; and (4) Never. Participants who answered (1) or (2) were defined as those who consult nutrition information. Respondents were asked, “Do you talk to your family and friends during meals about the meal or nutrition?” with four response choices: (1) Very often; (2) Often; (3) Not much; and (4) Never. People who answered (1) or (2) to this question were defined as having positive conversations during meals.The study asked about two health-risk behaviors: current smoking and alcohol consumption. Smoking habits were surveyed with the question, “How many cigarettes or cigars per day do you smoke?” Responses were: (1) I have never smoked; (2) I stopped smoking more than 1 year ago; (3) I stopped smoking less than 1 year ago; (4) I smoke 1–20 cigarettes or cigars per day (5) I smoke 21–40 cigarettes or cigars per day; and (6) I smoke more than 41 cigarettes or cigars per day. We categorized participants who responded (4) to (6) as current smokers. Alcohol consumption was surveyed by asking, “How many days per week do you consume alcohol?” Response choices were as follows: (1) Every day; (2) 5 or 6 days/week; (3) 3 or 4 days/week; (4) 1 or 2 days/week; (5) 1 to 3 days/month; (6) I stopped consuming alcohol more than 1 year ago; and (7) I hardly drink alcohol at all. We categorized responses (1) to (5) as current alcohol consumption.Demographic variables included sex, age, marital status, residence status, and employment status. Age was classified as 30–39, 40–49, and 50–59 years. Marital status was categorized as currently married or currently unmarried. Residence status was categorized as living with others or living alone. Employment status was categorized as employed or not employed.n = 52) or employment status  were not included in the analysis because these were important variables in this study. We considered responses of Other/Unknown to be missing values rather than lost data. Interpretation of the results would have been difficult if Other/Unknown responses were combined with the other choices, not least because there were very few such responses. We therefore excluded these data from the analysis.Data were analyzed for the 3,137 adults who provided complete information for the study variables. Respondents who did not provide education status , adjusted odds ratios (AOR), and 95 % confidence intervals (CI) were calculated for each variable. Associations between eating behaviors and household income or education were determined using binomial logistic regression analysis, with eating behaviors as the dependent variable and household income and education as independent variables. Household income as an independent variable was adjusted for age group; marital, residence and employment status; and education. Education as an independent variable was adjusted for age group; marital, residence and employment status; and household income. Previous studies have found that differences in eating behaviors were predicted by sex, age, marital status, and residence status , 17, 18,Table The prevalence of each eating behavior is shown in Table Table In the unadjusted analysis Table , taking p = 0.032; high-income households with men living with other people: AOR 0.70, 95 % CI 0.48—1.02, p = 0.065; and middle-income households with men living alone: AOR 0.82, 95 % CI 0.26–2.60, p = 0.730. It was impossible to estimate an AOR or 95 % CI for high-income households with men living alone because of the small sample size. Men with the highest levels of income were positively and significantly more likely than those with the lowest income levels to take care of their diet for the sake of their health, eat vegetables, use nutrition information, and converse during meals at the recommended levels. There was no significant association between frequency of eating breakfast and household income, but positive associations were seen between men’s household income, and the domains of taking care of one’s diet for health, eating vegetables, using nutrition information, conversations during meals, and negative association was seen between frequency of family breakfasts.Results of the logistic regression analysis of the association between household income and eating behaviors for men are shown in Table p = 0.225; high-income households with women living with others: AOR 0.65, 95 % CI 0.47–0.90, p = 0.010; and middle-income households with women living alone: AOR 1.90, 95 % CI 0.51–6.99, p = 0.337. The AOR and 95 % CI were impossible to estimate for high-income households with women living with others because of the small sample size. Women with the highest levels of income were positively and significantly more likely than those with the lowest levels of income to take care of their diet for the sake of their health, eat vegetables, use information on nutrition labels, and engage in conversation with family or friends during meals at the recommended levels. Frequency of eating breakfast and frequency of family breakfasts were not associated with household income. For household income among women, all domains showed significant trends.In the unadjusted analysis for women Table , most eaResults of the logistic regression analysis of the association between education and eating behaviors for men are shown in Table In the unadjusted analysis for women in Table In this study using an Internet-based survey, eating vegetables, using the information on nutrition labels, and