Cyberbullying and eating disorder symptoms in US early adolescents.

OBJECTIVE: The objective of this study was to determine the association between cyberbullying and eating disorder symptoms in a national sample of 10-14-year-old early adolescents. METHOD: We analyzed cross-sectional data from the Adolescent Brain Cognitive Development (ABCD) Study (Year 2, 2018-2020, N = 10,258/11,875, 49% female, 46% non-White). Data were collected using multi-stage probability sampling. Modified Poisson regression analyses examined the association between cyberbullying and self-reported eating disorder symptoms based on the Kiddie Schedule for Affective Disorders and Schizophrenia (KSADS-5). RESULTS: Cyberbullying victimization was associated with worry about weight gain (prevalence ratio [PR] 2.41, 95% confidence interval [CI] 1.48-3.91), self-worth tied to weight (PR 2.08, 95% CI 1.33-3.26), inappropriate compensatory behavior to prevent weight gain (PR 1.95, 95% CI 1.57-2.42), binge eating (PR 1.95, 95% CI 1.59-2.39), and distress with binge eating (PR 2.64, 95% CI 1.94-3.59), in models adjusting for potential confounders. Cyberbullying perpetration was associated with worry about weight gain (PR 3.52, 95% CI 1.19-10.37), self-worth tied to weight (PR 5.59, 95% CI 2.56-12.20), binge eating (PR 2.36, 95% CI 1.44-3.87), and distress with binge eating (PR 2.84, 95% CI 1.47-5.49). DISCUSSION: Cyberbullying victimization and perpetration in early adolescence are associated with eating disorder symptoms. Clinicians may consider assessing for cyberbullying and eating disorder symptoms in early adolescence and provide anticipatory guidance. PUBLIC SIGNIFICANCE STATEMENT: Eating disorders often onset in adolescence and have among the highest mortality rates of any psychiatric disorder. In addition, cyberbullying has increased in prevalence among adolescents and significantly impacts mental health. In a national study of early adolescents, we found that cyberbullying victimization and perpetration are associated with eating disorder symptoms. Screening for and providing anticipatory guidance on cyberbullying and eating disorder symptoms in early adolescents may be warranted.

Impact of Hypoglossal Nerve Stimulation on Consumer Sleep Technology Metrics and Patient Symptoms.

OBJECTIVES: Obstructive sleep apnea (OSA) is usually assessed at discrete and infrequent timepoints. Wearable consumer sleep technologies (CST) may allow for more granular and longitudinal assessments of OSA therapy responses and OSA-related symptoms. METHODS: In this case series, we enrolled hypoglossal nerve stimulator (HGNS) patients who had an effective treatment response for an 8-week study using a wearable CST. Participants started with "HGNS-on," were randomized to turn off HGNS therapy during either week 4 or 5 ("HGNS-off"), followed by a return to therapy, "HGNS-resume." Participants completed validated symptom questionnaires assessing sleepiness, insomnia symptoms, functional status, and overall sleep health (Satisfaction, Alertness, Timing, Efficiency, and Duration, SATED) each week. CST metrics and survey scores were compared between HGNS treatment phases. Associations between CST metrics and survey scores were assessed. RESULTS: Seven participants with a total of 304 nights of CST data showed no statistically significant changes in total sleep time (TST), wake time after sleep onset, or sleep efficiency (SE) across the study periods. During HGNS-off, survey scores indicated significantly worsened OSA-related symptom scores. Two participants had significantly higher heart rate variability (HRV) during HGNS-off (by 3.3 and 6.3 ms) when compared to HGNS active therapy periods. Amongst CST metrics, SATED scores correlated with TST (r = 0.434, p < 0.0001), HRV (r = -0.486, p < 0.0001), and SE (r = 0.320, = 0.0014). In addition, FOSQ-10 scores correlated with average HR during sleep (r = -0.489, p < 0.001). CONCLUSION: A 1-week HGNS therapy withdrawal period impacted OSA-related sleep symptoms. Sleep-related metrics measured by a wearable CST correlated with symptom scores indicating potential value in the use of CSTs for longitudinal sleep-tracking in OSA patients. LEVEL OF EVIDENCE: 4 Laryngoscope, 134:3406-3411, 2024.

Bedtime Screen Use Behaviors and Sleep Outcomes in Early Adolescents: A Prospective Cohort Study.

