Two-Photon Microscopy to Measure Calcium Signaling in the Living Brain.

Two-photon microscopy enables imaging of calcium signaling at cellular or subcellular resolution up to hundreds of microns deep in the living brain. Changes in the brightness of fluorescent calcium indicators provide a readout of calcium levels over time, affording information about neuronal activity and/or calcium-dependent subcellular signaling. Here, we describe a protocol for repeated two-photon imaging of calcium signals in mice expressing a genetically encoded calcium indicator that have been implanted with a chronic cranial window.

Metabolic engineering of yeast for de novo production of kratom monoterpene indole alkaloids.

Monoterpene indole alkaloids (MIAs) from Mitragyna speciosa ("kratom"), such as mitragynine and speciogynine, are promising novel scaffolds for opioid receptor ligands for treatment of pain, addiction, and depression. While kratom leaves have been used for centuries in South-East Asia as stimulant and pain management substance, the biosynthetic pathway of these psychoactives have only recently been partially elucidated. Here, we demonstrate the de novo production of mitragynine and speciogynine in Saccharomyces cerevisiae through the reconstruction of a five-step synthetic pathway from common MIA precursor strictosidine comprising fungal tryptamine 4-monooxygenase to bypass an unknown kratom hydroxylase. Upon optimizing cultivation conditions, a titer of ∼290 µg/L kratom MIAs from glucose was achieved. Untargeted metabolomics analysis of lead production strains led to the identification of numerous shunt products derived from the activity of strictosidine synthase (STR) and dihydrocorynantheine synthase (DCS), highlighting them as candidates for enzyme engineering to further improve kratom MIAs production in yeast. Finally, by feeding fluorinated tryptamine and expressing a human tailoring enzyme, we further demonstrate production of fluorinated and hydroxylated mitragynine derivatives with potential applications in drug discovery campaigns. Altogether, this study introduces a yeast cell factory platform for the biomanufacturing of complex natural and new-to-nature kratom MIAs derivatives with therapeutic potential.

Positive experiences with the use of an eHealth smartphone application during the treatment of an abdominal aortic aneurysm.

OBJECTIVE: To evaluate the patient perceived satisfaction and feasibility of a personalized eHealth application (app) for abdominal aortic aneurysm (AAA) patients undergoing surgery. METHODS: Patients were offered to download the app prior to undergoing AAA surgery, in a prospective single centre cohort study, using a mixed methods sequential explanatory design. It offers information via the timely delivery of push notifications with text, images, and videos. The information includes chapters regarding the AAA, surgical techniques (endovascular aneurysm repair and open surgical repair), and perioperative lifestyle advice such as; physical exercise programmes, healthy and protein rich diet, geriatric care, and to stop smoking or drinking alcohol. RESULTS: The app was installed by 59/65 patients (91%). After installation, six patients deactivated the app (10%). The mean age was 74 years (SD = 7) and 85% of patients were male. The app was opened a median of 67 times (interquartile range [IQR] 33-127) and with a median time interval of 50 hours (IQR 28-74). Overall, 90% (53/59) completed a satisfaction questionnaire. On a numeric rating scale from 0 to 10, the median scored satisfaction for guidance was 8 (IQR 6-8), provided information was 8 (IQR 6-8), usefulness was 7 (IQR 6-8.5), and for recommending it to others 8 (IQR 6-9). Using purposeful sampling, seven patients underwent a semi-structured interview on user-experience of the app. They described experiencing positive changes to lifestyle habits, and appreciating the ability to share it with loved ones or informal caregivers. Several areas of improvement were reported. CONCLUSION: The personalized eHealth app is feasible in older AAA patients and valued as a useful supplement to the standard of care. We argue that the app aids in managing a prehabilitation programme, aids in the digital transformation of healthcare, and thereby decreases the workload of hospital staff.

Serotype 15C Streptococcus pneumoniae resistant to classical complement deposition and agglutination by polyclonal rabbit anti-capsular 15B sera.

