Structural basis of sodium-dependent bile salt uptake into the liver.

The liver takes up bile salts from blood to generate bile, enabling absorption of lipophilic nutrients and excretion of metabolites and drugs1. Human Na+-taurocholate co-transporting polypeptide (NTCP) is the main bile salt uptake system in liver. NTCP is also the cellular entry receptor of human hepatitis B and D viruses2,3 (HBV/HDV), and has emerged as an important target for antiviral drugs4. However, the molecular mechanisms underlying NTCP transport and viral receptor functions remain incompletely understood. Here we present cryo-electron microscopy structures of human NTCP in complexes with nanobodies, revealing key conformations of its transport cycle. NTCP undergoes a conformational transition opening a wide transmembrane pore that serves as the transport pathway for bile salts, and exposes key determinant residues for HBV/HDV binding to the outside of the cell. A nanobody that stabilizes pore closure and inward-facing states impairs recognition of the HBV/HDV receptor-binding domain preS1, demonstrating binding selectivity of the viruses for open-to-outside over inward-facing conformations of the NTCP transport cycle. These results provide molecular insights into NTCP 'gated-pore' transport and HBV/HDV receptor recognition mechanisms, and are expected to help with development of liver disease therapies targeting NTCP.

The ion-coupling mechanism of human excitatory amino acid transporters.

Excitatory amino acid transporters (EAATs) maintain glutamate gradients in the brain essential for neurotransmission and to prevent neuronal death. They use ionic gradients as energy source and co-transport transmitter into the cytoplasm with Na+ and H+ , while counter-transporting K+ to re-initiate the transport cycle. However, the molecular mechanisms underlying ion-coupled transport remain incompletely understood. Here, we present 3D X-ray crystallographic and cryo-EM structures, as well as thermodynamic analysis of human EAAT1 in different ion bound conformations, including elusive counter-transport ion bound states. Binding energies of Na+ and H+ , and unexpectedly Ca2+ , are coupled to neurotransmitter binding. Ca2+ competes for a conserved Na+ site, suggesting a regulatory role for Ca2+ in glutamate transport at the synapse, while H+ binds to a conserved glutamate residue stabilizing substrate occlusion. The counter-transported ion binding site overlaps with that of glutamate, revealing the K+ -based mechanism to exclude the transmitter during the transport cycle and to prevent its neurotoxic release on the extracellular side.

Structure and allosteric inhibition of excitatory amino acid transporter 1.

Human members of the solute carrier 1 (SLC1) family of transporters take up excitatory neurotransmitters in the brain and amino acids in peripheral organs. Dysregulation of the function of SLC1 transporters is associated with neurodegenerative disorders and cancer. Here we present crystal structures of a thermostabilized human SLC1 transporter, the excitatory amino acid transporter 1 (EAAT1), with and without allosteric and competitive inhibitors bound. The structures reveal architectural features of the human transporters, such as intra- and extracellular domains that have potential roles in transport function, regulation by lipids and post-translational modifications. The coordination of the allosteric inhibitor in the structures and the change in the transporter dynamics measured by hydrogen-deuterium exchange mass spectrometry reveal a mechanism of inhibition, in which the transporter is locked in the outward-facing states of the transport cycle. Our results provide insights into the molecular mechanisms underlying the function and pharmacology of human SLC1 transporters.

Monophonic and Polyphonic Wheezing Classification Based on Constrained Low-Rank Non-Negative Matrix Factorization.

The appearance of wheezing sounds is widely considered by physicians as a key indicator to detect early pulmonary disorders or even the severity associated with respiratory diseases, as occurs in the case of asthma and chronic obstructive pulmonary disease. From a physician's point of view, monophonic and polyphonic wheezing classification is still a challenging topic in biomedical signal processing since both types of wheezes are sinusoidal in nature. Unlike most of the classification algorithms in which interference caused by normal respiratory sounds is not addressed in depth, our first contribution proposes a novel Constrained Low-Rank Non-negative Matrix Factorization (CL-RNMF) approach, never applied to classification of wheezing as far as the authors' knowledge, which incorporates several constraints (sparseness and smoothness) and a low-rank configuration to extract the wheezing spectral content, minimizing the acoustic interference from normal respiratory sounds. The second contribution automatically analyzes the harmonic structure of the energy distribution associated with the estimated wheezing spectrogram to classify the type of wheezing. Experimental results report that: (i) the proposed method outperforms the most recent and relevant state-of-the-art wheezing classification method by approximately 8% in accuracy; (ii) unlike state-of-the-art methods based on classifiers, the proposed method uses an unsupervised approach that does not require any training.

Wheezing Sound Separation Based on Informed Inter-Segment Non-Negative Matrix Partial Co-Factorization.