PURPOSE: To determine prospective associations between bedtime screen use behaviors and sleep outcomes one year later in a national study of early adolescents in the United States. METHODS: We analyzed prospective cohort data from 9,398 early adolescents aged 11-12 years (48.4% female, 45% racial/ethnic minority) in the Adolescent Brain Cognitive Development Study (Years 2-3, 2018-2021). Regression analyses examined the associations between self-reported bedtime screen use (Year 2) and sleep variables (Year 3; self-reported sleep duration; caregiver-reported sleep disturbance), adjusting for sociodemographic covariates and sleep variables (Year 2). RESULTS: Having a television or Internet-connected electronic device in the bedroom was prospectively associated with shorter sleep duration one year later. Adolescents who left their phone ringer activated overnight had greater odds of experiencing sleep disturbance and experienced shorter sleep duration one year later, compared to those who turned off their phones at bedtime. Talking/texting on the phone, listening to music, and using social media were all prospectively associated with shorter sleep duration, greater overall sleep disturbance, and a higher factor score for disorders of initiating and maintaining sleep one year later. DISCUSSION: In early adolescents, several bedtime screen use behaviors are associated with adverse sleep outcomes one year later, including sleep disturbance and shorter weekly sleep duration. Screening for and providing anticipatory guidance on specific bedtime screen behaviors in early adolescents may be warranted.

The use of technology to improve medication adherence in heart failure patients: a systematic review of randomised controlled trials.

BACKGROUND: Heart failure is an ever-growing contributor to morbidity and mortality in the ageing population. Medication adherence rates among the HF population vary widely in the literature, with a reported range of 10-98%. Technologies have been developed to improve adherence to therapies and other clinical outcomes. AIMS: This systematic review aims to investigate the effect of different technologies on medication adherence in patients with heart failure. It also aims to determine their impact on other clinical outcomes and examine the potential of these technologies in clinical practice. METHODS: This systematic review was conducted using the following databases: PubMed Central UK, Embase, MEDLINE, CINAHL Plus, PsycINFO and Cochrane Library until October 2022. Studies were included if they were randomised controlled trials that used technology to improve medication adherence as an outcome in heart failure patients. The Cochrane Collaboration's Risk of Bias tool was used to assess individual studies. This review was registered with PROSPERO (ID: CRD42022371865). RESULTS: A total of nine studies met the inclusion criteria. Two studies showed statistically significant improvement in medication adherence following their respective interventions. Eight studies had at least one statistically significant result in the other clinical outcomes it measured, including self-care, quality of life and hospitalisations. All studies that evaluated self-care management showed statistically significant improvement. Improvements in other outcomes, such as quality of life and hospitalisations, were inconsistent. CONCLUSION: It is observable that there is limited evidence for using technology to improve medication adherence in heart failure patients. Further studies with larger study populations and validated self-reporting methods for medication adherence are required.

Analysis of the ability of vitamin D3-metabolizing cytochromes P450 to act on vitamin D3 sulfate and 25-hydroxyvitamin D3 3-sulfate.

25-Hydroxyvitamin D3 (25(OH)D3) is present in the human circulation esterified to sulfate with some studies showing that 25(OH)D3 3-sulfate levels are almost as high as unconjugated 25(OH)D3. Vitamin D3 is also present in human serum in the sulfated form as are other metabolites. Our aim was to determine whether sulfated forms of vitamin D3 and vitamin D3 metabolites can be acted on by vitamin D-metabolizing cytochromes P450 (CYPs), one of which (CYP11A1) is known to act on cholesterol sulfate. We used purified, bacterially expressed CYPs to test if they could act on the sulfated forms of their natural substrates. Purified CYP27A1 converted vitamin D3 sulfate to 25(OH)D3 3-sulfate with a catalytic efficiency (kcat/Km) approximately half that for the conversion of vitamin D3 to 25(OH)D3. Similarly, the rate of metabolism of vitamin D3 sulfate was half that of vitamin D3 for CYP27A1 in rat liver mitochondria. CYP2R1 which is also a vitamin D 25-hydroxylase did not act on vitamin D3 sulfate. CYP11A1 was able to convert vitamin D3 sulfate to 20(OH)D3 3-sulfate but at a considerably lower rate than for conversion of vitamin D3 to 20(OH)D3. 25(OH)D3 3-sulfate was not metabolized by the activating enzyme, CYP27B1, nor by the inactivating enzyme, CYP24A1. Thus, we conclude that 25(OH)D3 3-sulfate in the circulation may act as a pool of metabolically inactive vitamin D3 to be released by hydrolysis at times of need whereas vitamin D3 sulfate can be metabolized in a similar manner to free vitamin D3 by CYP27A1 and to a lesser degree by CYP11A1.