BACKGROUND: S. pneumoniae (SP) serotypes 15B and 15C are frequent causative agents of invasive pneumococcal disease (IPD), otitis media, and nasopharyngeal colonization in the post PCV13 era. The principal difference between serotypes 15B and 15C is the presence of an O-acetyl group on the pentasaccharide repeating unit of 15B polysaccharide. It remains unclear if antibodies against SP15B polysaccharide demonstrate functional cross reaction with SP15C strains. We compared functional activity of polyclonal rabbit anti-capsular 15B sera against SP15B and SP15C isolates. METHODS: Using flow cytometry we measured complement factors C3c and C4d deposition on SP15B and 15C in the presence of polyclonal rabbit anti capsular 15B sera. We measured the binding of C3c, common to all complement pathways, and C4d, specific to classical pathway, on SP serotypes 15B and 15C when co-incubated with polyclonal rabbit anti capsular 15B sera and antibody depleted complement. Both the proportion of bacteria with complement deposition and the fluorescence intensity were measured. We also measured agglutination as the increase in forward and side scatter. RESULTS: Polyclonal rabbit anti-capsular 15B sera activated classical pathway resulting in deposition of C4d and C3c at high intensity on all SP15B cells but only achieved limited deposition and intensity of C4 with SP15C. Similarly, polyclonal rabbit anti-capsular 15B sera induced agglutination of SP15B strains in a dose dependent manner and limited agglutination of SP15C. CONCLUSIONS: Anti-capsular 15B sera induce limited C4d and C3c deposition, and minimal agglutination with SP15C strains, reflecting lower classical pathway activation in contrast to high C4d and C3c deposition and agglutination of SP15B. These observations support limited functional cross reactivity of anti-15B to SP15C strains and are consistent with the reduction in opsonophagocytic killing of SP15C reported following immunization with vaccines containing 15B polysaccharide.

Effective regulatory responses to predatory stem cell markets in Australia and Canada.

The rapid proliferation of businesses engaged in direct-to-consumer advertising of unproven stem cell interventions has raised troubling questions about whether government bodies can regulate this health market effectively. Recent developments in Australia and Canada suggest that such fears are unfounded and that targeted regulatory action can have meaningful effects.

A 1024-Channel 268 nW/pixel 36×36 µm2/channel Data-Compressive Neural Recording IC for High-Bandwidth Brain-Computer Interfaces.

This paper presents a data-compressive neural recording IC for single-cell resolution high-bandwidth brain-computer interfaces. The IC features wired-OR lossy compression during digitization, thus preventing data deluge and massive data movement. By discarding unwanted baseline samples of the neural signals, the output data rate is reduced by 146× on average while allowing the reconstruction of spike samples. The recording array consists of pulse position modulation-based active digital pixels with a global single-slope analog-to-digital conversion scheme, which enables a low-power and compact pixel design with significantly simple routing and low array readout energy. Fabricated in a 28-nm CMOS process, the neural recording IC features 1024 channels (i.e., 32 × 32 array) with a pixel pitch of 36 µm that can be directly matched to a high-density microelectrode array. The pixel achieves 7.4 µVrms input-referred noise with a -3 dB bandwidth of 300-Hz to 5-kHz while consuming only 268 nW from a single 1-V supply. The IC achieves the smallest area per channel (36 × 36 µm2) and the highest energy efficiency among the state-of-the-art neural recording ICs published to date.

PRMT1 Modulates Alternative Splicing to Enhance HPV18 mRNA Stability and Promote the Establishment of Infection.