engaging in positive conversations with family or friends during meals were positively associated with higher household incomes and education levels among both men and women. Lower frequency of family breakfasts and dinners were associated with higher household income among men; lower frequency of family dinners was associated with higher household income among women. These associations were not seen with education. We found that eating behaviors differed by SES, suggesting that supporting healthy eating behaviors could reduce health disparities. To our knowledge, this is the first study to investigate the association of SES with healthy eating behavior in Japanese adults.Our results showed that eating vegetables is a dietary behavior that is affected by SES. We set five small dishes of vegetables or about 350 g per day as a standard against which to assess frequency of vegetable intake. This is in line with a previous study in Japan . In bothWe found that participants with higher SES used the information on nutrition labels. Sinclair  reportedIn our sample of Japanese adults, few with a household income of 7,000,000 JPY or more reported frequent family meals. This is a new implication in this area of study. Frequent family meals in adolescence and young adulthood may have a lasting positive influence on dietary quality and meal patterns, such as greater intake of green, yellow, and other vegetables and fruit . Larson Despite participants with higher household incomes showing lower frequency of family meals, these participants conversed more about food during mealtimes. A previous study showed that Japanese children who engaged in mealtime conversations during meals had better dietary attitudes, eating behaviors, and quality of life; good health status; higher vegetable intake; and good table manners , 30, 46.This study had several limitations. First, our sample may not have been representative of the general population because we relied on an Internet-based survey. Caution should therefore be used in generalizing the results. Internet-based survey respondents may be more likely to have certain characteristics, such as being younger, having higher levels of education and income, and having better access to the Internet –49. The In this study, eating vegetables, using information found on nutrition labels, and engaging in positive conversations with family or friends during meals were significantly and positively associated with higher household income and education level among Japanese men and women. We identified a need to support eating behaviors that can help to reduce health disparities. SES should be considered in planning initiatives of health promotion and dietary improvement in Japan.


Recent studies have focused on the significant cytotoxicity of natural killer (NK) cells, cytokine-induced killer (CIK) cells, and gamma-delta (γδ) T cells in tumor cells. Nevertheless, the therapeutic features of these cell types have not been compared in the literature. The aim of this study was to evaluate the feasibility of activation and expansion of NK, CIK, and γδ T cells from cancer patients in vitro, and to clarify the differences in their antitumor capacities.NK, CIK, and γδ T cells were induced from the peripheral blood mononuclear cells of 20 cancer patients by using specific cytokines. Expression of CD69, NKG2D, CD16, granzyme B, perforin, IFN-γ, and IL-2 was measured by flow cytometry. Cytokine production and cytotoxicity were analyzed by enzyme-linked immunosorbent assay and Calcein-AM methods.NK cell proliferation was superior to that of CIK cells, but lower than that of γδ T cells. NK cells had a much stronger ability to secrete perforin, granzyme B, IFN-γ, and IL-2 than did CIK and γδ T cells, and imparted significantly higher overall cytotoxicity.Expanded NK cells from cancer patients are the most effective immune cells in the context of cytokine secretion and anti-tumor cytotoxicity in comparison to CIK and γδ T cells, making them an optimal candidate for adoptive cellular immunotherapy. Cellular immunity plays an essential role in anti-tumor activity. Immunocyte activity is often compromised in tumor patients, due to the inhibitory tumor environment. Therefore, numerous preliminary studies have demonstrated the safety and efficacy of adoptive cellular immunotherapy (ACI)—consisting of ex vivo expansion and activation of patient immunocytes followed by reinfusion . ACI has−CD56+ lymphocytes, can be rapidly activated to spontaneously attack certain abnormal cells in the body, particularly cancerous or virus-infected cells  × 100 %.NK, CIK, and γδ T cells were used as the effectors and leukemia cells , lymphoma cells (Raji), and multiple myeloma cells (U266) were used as targets. Briefly, target cells were collected, washed once with PBS, and suspended in PBS at 1 × 10Flow cytometry data were collected before and after induction and analyzed for correlation using Spearman’s test. Other data were analyzed by Wilcoxon’s rank-sum test in SPSS13.0 software. Results were considered significant at p ≤ 0.05.In this study, 20 tumor patients were enrolled. Informed consent in accordance with the Declaration of Helsinki was obtained from all patients, and approval was obtained from the Ethics Committee of the the First Hospital of Jilin University (protocol #2010–019). Inclusion criteria were no radiation or chemotherapy for at least one month before blood collection. Only patients with solid tumors were included. Patient characteristics are listed in the Table