Parents' perspectives of anorexia nervosa treatment in adolescents: a systematic review and metasynthesis of qualitative data.

BACKGROUND: Studies have established the central role of the family in the recognition, treatment, and recovery of anorexia nervosa. The objective of this study was to review, synthesize, and critically appraise the literature on parents' views on the treatment and recovery process of anorexia nervosa in their adolescent child. METHOD: A systematic search of Medline, PsychINFO, CINHAL, EMBASE, Cochrane library, and SSCI was conducted for qualitative studies published regarding parents' views about the treatment of anorexia nervosa. The quality of articles was assessed using the critical appraisal skills program (CASP) and findings were analysed using thematic synthesis. RESULTS: A total of 25 studies from nine countries reporting the views of 357 parents met the inclusion criteria. Four major themes were developed from the analysis: understanding the child and the disease, experience of services and treatment modalities, the role of professionals, and the experience of recovery. CONCLUSION: Parents report struggles with delays in finding help, judgmental attitudes of professionals, and uncertainty about the future. Recognition of the challenges faced by parents and families empowers clinicians to build stronger therapeutic relationships essential for long-term recovery from anorexia nervosa.

Interventions for adolescents with anorexia nervosa often focus on families. Although prior systematic reviews have incorporated adolescents' perspectives on their treatment for anorexia nervosa, none have previously reviewed parents' perspectives on their adolescents' treatment. We conducted a systematic review and metasynthesis of qualitative data capturing parents' perspectives of anorexia nervosa treatment in adolescents. Four themes emerged from the analysis: understanding the child and the disease, experience of services and treatment modalities, the role of professionals, and the experience of recovery. Parents report struggles with delays in finding help, judgmental attitudes of professionals, and uncertainty about the future. Recognition of the challenges faced by parents and families empowers clinicians to build stronger therapeutic relationships essential for long-term recovery from anorexia nervosa.

Nipsnap1-A regulatory factor required for long-term maintenance of non-shivering thermogenesis.

OBJECTIVE: The activation of non-shivering thermogenesis (NST) has strong potential to combat obesity and metabolic disease. The activation of NST however is extremely temporal and the mechanisms surrounding how the benefits of NST are sustained once fully activated, remain unexplored. The objective of this study is to investigate the role of 4-Nitrophenylphosphatase Domain and Non-Neuronal SNAP25-Like 1 (Nipsnap1) in NST maintenance, which is a critical regulator identified in this study. METHODS: The expression of Nipsnap1 was profiled by immunoblotting and RT-qPCR. We generated Nipsnap1 knockout mice (N1-KO) and investigated the function of Nipsnap1 in NST maintenance and whole-body metabolism using whole body respirometry analyses. We evaluate the metabolic regulatory role of Nipsnap1 using cellular and mitochondrial respiration assay. RESULTS: Here, we show Nipsnap1 as a critical regulator of long-term thermogenic maintenance in brown adipose tissue (BAT). Nipsnap1 localizes to the mitochondrial matrix and increases its transcript and protein levels in response to both chronic cold and ß3 adrenergic signaling. We demonstrated that these mice are unable to sustain activated energy expenditure and have significantly lower body temperature in the face of an extended cold challenge. Furthermore, when mice are exposed to the pharmacological ß3 agonist CL 316, 243, the N1-KO mice exhibit significant hyperphagia and altered energy balance. Mechanistically, we demonstrate that Nipsnap1 integrates with lipid metabolism and BAT-specific ablation of Nipsnap1 leads to severe defects in beta-oxidation capacity when exposed to a cold environmental challenge. CONCLUSION: Our findings identify Nipsnap1 as a potent regulator of long-term NST maintenance in BAT.

Ado-Mediated Depletion of Taurine Impairs Mitochondrial Respiratory Capacity and Alters the Chromatin Landscape of Inguinal Adipose Tissue.