Only persistent HPV infections lead to the development of cancer. Thus, understanding the virus-host interplay that influences the establishment of viral infection has important implications for HPV biology and human cancers. The ability of papillomaviruses to establish in cells requires the strict temporal regulation of viral gene expression in sync with cellular differentiation. This control primarily happens at the level of RNA splicing and polyadenylation. However, the details of how this spatio-temporal regulation is achieved still need to be fully understood. Until recently, it has been challenging to study the early events of the HPV lifecycle following infection. We used a single-cell genomics approach to identify cellular factors involved in viral infection and establishment. We identify protein arginine N-methyltransferase 1 (PRMT1) as an important factor in viral infection of primary human cervical cells. PRMT1 is the main cellular enzyme responsible for asymmetric dimethylation of cellular proteins. PRMT1 is an enzyme responsible for catalyzing the methylation of arginine residues on various proteins, which influences processes such as RNA processing, transcriptional regulation, and signal transduction. In this study, we show that HPV18 infection leads to increased PRMT1 levels across the viral lifecycle. PRMT1 is critical for the establishment of a persistent infection in primary cells. Mechanistically, PRMT1 inhibition leads to a highly dysregulated viral splicing pattern. Specifically, reduced PRMT1 activity leads to intron retention and a change in the E6 and E7 expression ratio. In the absence of PRMT1, viral transcripts are destabilized and subject to degradation via the nonsense-mediated decay (NMD) pathway. These findings highlight PRMT1 as a critical regulator of the HPV18 lifecycle, particularly in RNA processing, and position it as a potential therapeutic target for persistent HPV18 infections.

Atlantic bluefin tuna tagged off Norway show extensive annual migrations, high site-fidelity and dynamic behaviour in the Atlantic Ocean and Mediterranean Sea.

Atlantic bluefin tuna (ABFT; Thunnus thynnus) is a highly migratory species. To investigate the migrations and vertical behaviours of ABFT migrating to Nordic waters, we deployed pop-up satellite archival transmitting tags on 25 ABFT off Norway (curved fork length: 228-292 cm). We obtained 16 full-year migrations, which differed between individuals, and physically recovered 13 tags, which provided 4699 days of archival depth and temperature data. ABFT occupied waters from the Arctic Circle to as far south as Cabo Verde, Africa, and occupied depths down to 1190 m and temperatures from 0.5 to 27.8°C. During their annual migrations, ABFT spent, on average, 68 days in Norwegian waters, 65 days in the Newfoundland Basin, 35 days around the Canary Islands and 33 days in the West European Basin. Most ABFT entered the Mediterranean Sea with a mean entry date of 13 May and visited known spawning grounds, staying, on average, 44 days. All ABFT with full-year deployments returned to Norwegian waters. ABFT displayed high site-fidelity and dynamic vertical diving behaviours that varied between hotspots and seasons. These spatiotemporal data provide important ecological knowledge for sustainable management and the conservation of the recently recovered eastern ABFT stock.

Closed-Loop Therapy and Sleep in Young People Newly Diagnosed With T1D and Their Parents.

BACKGROUND: A diagnosis of type 1 diabetes in a young person can create vulnerability for sleep. Historically it has been rare for young people to be offered a closed-loop system soon after diagnosis meaning that studies examining sleep under these circumstances in comparison with standard treatment have not been possible. In this study, we examine sleep in young people (and their parents) who were provided with hybrid closed-loop therapy at diagnosis of type 1 diabetes versus those who receive standard treatment over a 2-year period. METHODS: The sample comprised 97 participants (mean age = 12.0 years; SD = 1.7) from a multicenter, open-label, randomized, parallel trial, where young people were randomized to either hybrid closed-loop insulin delivery or standard care at diagnosis. Sleep was measured using actigraphy and the Pittsburgh Sleep Quality Index (PSQI) in the young people, and using the PSQI in parents. RESULTS: Sleep in young people using hybrid closed-loop insulin delivery did not differ significantly compared with those receiving standard care (although there were nonsignificant trends for better sleep in the closed-loop group for 4 of the 5 sleep actigraphy measures and PSQI). Similarly, there were nonsignificant differences for sleep between the groups at 24 months (with mixed direction of effects). CONCLUSIONS: This study assessed for the first time sleep in young people using a closed-loop system soon after diagnosis. Although sleep was not significantly different for young people using closed-loop insulin delivery as compared with those receiving standard care, the direction of effects of the nonsignificant results indicates a possible tendency for better sleep quality in the hybrid closed-loop insulin delivery group at the beginning of the treatment.