Risk adjusted mortality for intensive care units (ICU) is usually estimated via logistic regression. Random effects (RE) or hierarchical models have been advocated to estimate provider risk-adjusted mortality on the basis that standard estimators increase false outlier classification. The utility of fixed effects (FE) estimators (separate ICU-specific intercepts) has not been fully explored.Using a cohort from the Australian and New Zealand Intensive Care Society Adult Patient Database, 2009–2010, the model fit of different logistic estimators  was characterised: Bayesian Information Criterion , receiver-operator characteristic curve area (AUC) and Hosmer-Lemeshow (H-L) statistic. ICU standardised hospital mortality ratios (SMR) and 95%CI were compared between models. ICU site performance (FE), relative to the grand observation-weighted mean (GO-WM) on odds ratio (OR), risk ratio (RR) and probability scales were assessed using model-based average marginal effects (AME).P-value = 0.22. The best-fitting random-intercept model had a BIC = 64457, AUC = 0.90 and H-L statistic P-value = 0.32 and random-coefficient model, BIC = 64556, AUC = 0.90 and H-L statistic P-value = 0.28. Across ICUs and over years no outliers  were identified and no model difference in SMR spread or 95%CI span was demonstrated. Using AME , ICU site-specific estimates diverged from the GO-WM, and the effect spread decreased over calendar years. On the probability scale, a majority of ICUs demonstrated calendar year decrease, but in the for-profit sector, this trend was reversed.The data set consisted of 145355 patients in 128 ICUs, years 2009 (47.5%) & 2010 (52.5%), with mean(SD) age 60.9(18.8) years, 56% male and ICU and hospital mortalities of 7.0% and 10.9% respectively. The FE model had a BIC = 64058, AUC = 0.90 and an H-L statistic The FE estimator had model advantage compared with conventional RE models. Using AME, between and over-year ICU site-effects were easily characterised. Risk-adjusted mortality has been used to characterise the performance of health care providers for a number of years However, such recommendation must also address certain cautions recently advanced regarding the latter methods The Australian and New Zealand Intensive Care Society (ANZICS) adult patient data base (APD) Access to the data was granted by the ANZICS Database Management Committee in accordance with standing protocols; local hospital  Ethics of Research Committee approval was waived. The data set analysed is the property of the ANZICS Data base and contributing ICUs and is not in the public domain. The data are available to personnel of the ANZICS Data base and contributing ICUs under specific conditions and upon written request.As previously described Analyses were performed using Stata ; continuous variables were reported as mean (SD), except where otherwise indicated, and statistical significance was ascribed at P≤0.05.Three separate models were estimated:i in provider k the logit (log-odds) of hospital mortality probability . The H the logistic cumulative distribution function.random effects  models, random intercept and random coefficient, as In the random intercept model, In the random coefficient (“slope”) model, the centred APACHE III score  adaptive quadrature, a computational method used to approximate the marginal likelihood by numerical integration Seasonality of mortality was addressed using trigonometric (sine and cosine) terms for yearly, 6 monthly and weekly effects after Stolwijk n was the number of points per bin, indicated ± 2SE bounds, within which one would expect about 95% of the binned residuals to fall.For fixed model variables, detailed above in “Methods”, sets of parameter coefficients were tested using a global Wald test Confidence intervals (CI) of the ICU standardised mortality ratio (SMR) were calculated by back-transformation from the variance of the (log) observed / predicted mortality using the Taylor series approximation X) and the values of  other model covariates i-th observation and x, coded (0/1), the marginal mean for x = 1 is obtained by considering all the observations of x wherever “x” appears in the model ; that is ith individual in the kth group and β, the Exploration of comparative ICU site performance, by ICU level and calendar year, relative to the grand observation-weighted mean The following effect computations with 95% CI were undertaken: (i) OR contrasts; using linear predictions via the “predict(xb)” option of the “margins” command and (ii) RR; as the ratio of the provider predictive margins divided by the grand weighted mean of the predictive margins, via nonlinear combination of estimates  & 2010 (52.5%), with mean(SD) age 60.9(18.8) years, APACHE III score 51.4(28.0) and ICU and hospital mortalities of 7.0% and 10.9% respectively. Fifty six percent were male and 38% were ventilated in the first 24 hours. The mean annual patient volume was 758(404); median 623, range 168–1701. The largest percentage of patients (42.8%) were in tertiary hospitals and resided in the Australian state of New South Wales (31.4%). Patient demographics over calendar year by hospital level and patient surgical status is seen in P = 0.22; grouping number  = 40). The continuous variable “annual-volume” and the