Longitudinal Outcomes in Adolescents After Referral for Metabolic and Bariatric Surgery.

BACKGROUND: Metabolic and bariatric surgery (MBS) can be a well tolerated and effective treatment option for severe obesity in adolescents. We compared outcomes for adolescents that did and did not proceed to surgery. METHODS: A single-center longitudinal study (2015-2020). Patients were identified as LSG if they completed laparoscopic sleeve gastrectomy within 6 months of initial visit and NoLSG if they did not. Chi-square, Fisher exact, nonparametric Kruskal-Wallis tests, and Linear Mixed Models (LMM) were used to compare outcomes over 2 years. RESULTS: Three hundred fifty-two adolescents were referred with a mean age of 15.6 ±â€Š1.4, 69% girls, 38% Hispanic, and 78% had noncommercial insurance. The median baseline weight was 135 kg and body mass index (BMI) was 48 kg/m2; 42% had a BMI >50. Seventy-nine (22%) underwent LSG whereas 273 (78%) did not complete MBS primarily because of lack of interest. LSG patients had 21% total weight loss and 22% total BMI loss at 24 months whereas NoLSG patients had 4% total weight gain and 3% BMI gain (P < 0.01). Obesity-associated conditions improved in the LSG group (P < 0.01). Follow-up in both groups was poor (≤30% at 24 months). Patients with public insurance and those with BMI from 50 to 59.9 kg/m2 were high performing LSG patients. CONCLUSIONS: A minority (22%) of adolescents referred for MBS proceeded to surgery, despite its demonstrated efficacy and safety in adolescence. Those that did not undergo surgery continued to gain weight. Further research is needed to understand patient preferences or concerns related to MBS utilization during adolescence.

Copper and zinc ions govern the trans-directed dimerization of APP family members in multiple ways.

The amyloid precursor protein (APP) and its homologs amyloid precursor-like protein 1 (APLP1) and APLP2 have central physiological functions in transcellular adhesion that depend on copper and zinc mediated trans-directed dimerization of the extracellular domains E1 and E2. Copper binds to three distinct sites in APP, one in the copper binding (CuBD) and growth factor-like (GFLD) domains each within E1, and one in the E2 domain. For APLP1 and APLP2, metal binding has so far only been shown for the E2 domain. Zinc binding has been reported for all APP family members to a unique site in the E2 domain and an additional site essential for APLP1 E2 domain trans-dimerization. Using isothermal titration calorimetry, co-immunoprecipitation, and in vitro bead aggregation assays, we show that copper promotes cis- as well as trans-directed dimerization of APLP1 and APLP2, similar as reported previously for APP. Furthermore, we report a APP-specific zinc binding site with nanomolar affinity located in the E1 domain, whereas no binding of zinc to the individual subdomains GFLD or CuBD was detected. Zinc binding did not affect the cis- but trans-dimerization of APP and APLP1. Furthermore, zinc binding inhibited copper-induced trans-directed dimerization of APP. Together, we identified a high-affinity APP-specific zinc binding site in the E1 domain and revealed contrasting cis- and trans-directed dimerization properties of APP, APLP1, and APLP2 in dependence on zinc and copper ions. Consequently, changes in metal ion homeostasis, as reported in the context of synaptic activity and neurodegenerative diseases, appear as key modulators of homo- and heterotypic trans-cellular APP/APLPs complexes.

Characterising protein/detergent complexes by triple-detection size-exclusion chromatography.

BACKGROUND: In vitro investigations of membrane proteins usually depend on detergents for protein solubilisation and stabilisation. The amount of detergent bound to a membrane protein is relevant to successful experiment design and data analysis but is often unknown. Triple-detection size-exclusion chromatography enables simultaneous separation of protein/detergent complexes and protein-free detergent micelles and determination of their molar masses in a straightforward and absolute manner. Size-exclusion chromatography is used to separate different species, while ultraviolet absorbance, static light scattering, and refractive index measurements allow molar mass determination of protein and detergent components. RESULTS: We refined standard experimental and data-analysis procedures for challenging membrane-protein samples that elude routine approaches. The general procedures including preparatory steps, measurements, and data analysis for the characterisation of both routine and complex samples in difficult solvents such as concentrated denaturant solutions are demonstrated. The applicability of the protocol but also its limitations and possible solutions are discussed, and an extensive troubleshooting section is provided. CONCLUSIONS: We established and validated a protocol for triple-detection size-exclusion chromatography that enables the inexperienced user to perform and analyse measurements of well-behaved protein/detergent complexes. More experienced users are provided with an example of a more sophisticated analysis procedure allowing mass determination under challenging separation conditions.