Occurrence and predictors of laboratory abnormalities during outpatient parenteral antimicrobial therapy - A multicenter cohort study to inform laboratory test monitoring.

OBJECTIVES: Evidence on the optimal frequency of laboratory testing during outpatient parenteral antimicrobial therapy (OPAT) is lacking. Therefore, we investigated how often and when laboratory abnormalities occur during OPAT and which factors are associated with these abnormalities. METHODS: We performed a multicenter cohort study in four Dutch hospitals among adult patients receiving OPAT and collected routinely obtained laboratory test results. Incidence and incidence rates were calculated for various laboratory abnormalities. Survival analysis was performed to visualize the time to the first occurrence of laboratory abnormalities and Poisson regression analysis to compare the number of abnormalities in the first and second 30 OPAT days among patients receiving OPAT for ≥60 days. Predictors were identified using a multivariable Cox proportional hazard regression model. RESULTS: 45.1% of 1152 included patients developed laboratory abnormalities, but only 2% led to OPAT discontinuation. Hepatotoxicity was most common (33.9 events/1000 OPAT days), with a time-dependent decrease in the occurrence of the first hepatotoxic event, while hypokalemia was rare (1.7 events/1000 OPAT days). In the subgroup of patients receiving ≥60 days of OPAT, nephrotoxicity was more common in days 31-60. We observed partly toxicity-specific associations between antibiotic type, concomitant medication, baseline laboratory values, patient characteristics, and the occurrence of laboratory abnormalities. CONCLUSIONS: While laboratory abnormalities are frequently observed during OPAT, they rarely lead to discontinuation of OPAT. Specific patient, treatment and laboratory characteristics were associated with the occurrence of laboratory abnormalities. Based on our results, we recommend a more personalized laboratory monitoring policy with less blood sampling.

Molecular mechanisms of Coxiella burnetii formalin-fixed cellular vaccine reactogenicity.

Local and systemic reactogenic responses to Q-VAX have prevented licensing of this vaccine outside of Australia. These reactogenic responses occur in previously sensitized individuals and have not been well defined at the cellular level, in part because many studies have been done in guinea pigs that have limited molecular tools. We previously characterized a mouse model of reactogenicity where local reaction sites showed an influx of CD8+ and IFNγ-expressing IL17a+ CD4+ T cells consistent with a Th1 delayed-type hypersensitivity. In this study, we determined, using depletion and adoptive transfer experiments, that both anti-Coxiella antibodies and CD4+ T cells were essential for localized reactions at the site of vaccination. Furthermore, IFNγ depletion showed significant histological changes at the local reaction sites demonstrating the essential nature of this cytokine to reactogenicity. In addition to the cells and cytokines required for this response, we determined that whole cell vaccine (WCV) material remained at the site of vaccination for at least 26 weeks post-injection. Transmission electron microscopy (TEM) of these sites demonstrated intact rod-shaped bacteria at 2 weeks post-injection and partially degraded bacteria within macrophages at 26 weeks post-injection. Finally, because small cell variants (SCVs) are an environmentally stable form, we determined that local reactions were more severe when the WCV material was prepared with higher levels of SCVs compared to typical WCV or with higher levels of large cell variant (LCV). These studies support the hypothesis that antigen persistence at the site of injection contributes to this reactogenicity and that anti-Coxiella antibodies, CD4+ T cells, and IFNγ each contribute to this process.

Successful Delayed Amputated Split-Thickness Dermis Reattachment of a Fingertip Injury in the Home Setting without Sutures.

Presentation: This case reports details a fingertip amputation injury. The patient was vitally stable post-injury and blood loss was controlled with direct pressure. Diagnosis: The injury was inspected and found to involve the finger pulp and nailbed, without exposure of the terminal phalanx (Allen 2). Treatment: The avulsed tissue was initially placed in situ at the site of the injury. At day 3 the viable dermis from the avulsed tissue was dissected away and a split-thickness dermal graft was performed. The graft was held in place with antibacterial dressings. Epithelialisation was complete at two weeks, sensation returned to normal at five months and progress was tracked with interval photography. Overall there was an excellent cosmetic and functional outcome. Discussion: Split-thickness grafting of the dermis has been previously described, but there are no reports of this technique being applied to fingertip injuries. De-epithelialisation may enhance the likelihood of graft survival when compared to composite grafting techniques.