Non-shivering thermogenesis (NST) has strong potential to combat obesity; however, a safe molecular approach to activate this process has not yet been identified. The sulfur amino acid taurine has the ability to safely activate NST and confer protection against obesity and metabolic disease in both mice and humans, but the mechanism of this action is unknown. In this study, we discover that a suite of taurine biosynthetic enzymes, especially that of cysteamine dioxygenase (ADO), significantly increases in response to ß3 adrenergic signaling in inguinal adipose tissue (IWAT) in order to increase intracellular concentrations of taurine. We further show that ADO is critical for thermogenic mitochondrial respiratory function as its ablation in adipocytes significantly reduces taurine levels, which leads to declines in mitochondrial oxygen consumption rates. Finally, we demonstrate via assay for transposase-accessible chromatin with sequencing (ATAC-seq) that taurine supplementation in beige adipocytes has the ability to remodel the chromatin landscape to increase the chromatin accessibility and transcription of genes, such as glucose-6-phosphate isomerase 1 (Gpi1), which are critical for NST. Taken together, our studies highlight a potential mechanism for taurine in the activation of NST that can be leveraged toward the treatment of obesity and metabolic disease.

Body weight distortions in an auditory-driven body illusion in subclinical and clinical eating disorders.

Previous studies suggest a stronger influence of visual signals on body image in individuals with eating disorders (EDs) than healthy controls; however, the influence of other exteroceptive sensory signals remains unclear. Here we used an illusion relying on auditory (exteroceptive) signals to manipulate body size/weight perceptions and investigated whether the mechanisms integrating sensory signals into body image are altered in subclinical and clinical EDs. Participants' footstep sounds were altered to seem produced by lighter or heavier bodies. Across two experiments, we tested healthy women assigned to three groups based on self-reported Symptomatology of EDs (SED), and women with Anorexia Nervosa (AN), and used self-report, body-visualization, and behavioural (gait) measures. As with visual bodily illusions, we predicted stronger influence of auditory signals, leading to an enhanced body-weight illusion, in people with High-SED and AN. Unexpectedly, High-SED and AN participants displayed a gait typical of heavier bodies and a widest/heaviest visualized body in the 'light' footsteps condition. In contrast, Low-SED participants showed these patterns in the 'heavy' footsteps condition. Self-reports did not show group differences. The results of this pilot study suggest disturbances in the sensory integration mechanisms, rather than purely visually-driven body distortions, in subclinical/clinical EDs, opening opportunities for the development of novel diagnostic/therapeutic tools.

Electrographic Features of Spontaneous Recurrent Seizures in a Mouse Model of Extended Hippocampal Kindling.

Epilepsy is a chronic neurological disorder characterized by spontaneous recurrent seizures (SRS) and comorbidities. Kindling through repetitive brief stimulation of a limbic structure is a commonly used model of temporal lobe epilepsy. Particularly, extended kindling over a period up to a few months can induce SRS, which may simulate slowly evolving epileptogenesis of temporal lobe epilepsy. Currently, electroencephalographic (EEG) features of SRS in rodent models of extended kindling remain to be detailed. We explored this using a mouse model of extended hippocampal kindling. Intracranial EEG recordings were made from the kindled hippocampus and unstimulated hippocampal, neocortical, piriform, entorhinal, or thalamic area in individual mice. Spontaneous EEG discharges with concurrent low-voltage fast onsets were observed from the two corresponding areas in nearly all SRS detected, irrespective of associated motor seizures. Examined in brain slices, epileptiform discharges were induced by alkaline artificial cerebrospinal fluid in the hippocampal CA3, piriform and entorhinal cortical areas of extended kindled mice but not control mice. Together, these in vivo and in vitro observations suggest that the epileptic activity involving a macroscopic network may generate concurrent discharges in forebrain areas and initiate SRS in hippocampally kindled mice.

Analysing coherence of oral discourse among Cantonese speakers in Mainland China with traumatic brain injury and cerebrovascular accident.

Purpose: Coherence can reflect subtle language deficits in individuals with traumatic brain injury (TBI) and cerebrovascular accident (CVA). This study aimed at investigating whether global and local coherence in Cantonese-speaking adults with CVA and TBI differ from non-brain-injured (NBI) speakers. Factors contributing to the coherence ratings and impacts of elicitation tasks on coherence were examined.Method: Two clinical groups with fluent aphasia (7 CVA and 11 TBI) and 18 controls matched in age and education, who were Cantonese speakers living in China participated. Language samples of single and sequential picture description and storytelling were elicited, and subsequently analysed on global and local coherence, content sequence, and informativeness.Result: TBI speakers had impaired global and local coherence, while CVA speakers had poor global coherence. Sequence of main events produced by the three groups correlated significantly with global coherence. Attention and visuospatial skills were also significantly related to global coherence in both clinical groups. Finally, impaired language integrity was associated with problems of local coherence.Conclusion: The results were consistent with previous studies. Linguistic deficits of coherence in discourse in the two clinical groups and possible impacts of elicitation tasks on the cognitive demands and coherence ratings were discussed.