categorical variables “ICU-level” and geographical-location” were identified as collinear and removed from the dependent variable list. When “annual-volume” was parameterised as a decile categorical variable, there was a model AIC increment of 5  were >0.1) and the variable was not further considered. A global Wald test of the 6 trigonometric seasonality parameters was significant at P = 0.004. A random intercept model with the identical independent variable list  had a BIC  = 64457, an AUC  = 0.90 and an H-L statistic  = 41.6 . A random coefficient model  as centred APACHE III score; unstructured covariance), including the variables “annual-volume” (continuous) and “ICU-level” and “geographical-location”  had a BIC  = 64555.8, an AUC  = 0.90 and an H-L statistic  = 42.8 . Both RE models satisfied the assumption of normality of random effects estimates  had a BIC  = 64057.9, an AUC  = 0.90 and an H-L statistic  = 18.3  of ICUs relative to the grand mean, by hospital level for the calendar years 2009 and 2010 is seen in Using a fixed-effects logistic model to generate provider mortality probabilities in a large data-base over a two year period we were unable to demonstrate (i) substantive advantage for a conventional random effects approach and (ii) outlier status for any of the ICUs. These findings deserve further comment.Multiple studies have compared fixed and random effects estimators in assessing provider performance, the key performance indicator usually being the (log)-SMR A number of papers have suggested that “non-hierarchical” estimators increase the possibility of false outlier classification The current study would appear to be one of the first to assess provider performance exploiting predictive margins, the use of the which has distinct advantages: the ability to resolve problems inherent in prediction across categorical predictors  were unable to be explicitly fitted in the FE model due to confounding / collinearity, but were “recovered” within the model based predictive margins analysis. Such confounding does not arise with RE modelling and FE and RE modelling approaches might be best characterised as complementary, rather than comparative.Conventional one-stage RE (and FE) estimation considers both “usually” and “unusually” performing providers, leading to inflated random effect variance estimates and the inability to properly account for the latter provider-type  in estimation. A staged approach to estimation which includes a "null" model describing the behaviour of "usual" providers would appear to be apposite The developed FE model had advantage compared with the conventional RE models and disclosed no ICU performance outliers in calendar years 2009–2010. Current developments in RE estimation, which embrace a “null” model and adjust for the false discovery rate and regression to the mean, are superior to a single application of a RE (or FE) model, but are considerably more complex statistically and computationally. Analysis using predictive margins allows substantial inferential insight into provider performance.File S1This file contains supporting information including Table S1-Table S3, Figure S1, and Figure S2. Table S1, Model estimates: fixed effects. Table S2, Model estimates: random intercept. Table S3, Model estimates: random coefficient. Figure S1, Standardized normal probability plots (P–P plot) of the random effects; random intercept model. Figure S2, Standardized normal probability plots (P–P plot) of the random effects; random coefficient model. a) Random effects for ICU site: APACHE III score. b) Random effects for ICU site: constant.(DOCX)Click here for additional data file.


The protein kinase, p38α MAPK, is a key intracellular transducer of stressor-induced neuroinflammatory responses and, as such, is of high interest as a potential therapeutic target. We recently reported the synthesis and evaluation of first-in-class CNS-penetrant and highly specific p38 MAPK inhibitors that avoid target crossover issues seen in popular small molecule p38 MAPK inhibitors used in hundreds of previous reports. The novel p38 MAPK inhibitors, represented in this study by MW181, are efficacious in vivo. Pharmacodynamic actions include attenuation of stressor-induced increases in brain proinflammatory cytokine levels. We report here more detailed analyses of MW181 target engagement and specific linkage to the downstream increase in glia proinflammatory cytokine production. In vivo validation included demonstration that oral administration of MW181 suppresses lipopolysaccharide-induced increases in mouse brain IL-1β, TNFα, IL-6, IL-10, and CXCL1 but not in a drug-resistant p38α MAPK mutant mouse. Neuroinflammation is a complex process that can be profoundly influenced by the cellular and environmental context, disease stage, and inciting stimuli. Microglia, as the resident tissue macrophage, are the archetypal cell in the CNS neuroinflammatory response. In addition, astrocytes, along with microglia, represent the resident cells in the nervous system responsible for neuroinflammation. In response to injury, infection, or other disturbances, microglia and astrocytes activate a patterned response to defend against and isolate inducing stimuli, which is followed by healing, repair, and resolution of the neuroinflammation. The reactive response of glia is fundamental for CNS homeostasis. However, the reactive glia response is