Structural basis of thiamine transport and drug recognition by SLC19A3.

Thiamine (vitamin B1) functions as an essential coenzyme in cells. Humans and other mammals cannot synthesise this vitamin de novo and thus have to take it up from their diet. Eventually, every cell needs to import thiamine across its plasma membrane, which is mainly mediated by the two specific thiamine transporters SLC19A2 and SLC19A3. Loss of function mutations in either of these transporters lead to detrimental, life-threatening metabolic disorders. SLC19A3 is furthermore a major site of drug interactions. Many medications, including antidepressants, antibiotics and chemotherapeutics are known to inhibit this transporter, with potentially fatal consequences for patients. Despite a thorough functional characterisation over the past two decades, the structural basis of its transport mechanism and drug interactions has remained elusive. Here, we report seven cryo-electron microscopy (cryo-EM) structures of the human thiamine transporter SLC19A3 in complex with various ligands. Conformation-specific nanobodies enable us to capture different states of SLC19A3's transport cycle, revealing the molecular details of thiamine recognition and transport. We identify seven previously unknown drug interactions of SLC19A3 and present structures of the transporter in complex with the inhibitors fedratinib, amprolium and hydroxychloroquine. These data allow us to develop an understanding of the transport mechanism and ligand recognition of SLC19A3.

Adiabatic light guide with S-shaped strips.

A light guide is an essential part of many scintillator counters and light collection systems. Our main interest is a light guide for a thin, wide scintillator that has high light transmission while converting the area of the light source to the shape of a photo-detector. We propose a variation of the light guide that avoids a 90° twist of the strips, reduces the length of the light pipe, and reduces the complexity of production. Detailed Monte Carlo simulation studies have been performed for a three-strip S-shaped light-guide system.

Considering best practice standards for routine whole-genome sequencing for TB care and control.

TB is a priority pathogen for the application of whole-genome sequencing (WGS) into routine public health practice. In low-incidence settings, a growing number of services have begun to incorporate routine WGS into standard practice. The increasing availability of real-time genomic information supports a variety of aspects of the public health response, including the detection of drug resistance, monitoring of laboratory and clinical practices, contact tracing investigations and active case finding. Optimal structures and approaches are needed to support the rapid translation of genomic information into practice and to evaluate outcomes and impact. In this consensus paper, we outline the elements needed to systemically incorporate routine WGS into the TB public health response, including the sustainability of services, multidisciplinary team models and monitoring and evaluation frameworks. If integrated in an efficient and thoughtful manner, routine WGS has the potential to significantly improve clinical TB care for individuals and the overall public health response.

La TB est un agent pathogène prioritaire pour l'application du séquençage du génome entier (WGS, pour l'anglais « whole-genome sequencing ¼) dans les pratiques courantes de santé publique. Dans des contextes à faible incidence, un nombre croissant de services ont commencé à intégrer le WGS de manière routinière dans la pratique standard. La disponibilité croissante de l'information génomique en temps réel soutient divers aspects de l'intervention de santé publique, notamment la détection de la résistance aux médicaments, la surveillance des pratiques de laboratoire et cliniques, les enquêtes de recherche des contacts et la recherche active des cas. Des structures et des approches optimales sont nécessaires pour soutenir l'application rapide de l'information génomique dans la pratique et pour évaluer les résultats et l'impact. Dans ce document de consensus, nous décrivons les éléments nécessaires à l'intégration systématique du WGS dans la riposte de santé publique à la TB, notamment la durabilité des services, les modèles d'équipe multidisciplinaire et les cadres de suivi et d'évaluation. S'il est intégré de manière efficace et réfléchie, le WGS de routine a le potentiel d'améliorer considérablement les soins cliniques de la TB pour les individus et la réponse globale de la santé publique.