Modulating bilayer mechanical properties to promote the coupled folding and insertion of an integral membrane protein.

Bilayer mechanical properties are not only of crucial importance to the mechanism of action of mechanosensation in lipid membranes but also affect preparative laboratory tasks such as membrane-protein refolding. We report this for coupled refolding and bilayer insertion of outer membrane phospholipase A (OmpLA), an integral membrane enzyme that catalyses the hydrolytic cleavage of glycerophospholipids. OmpLA can be refolded into a variety of detergent micelles and unilamellar vesicles composed of short-chain phospholipids but, in the absence of chemical or molecular chaperones, not into thicker membranes. Controlled modulation of bilayer mechanical properties by judicious use of subsolubilising concentrations of detergents induces monolayer curvature strain, acyl chain fluidisation, membrane thinning, and transient aqueous bilayer defects. This enables quantitative and functional refolding of OmpLA even into bilayer membranes composed of long-chain phospholipids to yield enzymatically active proteoliposomes without requiring membrane solubilisation.

Mental health factors associated with progression to adolescent metabolic and bariatric surgery.

BACKGROUND: While bariatric surgery has demonstrated significant weight loss for adolescents with severe obesity, only a limited number of adolescents referred to surgery successfully complete the surgical program. Better identification of pre-surgical factors, especially mental health factors, associated with completing bariatric surgery may determine successful referrals to surgical programs versus alternative behavioral health interventions. OBJECTIVES: The primary objective of this article was to investigate the relationship between presurgical mental health factors and whether or not a patient received bariatric surgery within the first six months of entering the program. SETTING: Pediatric Children's Hospital, Bariatric Surgery Program within Pediatric Surgery Department. METHODS: A retrospective medical record review of all patients referred from 2016 to 2019 to an adolescent surgical weight loss program was conducted. Patients were determined as completers if they received a laparoscopic sleeve gastrectomy within 6 months of entering the program. Demographic and preoperative mental health factors among completers (n = 30) were compared with surgery noncompleters (n = 44) using enter method logistic regression. RESULTS: Regression analyses revealed that younger patients, White patients, patients with lower internalizing symptoms on a standardized measure, and patients with greater self-reported motivation for surgery at their initial surgery visit were more likely to obtain surgery within 6 months of starting the program. CONCLUSION: The findings underscore the importance of assessing patients' motivation early in the bariatric surgery referral process. This study highlights potential treatment implications in how best to mitigate patient barriers, such as internalizing symptoms and low motivation, to pursue bariatric surgery and remain committed to weight loss goals through techniques such as motivational interviewing.

Real-time Visualization of Phospholipid Degradation by Outer Membrane Phospholipase A using High-Speed Atomic Force Microscopy.

Phospholipases are abundant in various types of cells and compartments, where they play key roles in physiological processes as diverse as digestion, cell proliferation, and neural activation. In Gram-negative bacteria, outer membrane phospholipase A (OmpLA) is involved in outer-membrane lipid homeostasis and bacterial virulence. Although the enzymatic activity of OmpLA can be probed with an assay relying on an artificial monoacyl thioester substrate, only little is known about its activity on diacyl phospholipids. Here, we used high-speed atomic force microscopy (HS-AFM) to directly image enzymatic phospholipid degradation by OmpLA in real time. In the absence of Ca2+, reconstituted OmpLA diffused within a phospholipid bilayer without revealing any signs of phospholipase activity. Upon the addition of Ca2+, OmpLA was activated and degraded the membrane with a turnover of ~2 phospholipid molecules per second and per OmpLA dimer until most of the membrane phospholipids were hydrolyzed and the protein became tightly packed.