context specific and highly variable, which can cause beneficial and/or detrimental forms of neuroinflammation . The novel CNS paradigm stands in contrast to previous efforts at targeting p38 MAPK in peripheral tissue disorders where preclinical studies and clinical trials showed a pharmacodynamics effect of attenuating blood proinflammatory cytokine levels, but the direct linkage of the proinflammatory cytokine levels to end organ damage was less than ideal or engagement of target tissue p38 MAPK activity was not established.The major challenge for p38 MAPK inhibitor in vivo studies in CNS disorders was, until recently, the issues found generally for targeting protein kinases in the CNS  mice but not in genetically engineered p38α MAPK inhibitor-resistant mice. These results provide further validation of the utility of MW181 as a highly selective chemical biology tool appropriate to study the role of p38α MAPK in CNS disorders that have neuroinflammation as a component of the disease mechanism.MW01-10-181SRM (=MW181) was synthesized as described -induced responses was tested in the murine microglial BV-2 cell line as previously described  mice (p38α KI mice) were generated as previously described , and cytokine/chemokine levels in mouse cortex lysates were measured using the MSD V-plex assay as previously described . For Fig. 50 values were made using a nonlinear regression with a variable Hill slope, with the data normalized to the positive control to fit the top and bottom plateaus. For Fig. n = 3. Significance was defined as a p < 0.05. Unless otherwise indicated, values are expressed as mean ± SEM.Statistical analysis used GraphPad prism software version 6  that comprise tissue barriers. If the drug is a PgP substrate, it is pumped back out (B→A direction) and eventually excreted from the body. The Pgp efflux pump status is especially important for CNS studies, as the pump removes susceptible small molecules from cells and is a major contributor to blood brain permeability problems for the vast majority of small molecule drugs and inhibitors. In addition, Pgp status can change with disease, drug treatment or innate immune status. The caco-2 bi-directional transport assay is a calibrated standard approach used to address permeability, which is a reflection of the drug’s molecular properties, and to determine if the drug is a PgP substrate. Briefly, cells are grown as a monolayer in a transwell, allowing directional measurement of transport. The net flux is determined as a ratio of B→A/A→B. If the ratio is >2, then more of the drug is pumped back out than enters by passive diffusion, usually due to the drug being a PgP substrate. Status is confirmed by incubating in the presence of a saturating concentration of a specific PgP substrate, verapamil. The competition for the PgP transporter results in the efflux ratio for the drug changing from >2 to <2. As summarized in Table 50 of 1,197 nM  and mitogen- and stress-activated kinase 1 (MSK1) are downstream substrates of p38α MAPK. To assess target engagement by MW181, we measured the levels of phosphorylated (p)-MK2 and pMSK1 in LPS-stimulated glia treated with MW181 at different concentrations. In our culture conditions without LPS, a basal activation of pMK2 is observed, which is further enhanced by LPS stimulation. MW181 was able to suppress both basal and LPS-induced levels of pMK2 with an IC nM Fig. . MW181 a nM Fig. .50 of 599 nM  = 4.041; p = 0.024). MW181 at 20 mg/kg suppressed IL-1β levels in LPS-challenged mice (p = 0.031), and at 5 mg/kg showed a trend, but when corrected for multiple comparisons the decrease did not reach significance. MW181 showed a strong inhibition of TNFα up-regulation  = 16.39; p < 0.0001), with both the 5 mg/kg and 20 mg/kg doses significantly suppressing TNFα levels . There was also a significant effect of MW181 on IL-6 levels in LPS-treated WT mice  = 5,808; p = 0.0056); again, 20 mg/kg of MW181 inhibited IL-6 levels (p = 0.0071), but the inhibition by MW181 at 5 mg/kg did not quite reach significance (p = 0.055). At this time window, MW181 also decreased the anti-inflammatory cytokine IL-10  = 5.436; p = 0.0075) at both compound concentrations . However, under the conditions of this study it cannot be stated if this is a direct effect of MW181 or feedback loop responses to blockage of proinflammatory cytokine increases. The cortical levels of CXCL1, a chemokine involved in neutrophil and oligodendrocyte precursor cell recruitment  = 5.834; p = 0.0055). Both the 5 mg/kg and 20 mg/kg doses of MW181 significantly suppressed CXCL1 levels compared to the LPS-stimulated WT mice treated with vehicle .Replicating the IL-1β result in vivo and extending the data of Watterson et al. , we founT106M KI mice (O’Keefe et al. To further assess in vivo target specificity of MW181 and better link kinase target engagement by the inhibitor to in vivo pharmacodynamic endpoints, we took advantage of the p38αThe studies reported here demonstrate that MW181 engages the p38α MAPK target in its glia cellular context and links the target engagement to the downstream pharmacodynamic effect of normalized CNS proinflammatory cytokine levels. The in vivo selectivity of MW181 action is further indicated by the failure to bring about its pharmacodynamic effects in p38α MAPK inhibitor resistant mice. Taken in its entirety, the results presented here and previously (Bachstetter et al.