Allogeneic mesenchymal stromal cell therapy in kidney transplantation: should repeated human leukocyte antigen mismatches be avoided?

Mesenchymal stromal cells (MSCs) have immunomodulatory properties and are therefore considered promising tools in kidney transplantation. Although most studies have been conducted with autologous MSCs, using allogeneic MSCs as an off-the-shelf product is more feasible in clinical settings. However, allogeneic MSCs could potentially induce an immune response, which might eventually be directed towards the kidney allograft because of shared human leukocyte antigen (HLA) epitope mismatches between the kidney and MSC donor. In this study, we performed in-depth analyses of two cohorts (n = 20) that received third-party MSC therapy after kidney transplantation. While the Neptune Study from Leiden University Medical Center specifically selected MSC to avoid repeated HLA antigen mismatches between kidney and MSC donors, the study from the University of Liège did not perform specific MSC selection. The comparative analyses of amino acid mismatches between these cohorts showed that MSC selection to avoid repeated HLA mismatches at the split antigen level was not sufficient to prevent repeated mismatches at the amino acid level. However, repeated amino acid mismatches were not associated with the occurrence of donor-specific antibodies (DSAs). Thus, the clinical relevance of repeated amino acid mismatches seems to be limited with regard to the risk of DSA formation. Since DSA formation was limited (3 of 20 patients) in this study, larger studies are required to investigate the relevance of preventing repeated HLA mismatches in allogeneic MSC therapy in kidney transplantation.

Effectiveness of a Person-Centered Interdisciplinary Rehabilitation Treatment of Post-COVID-19 Condition: Protocol for a Single-Case Experimental Design Study.

BACKGROUND: Patients with post-COVID-19 condition (PCC) experience a wide range of complaints (physical, cognitive, and mental), sometimes with high levels of disability in daily activities. Evidence of effective interdisciplinary rehabilitation treatment is lacking. A person-centered, biopsychosocial, interdisciplinary rehabilitation program, adapted to expert opinions and the patient's needs, was therefore developed. OBJECTIVE: This study aims to present a study protocol for a clinical trial to evaluate the effect of a new, person-centered, interdisciplinary rehabilitation treatment for PCC. It is aimed at improving participation in society and health-related quality of life in patients with PCC who perceive a high level of disability in daily activities or participation. METHODS: A total of 20 Dutch adults, aged 18 years or older, with high levels of disability in daily activities and participation in society will be included in this replicated and randomized single-case experimental design study, from October 2023 onward. The replicated and randomized single-case experimental design consists of 3 phases. The baseline phase is the observational period, in which no specific treatment will be given. In the intervention phase, patients will receive the new outpatient treatment 3 times a week for 12 weeks, followed by a 12-week follow-up phase. During the intervention phase, the treatment will be personalized according to the patient's physical, mental, and cognitive symptoms and goals. The treatment team can consist of a rehabilitation physician, physiotherapist, occupational therapist, speech therapist, and psychologist. The primary outcomes of the study are daily diaries, which consist of 8 questions selected from validated questionnaires (Utrecht Scale for Evaluation of Rehabilitation-Participation, EQ-5D-5L, and the Hospital Anxiety and Depression Scale). The other primary outcome measurements are participation in society (Utrecht Scale for Evaluation of Rehabilitation-Participation) and health-related quality of life (EQ-5D-5L). The secondary outcomes are physical tests and validated questionnaires aimed at physical, mental, and cognitive complaints. Effect evaluation based on daily assessments will include visual analysis, calculation of effect sizes (Nonoverlap of All Pairs), randomization tests, and multilevel analysis. In addition, other analyses will be based on ANOVA or a 2-tailed Student t test. RESULTS: Data collection for this study started in October 2023 and is planned to be completed in July 2024. The results will be published in peer-reviewed journals and presented at international conferences. CONCLUSIONS: This is the first study investigating the effect of an interdisciplinary rehabilitation treatment with a person-centered, biopsychosocial approach in patients with PCC. Our findings will help to improve the treatment and support of patients with PCC. TRIAL REGISTRATION: German Clinical Trials Register DRKS00032636; https://drks.de/search/en/trial/DRKS00032636. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): DERR1-10.2196/63951.