Investigating PSMA-Targeted Radioligand Therapy Efficacy as a Function of Cellular PSMA Levels and Intratumoral PSMA Heterogeneity.

PURPOSE: Prostate-specific membrane antigen (PSMA) targeting radioligands deliver radiation to PSMA-expressing cells. However, the relationship between PSMA levels and intralesion heterogeneity of PSMA expression, and cytotoxic radiation by radioligand therapy (RLT) is unknown. Here we investigate RLT efficacy as function of PSMA levels/cell, and the fraction of PSMA+ cells in a tumor. EXPERIMENTAL DESIGN: RM1 cells expressing different levels of PSMA (PSMA-, PSMA+, PSMA++, PSMA+++; study 1) or a mix of PSMA+ and PSMA- RM1 (study 2, 4) or PC-3/PC-3-PIP (study 3) cells at various ratios were injected into mice. Mice received 177Lu- (studies 1-3) or 225Ac- (study 4) PSMA617. Tumor growth was monitored. Two days post-RLT, tumors were resected in a subset of mice. Radioligand uptake and DNA damage were quantified. RESULTS: 177Lu-PSMA617 efficacy increased with increasing PSMA levels (study 1) and fractions of PSMA positive cells (studies 2, 3) in both, the RM1 and PC-3-PIP models. In tumors resected 2 days post-RLT, PSMA expression correlated with 177Lu-PSMA617 uptake and the degree of DNA damage. Compared with 177Lu-PSMA617, 225Ac-PSMA617 improved overall antitumor effectiveness and tended to enhance the differences in therapeutic efficacy between experimental groups. CONCLUSIONS: In the current models, both the degree of PSMA expression and the fraction of PSMA+ cells correlate with 177Lu-/225Ac-PSMA617 tumor uptake and DNA damage, and thus, RLT efficacy. Low or heterogeneous PSMA expression represents a resistance mechanism to RLT.See related commentary by Ravi Kumar and Hofman, p. 2774.

The serum vitamin D metabolome: What we know and what is still to discover.

Vitamin D, referring to the two forms, D2 from the diet and D3 primarily derived from phototransformation in the skin, is a prohormone important in human health. The most hormonally active form, 1α,25-dihydroxyvitamin D (1α,25(OH)2D), formed from vitamin D via 25-hydroxyvitamin D (25(OH)D), is not only important for regulating calcium metabolism, but has many pleiotropic effects including regulation of the immune system and has anti-cancer properties. The major circulating form of vitamin D is 25(OH)D and both D2 and D3 forms are routinely measured by LC/MS/MS to assess vitamin D status, due to their relatively long half-lives and much higher concentrations compared to 1α,25(OH)2D. Inactivation of both 25(OH)D and 1α,25(OH)2D is catalyzed by CYP24A1 and 25-hydroxyvitamin D3 3-epimerase. Initial products from these enzymes acting on 25(OH)D3 are 24R,25(OH)2D3 and 3-epi-25(OH)D3, respectively, and both of these can also be measured routinely in some clinical laboratories to further document vitamin D status. With advances in LC/MS/MS and its increased availability, and with the help of studies with recombinant vitamin D-metabolizing enzymes, many other vitamin D metabolites have now been detected and in some cases quantitated, in human serum. CYP11A1 which catalyzes the first step in steroidogenesis, has been found to also act on vitamins D3 and D2 hydroxylating both at C20, but with some secondary metabolites produced by subsequent hydroxylations at other positions on the side chain. The major vitamin D3 metabolite, 20S-hydroxyvitamin D3 (20S(OH)D3), shows biological activity, often similar to 1α,25(OH)2D3 but without calcemic effects. Using standards produced enzymatically by purified CYP11A1 and characterized by NMR, many of these new metabolites have been detected in human serum, with semi-quantitative measurement of 20S(OH)D3 indicating it is present at comparable concentrations to 24R,25(OH)2D3 and 3-epi-25(OH)D3. Recently, vitamin D-related hydroxylumisterols derived from lumisterol3, a previtamin D3 photoproduct, have also been measured in human serum and displayed biological activity in initial in vitro studies. With the current extensive knowledge on the reactions and pathways of metabolism of vitamin D, especially those catalyzed by CYP24A1, CYP27A1, CYP27B1, CYP3A4 and CYP11A1, it is likely that many other of the resulting hydroxyvitamin D metabolites will be measured in human serum in the future, some contributing to a more detailed understanding of vitamin D status in health and disease.