Wheezing reveals important cues that can be useful in alerting about respiratory disorders, such as Chronic Obstructive Pulmonary Disease. Early detection of wheezing through auscultation will allow the physician to be aware of the existence of the respiratory disorder in its early stage, thus minimizing the damage the disorder can cause to the subject, especially in low-income and middle-income countries. The proposed method presents an extended version of Non-negative Matrix Partial Co-Factorization (NMPCF) that eliminates most of the acoustic interference caused by normal respiratory sounds while preserving the wheezing content needed by the physician to make a reliable diagnosis of the subject's airway status. This extension, called Informed Inter-Segment NMPCF (IIS-NMPCF), attempts to overcome the drawback of the conventional NMPCF that treats all segments of the spectrogram equally, adding greater importance for signal reconstruction of repetitive sound events to those segments where wheezing sounds have not been detected. Specifically, IIS-NMPCF is based on a bases sharing process in which inter-segment information, informed by a wheezing detection system, is incorporated into the factorization to reconstruct a more accurate modelling of normal respiratory sounds. Results demonstrate the significant improvement obtained in the wheezing sound quality by IIS-NMPCF compared to the conventional NMPCF for all the Signal-to-Noise Ratio (SNR) scenarios evaluated, specifically, an SDR, SIR and SAR improvement equals 5.8 dB, 4.9 dB and 7.5 dB evaluating a noisy scenario with SNR = -5 dB.

Smart Containers Schedulers for Microservices Provision in Cloud-Fog-IoT Networks. Challenges and Opportunities.

Docker containers are the lightweight-virtualization technology prevailing today for the provision of microservices. This work raises and discusses two main challenges in Docker containers' scheduling in cloud-fog-internet of things (IoT) networks. First, the convenience to integrate intelligent containers' schedulers based on soft-computing in the dominant open-source containers' management platforms: Docker Swarm, Google Kubernetes and Apache Mesos. Secondly, the need for specific intelligent containers' schedulers for the different interfaces in cloud-fog-IoT networks: cloud-to-fog, fog-to-IoT and cloud-to-fog. The goal of this work is to support the optimal allocation of microservices provided by the main cloud service providers today and used by millions of users worldwide in applications such as smart health, content delivery networks, smart health, etc. Particularly, the improvement is studied in terms of quality of service (QoS) parameters such as latency, load balance, energy consumption and runtime, based on the analysis of previous works and implementations. Moreover, the scientific-technical impact of smart containers' scheduling in the market is also discussed, showing the possible repercussion of the raised opportunities in the research line.

Systematic study of the cross polarization introduced by broadband antireflection layers at microwave frequencies.

Implementation of antireflection layers using structured materials is of common use in millimeter- and submillimeter-wave refractive optic systems. In this work we have systematically studied the effect of such structures in the optical propagation with special emphasis on the cross polarization they introduce. We have performed extensive simulations and experimental verification of several commonly used structures: concentric grooves, parallel grooves, an array of boxes, an array of cylinders, and rectangular- versus triangular-shaped grooves. As a result, we propose optimal structures for demanding applications in terms of polarization and return losses over large fractional bandwidths.

Transport mechanism of a bacterial homologue of glutamate transporters.

Glutamate transporters are integral membrane proteins that catalyse a thermodynamically uphill uptake of the neurotransmitter glutamate from the synaptic cleft into the cytoplasm of glia and neuronal cells by harnessing the energy of pre-existing electrochemical gradients of ions. Crucial to the reaction is the conformational transition of the transporters between outward and inward facing states, in which the substrate binding sites are accessible from the extracellular space and the cytoplasm, respectively. Here we describe the crystal structure of a double cysteine mutant of a glutamate transporter homologue from Pyrococcus horikoshii, Glt(Ph), which is trapped in the inward facing state by cysteine crosslinking. Together with the previously determined crystal structures of Glt(Ph) in the outward facing state, the structure of the crosslinked mutant allows us to propose a molecular mechanism by which Glt(Ph) and, by analogy, mammalian glutamate transporters mediate sodium-coupled substrate uptake.

Medium-term effects of septal and apical pacing in pacemaker-dependent patients: a double-blind prospective randomized study.

BACKGROUND: Pacing the right ventricle is established practice, but there remains controversy as to the optimal site to preserve hemodynamic function. AIMS: To evaluate clinical and hemodynamic differences between apical and septal pacing in pacemaker-dependent patients. METHODS: Patients receiving their first pacemaker for advanced atrioventricular block, with the atria in sinus rhythm, were randomized to receive apical (Group A) or septal (Group S) ventricular leads. After implant, with the device programmed VVI 70 beats/min fixed rate, patients underwent a 6-minute walk test and a transthoracic echocardiogram. Then, DDDR was programmed at nominal settings. The same tests were performed at 6 months and 12 months follow-up. If ventricular pacing was less than 98%, the patient was excluded. RESULTS: A total of 142 patients were included in the study. During the study year, 71 (50%) were excluded for not fulfilling the condition of 98% ventricular pacing. Groups A and S had 34 and 37 patients, respectively. Age and gender were similar in the groups. At implant, QRS duration was significantly greater in Group A (158 ms) than Group S (146 ms; P = 0.018), and the QRS axis was different: -74.5° in Group A and 1° in Group S (P < 0.001). At 1 year, the 6-minute walk improved significantly in both groups: Group A 15% (P = 0.048) and Group S 24% (P = 0.001). Left ventricular ejection fraction (LVEF) increased from 0.57 to 0.61 (P = 0.008) in Group S, without significant change in Group A. CONCLUSIONS: After 1 year, pacemaker-dependent patients with septal ventricular leads have better clinical and functional (LVEF) outcome.