A service evaluation of weight management for glycaemic control and remission of type 2 diabetes using traditional food in Nepal (Ho-DIRECT NEPAL): a single-arm trial.

Background: Remission of early type 2 diabetes (T2D) is possible; however, diet programmes proven effective are unaffordable in many southeast Asian populations where T2D is more frequent and more aggressive at lower body weight and younger age. We evaluate an entirely food-based service. Methods: This study employed a single-arm intervention and follow-up design for intervention evaluation in existing hospital people with T2D of under 5 years known duration. Individuals attending a diabetes clinic in Kathmandu with early T2D (<5 years) aged 30-70 years, BMI ≥23 kg/m2, were offered a low-cost nutritionally complete diet-programme, using traditional Nepali foods to provide 8-weeks ∼850 kcal/day weight loss induction, and then weight maintenance. The participants received 4-weekly dietetic appointments (30-45 min) and verbo-pictorial leaflets using household measures. Glucose-lowering medications (49/70 at baseline) were stopped at baseline or soon after. The study was registered as ISRCTN10671396, testing a traditional food-based intervention for weight loss and T2D remission. Findings: For 70 individuals (45 female) invited between March 19, 2022 and September 19, 2023, baseline mean (SD) age was 48.6 (9.9) years, bodyweight 74.6 (9.5) kg, BMI 29.7 (3.6) kg/m2, known diabetes duration 2.5 (1.9) years, HbA1c on treatment 8.1 (1.6) %. At 12, 24 and 52 weeks respectively, evaluating n = 44, 46, 45, bodyweight was 70.1 (8.5), 69.8 (8.9), 70.0 (8.8) kg, HbA1c 6.8 (0.9), 6.9 (1.5), 7.1 (1.3) %; HbA1c <6.5% was recorded for 46%, 48% and 36% and remission of T2D (HbA1c <6.5% off medication >3 months) in 43%, 39% and 29%. The main reported adherence barriers were fears of weakness, hunger, and inconvenience during travel. Incentives were ease of the diet, reduced doses and costs of medications, and improved appearance. Interpretation: Traditional food-based weight management can valuably improve control, reduce medication needs, and generate remissions of established T2D, but adherence barriers must be overcome to optimise outcomes. Funding: All Saints Educational Trust, England.

Maternal Complications After Laser Surgery for Twin-to-Twin Transfusion Syndrome, a Cohort Study.

OBJECTIVES: To assess maternal complications after fetoscopic laser surgery (FLS) for the twin-to-twin transfusion syndrome (TTTS). METHODS: All consecutive cases treated with FLS for TTTS between 2008 and 2021 at the Leiden University Medical Center (LUMC) were included. We allocated complications in three timeframes: "Admission for laser surgery," "pregnancy after laser," and "delivery and third stage of labor." Maternal complications were graded according to the Maternal and Fetal Adverse Event Terminology (MFAET) and for intra-abdominal hemorrhage, the Common Terminology Criteria for Adverse Events (CTCAE). RESULTS: In the study period, 637 mothers were treated for TTTS with FLS. There were 1559 occurrences of maternal complications. The rate of severe maternal complications (grade 3 or 4) was 8.0%. Severe complications consisted of six cases of severe intra-abdominal hemorrhage, nine cases of severe hemorrhage in pregnancy, one with severe chorioamnionitis, 10 with severe preeclampsia/HELLP syndrome, and 25 with a severe postpartum hemorrhage. CONCLUSIONS: Even though it is the gold standard for treating TTTS, FLS comes at a risk to the mother which should not be neglected. And even though not all complications have serious consequences to the mother, the severe maternal complication rate of 8.0% should be added to the inherent risks for the fetus, and should be discussed with patients eligible for surgery in order to make an informed decision on treatment options.