Impaired Spatial Learning and Memory in Middle-Aged Mice with Kindling-Induced Spontaneous Recurrent Seizures.

Temporal lobe epilepsy is the most common and often drug-resistant type of epilepsy in the adult and aging populations and has great diversity in etiology, electro-clinical manifestations, and comorbidities. Kindling through repeated brief stimulation of limbic structures is a commonly used model of temporal lobe epilepsy. Particularly, extended kindling can induce spontaneous recurrent seizures in several animal species. However, kindling studies in middle-aged, aging, or aged animals remain scarce, and currently, little is known about kindling-induced behavioral changes in middle-aged/aging animals. We therefore attempted to provide more information in this area using a mouse model of extended hippocampal kindling. We conducted experiments in middle-aged mice (C57BL/6, male, 12-14 months of age) to model new-onset epilepsy in adult/aging populations. Mice experienced twice daily hippocampal stimulations or handling manipulations for 60-70 days and then underwent continuous electroencephalogram (EEG)-video monitoring to detect spontaneous recurrent seizures. Extended kindled mice consistently exhibited spontaneous recurrent seizures with mean incidences of 6-7 events per day, and these seizures featured EEG discharges and corresponding convulsions. The handling control mice showed neither seizure nor aberrant EEG activity. The two groups of mice underwent the Morris water maze test of spatial learning and memory 1-2 weeks after termination of the kindling stimulation or handling manipulation. During visible platform trials, the kindled mice took a longer distance and required more time than the control mice to find the platform. During hidden platform trials, the kindled mice showed no improvement over 5-day trials in finding the platform whereas the control mice improved significantly. During probe tests in which the hidden platform was removed, the kindled mice spent less time than the controls searching in the correct platform location. There were no significant differences between the kindled and control mice with respect to swim speed or total locomotor activity in an open-field test. Together, these observations indicate that the extended kindled mice with spontaneous recurrent seizures are impaired in spatial learning and memory as assessed by the Morris water maze test. We postulate that the extended hippocampal kindling in middle-aged mice may help explore epileptogenic mechanisms and comorbidities potentially relevant to new-onset temporal lobe epilepsy in adult and aging patients. Limitations and confounds of our present experiments are discussed to improve future examinations of epileptic comorbidities in extended kindled mice.

Properties of purified CYP2R1 in a reconstituted membrane environment and its 25-hydroxylation of 20-hydroxyvitamin D3.

Recent studies indicate that CYP2R1 is the major 25-hydroxylase catalyzing the first step in vitamin D activation. Since the catalytic properties of CYP2R1 have been poorly studied to date and it is a membrane protein, we examined the purified enzyme in a membrane environment. CYP2R1 was expressed in E. coli and purified by nickel affinity- and hydrophobic interaction-chromatography and assayed in a reconstituted membrane system comprising phospholipid vesicles plus purified human NADPH-P450 oxidoreductase. CYP2R1 converted vitamin D3 in the vesicle membrane to 25-hydroxyvitamin D3 [25(OH)D3] with good adherence to Michaelis-Menten kinetics. The kinetic parameters for 25-hydroxylation of vitamin D3 by the two major vitamin D 25-hydroxylases, CYP2R1 and CYP27A1, were examined in vesicles under identical conditions. CYP2R1 displayed a slightly lower kcat than CYP27A1 but a much lower Km for vitamin D3, and thus an overall 17-fold higher catalytic efficiency (kcat/Km), consistent with CYP2R1 being the major vitamin D 25-hydroxylase. 20-Hydroxyvitamin D3 [20(OH)D3], the main product of vitamin D3 activation by an alternative pathway catalyzed by CYP11A1, was metabolized by CYP2R1 to 20,25-dihydroxyvitamin D3 [20,25(OH)2D3], with catalytic efficiency similar to that for the 25-hydroxylation of vitamin D3. 20,25(OH)2D3 retained full, or somewhat enhanced activity compared to the parent 20(OH)D3 for the inhibition of the proliferation of melanocytes and dermal fibroblasts, with a potency comparable to 1,25-dihydroxyvitamin D3 [1,25(OH)2D3]. The 20,25(OH)2D3 was also able to act as an inverse agonist on retinoic acid-related orphan receptor α, like its parent 20(OH)D3. Thus, the major findings of this study are that CYP2R1 can metabolize substrates in a membrane environment, the enzyme displays higher catalytic efficiency than CYP27A1 for the 25-hydroxylation of vitamin D, it efficiently hydroxylates 20(OH)D3 at C25 and this product retains the biological activity of the parent compound.

Diabetes Outcomes More than a Decade Following Sustained Weight Loss After Laparoscopic Adjustable Gastric Band Surgery.

BACKGROUND: Long-term outcome data are needed to define the role of bariatric surgery in type 2 diabetes (T2D). To address this, we collated diabetes outcomes more than a decade after laparoscopic adjustable gastric band (LAGB) surgery. METHOD: Clinical and biochemical measures from 113 obese T2D patients who underwent LAGB surgery in 2003 and 2004 were analyzed. Diabetes remission was defined as HbA1c < 6.2% (44 mmol/mol) and fasting glucose < 7.0 mmol/L. RESULTS: Seventy-nine patients had weight data at 10 years and attained a median [Q1, Q3] weight loss of 16 [10, 21] percent. Sixty patients attended a follow-up assessment. Their baseline HbA1c of 7.8 [7.1, 9.3] percentage units (62 [54, 78] mmol/mol) had decreased to 6.6 [6.1, 8.4] (49 [43, 68] mmol/mol) despite no significant change in glucose-lowering therapy. Eleven patients (18%) were in diabetes remission and another 18 had HbA1c ≤ 6.5%. Significant improvements in physical measures of quality of life, blood pressure, and lipid profile were also observed but there was no change in the proportion of patients with albuminuria and a significant decline in estimated glomerular filtration rate. Twelve patients in the follow-up cohort (20%) required anti-reflux medication after surgery and 26 (43%) underwent gastric band revision surgery. CONCLUSION: Weight loss for over 10 years after LAGB surgery delivers clinically meaningful improvements in HbA1c, blood pressure, lipids, and quality of life at the cost of a high rate of revision surgery and increased use of anti-reflux medication. These findings support the use of bariatric surgery as a long-term treatment for weight loss and wellbeing in patients with T2D. STUDY REGISTRATION: Registered with the Australian Clinical trials registry as ACTRN12615000089538.

CYP27A1 acts on the pre-vitamin D3 photoproduct, lumisterol, producing biologically active hydroxy-metabolites.

Prolonged exposure of the skin to UV radiation causes previtamin D3, the initial photoproduct formed by opening of the B ring of 7-dehydrocholesterol, to undergo a second photochemical reaction where the B-ring is reformed giving lumisterol3 (L3), a stereoisomer of 7-dehydrocholesterol. L3 was believed to be an inactive photoproduct of excessive UV radiation whose formation prevents excessive vitamin D production. Recently, we reported that L3 is present in serum and that CYP11A1 can act on L3 producing monohydroxy- and dihydroxy-metabolites which inhibit skin cell proliferation similarly to 1α,25-dihydroxyvitamin D3. In this study we tested the ability of human CYP27A1 to hydroxylate L3. L3 was metabolized by purified CYP27A1 to 3 major products identified as 25-hydroxyL3, (25R)-27-hydroxyL3 and (25S)-27-hydroxyL3, by NMR. These three products were also seen when mouse liver mitochondria containing CYP27A1 were incubated with L3. The requirement for CYP27A1 for their formation by mitochondria was confirmed by the inhibition of their synthesis by 5ß-cholestane-3α,7α,12α-triol, an intermediate in bile acid synthesis which serves as an efficient competitive substrate for CYP27A1. CYP27A1 displayed a high kcat for the metabolism of L3 (76 mol product/min/mol CYP27A1) and a catalytic efficiency (kcat/Km) that was 260-fold higher than that for vitamin D3. The CYP27A1-derived hydroxy-derivatives inhibited the proliferation of cultured human melanoma cells and colony formation with IC50 values in the nM range. Thus, L3 is metabolized efficiently by CYP27A1 with hydroxylation at C25 or C27 producing metabolites potent in their ability to inhibit melanoma cell proliferation, supporting that L3 is a prohormone which can be activated by CYP-dependent hydroxylations.

Effects of Antiepileptic Drugs on Spontaneous Recurrent Seizures in a Novel Model of Extended Hippocampal Kindling in Mice.

Epilepsy is a common neurological disorder characterized by naturally-occurring spontaneous recurrent seizures and comorbidities. Kindling has long been used to model epileptogenic mechanisms and to assess antiepileptic drugs. In particular, extended kindling can induce spontaneous recurrent seizures without gross brain lesions, as seen clinically. To date, the development of spontaneous recurrent seizures following extended kindling, and the effect of the antiepileptic drugs on these seizures are not well understood. In the present study we aim to develop a mouse model of extended hippocampal kindling for the first time. Once established, we plan to evaluate the effect of three different antiepileptic drugs on the development of the extended-hippocampal-kindled-induced spontaneous recurrent seizures. Male C57 black mice were used for chronic hippocampal stimulations or handling manipulations (twice daily for up to 70 days). Subsequently, animals underwent continuous video/EEG monitoring for seizure detection. Spontaneous recurrent seizures were consistently observed in extended kindled mice but no seizures were detected in the control animals. The aforementioned seizures were generalized events characterized by hippocampal ictal discharges and concurrent motor seizures. Incidence and severity of the seizures was relatively stable while monitored over a few months after termination of the hippocampal stimulation. Three antiepileptic drugs with distinct action mechanisms were tested: phenytoin, lorazepam and levetiracetam. They were applied via intra-peritoneal injections at anticonvulsive doses and their effects on the spontaneous recurrent seizures were analyzed 10-12 h post-injection. Phenytoin (25 mg/kg) and levetiracetam (400 mg/kg) abolished the spontaneous recurrent seizures. Lorazepam (1.5 mg/kg) decreased motor seizure severity but did not reduce the incidence and duration of corresponding hippocampal discharges, implicating its inhibitory effects on seizure spread. No gross brain lesions were observed in a set of extended hippocampal kindled mice submitted to histological evaluation. All these data suggests that our model could be considered as a novel mouse model of extended hippocampal kindling. Some limitations remain to be considered.

Investigation of 20S-hydroxyvitamin D3 analogs and their 1α-OH derivatives as potent vitamin D receptor agonists with anti-inflammatory activities.

20S-hydroxyvitamin D3 [20S(OH)D3] is anti-inflammatory and not hypercalcemic, suggesting its potential as a lead compound. In this study, side chain modified 20S(OH)D3 analogs (4, 13, 23 and 33) together with their 1α-OH derivatives were synthesized and their metabolism and biological activities tested. 4, 13 and 23 are good substrates for CYP27B1, enabling enzymatic synthesis of their 1α-OH derivatives 5, 14 and 24. However, 33 could not be hydroxylated by CYP27B1 and acts as an inhibitor. All analogs were poorer substrates for CYP24A1 than calcitriol, indicating improved catabolic stability. While the parent analogs showed minimal VDR stimulating activity, their 1α-OH derivatives were potent VDR agonists. 4, 5, 14 and 24 significantly upregulated the expression of CYP24A1 at the mRNA level, consistent with their VDR activation abilities and indicating that 1α-hydroxylation is required to produce analogs with strong activity. These analogs have anti-inflammatory activities that are influenced by side chain composition and by 1α-hydroxylation. To understand their molecular interactions with the VDR, 20S(OH)D3, 4 and 33 were co-crystalized with the VDR ligand binding domain, which revealed subtle differences to the calcitriol-bound receptor. This study demonstrates the potential of the 20S(OH)D3 scaffold for the development of novel anti-inflammatory agents.

ATR inhibition facilitates targeting of leukemia dependence on convergent nucleotide biosynthetic pathways.

Leukemia cells rely on two nucleotide biosynthetic pathways, de novo and salvage, to produce dNTPs for DNA replication. Here, using metabolomic, proteomic, and phosphoproteomic approaches, we show that inhibition of the replication stress sensing kinase ataxia telangiectasia and Rad3-related protein (ATR) reduces the output of both de novo and salvage pathways by regulating the activity of their respective rate-limiting enzymes, ribonucleotide reductase (RNR) and deoxycytidine kinase (dCK), via distinct molecular mechanisms. Quantification of nucleotide biosynthesis in ATR-inhibited acute lymphoblastic leukemia (ALL) cells reveals substantial remaining de novo and salvage activities, and could not eliminate the disease in vivo. However, targeting these remaining activities with RNR and dCK inhibitors triggers lethal replication stress in vitro and long-term disease-free survival in mice with B-ALL, without detectable toxicity. Thus the functional interplay between alternative nucleotide biosynthetic routes and ATR provides therapeutic opportunities in leukemia and potentially other cancers.Leukemic cells depend on the nucleotide synthesis pathway to proliferate. Here the authors use metabolomics and proteomics to show that inhibition of ATR reduced the activity of these pathways thus providing a valuable therapeutic target in leukemia.