The ion pathway through the opened Na(+),K(+)-ATPase pump.

P-type ATPases pump ions across membranes, generating steep electrochemical gradients that are essential for the function of all cells. Access to the ion-binding sites within the pumps alternates between the two sides of the membrane to avoid the dissipation of the gradients that would occur during simultaneous access. In Na(+),K(+)-ATPase pumps treated with the marine agent palytoxin, this strict alternation is disrupted and binding sites are sometimes simultaneously accessible from both sides of the membrane, transforming the pumps into ion channels (see, for example, refs 2, 3). Current recordings in these channels can monitor accessibility of introduced cysteine residues to water-soluble sulphydryl-specific reagents. We found previously that Na(+),K(+) pump-channels open to the extracellular surface through a deep and wide vestibule that emanates from a narrower pathway between transmembrane helices 4 and 6 (TM4 and TM6). Here we report that cysteine scans from TM1 to TM6 reveal a single unbroken cation pathway that traverses palytoxin-bound Na(+),K(+) pump-channels from one side of the membrane to the other. This pathway comprises residues from TM1, TM2, TM4 and TM6, passes through ion-binding site II, and is probably conserved in structurally and evolutionarily related P-type pumps, such as sarcoplasmic- and endoplasmic-reticulum Ca(2+)-ATPases and H(+),K(+)-ATPases.

Receptor-recognition and antiviral mechanisms of retrovirus-derived human proteins.

Human syncytin-1 and suppressyn are cellular proteins of retroviral origin involved in cell-cell fusion events to establish the maternal-fetal interface in the placenta. In cell culture, they restrict infections from members of the largest interference group of vertebrate retroviruses, and are regarded as host immunity factors expressed during development. At the core of the syncytin-1 and suppressyn functions are poorly understood mechanisms to recognize a common cellular receptor, the membrane transporter ASCT2. Here, we present cryo-electron microscopy structures of human ASCT2 in complexes with the receptor-binding domains of syncytin-1 and suppressyn. Despite their evolutionary divergence, the two placental proteins occupy similar positions in ASCT2, and are stabilized by the formation of a hybrid ß-sheet or 'clamp' with the receptor. Structural predictions of the receptor-binding domains of extant retroviruses indicate overlapping binding interfaces and clamping sites with ASCT2, revealing a competition mechanism between the placental proteins and the retroviruses. Our work uncovers a common ASCT2 recognition mechanism by a large group of endogenous and disease-causing retroviruses, and provides high-resolution views on how placental human proteins exert morphological and immunological functions.

Lack of current implantable cardioverter defibrillator guidelines application for primary prevention of sudden cardiac death in Latin American patients with heart failure: a cross-sectional study.

AIMS: This cross-sectional study evaluated the application of accepted international implantable cardioverter defibrillator (ICD) guidelines for primary prevention of sudden cardiac death in patients with heart failure. METHODS AND RESULTS: The PLASMA (Probabilidad de Sufrir Muerte Arritmica) study was designed to characterize management of cardiac patients in Latin America. Twelve centres included 1958 consecutively admitted patients in cardiology units in 2008 and 2009. Discharged patients were evaluated for primary prevention, ICD indication and prescription by general cardiologists. Of 1711 discharged patients, 1525 (89%) had data available for evaluating indication status. Class I indications for ICD therapy were met for 153 (10%) patients based on collected data. Only 20 (13%, 95% confidence interval: 7.7-18.4%) patients with indication were prescribed an ICD. Patients prescribed an ICD were younger than patients who were not prescribed an ICD (62 vs. 68 years, P < 0.01). The reasons given by cardiologists for not prescribing an ICD for 133 patients with an indication were: indication criteria not met (75%), life expectancy <1 year (9.7%), rejection by the patient (5.2%), no medical coverage paying for the device (3.7%), psychiatric patient (2.2%), and other reasons (4.2%). CONCLUSIONS: In Latin America, international guidelines for primary prevention ICD implantation are not well followed. The main reason is that cardiologists believe that patients do not meet indication criteria, even though study data confirm that criteria are met. This poses a significant challenge and underlines the importance of continuous and improved medical education.

Termination of Cardiopulmonary Resuscitation in Mountain Rescue: A Scoping Review and ICAR MedCom 2023 Recommendations.

Lugnet, Viktor, Miles McDonough, Les Gordon, Mercedes Galindez, Nicolas Mena Reyes, Alison Sheets, Ken Zafren, and Peter Paal. Termination of cardiopulmonary resuscitation in mountain rescue: a scoping review and ICAR MedCom 2023 recommendations. High Alt Med Biol. 24:274-286, 2023. Background: In 2012, the International Commission for Mountain Emergency Medicine (ICAR MedCom) published recommendations for termination of cardiopulmonary resuscitation (CPR) in mountain rescue. New developments have necessitated an update. This is the 2023 update for termination of CPR in mountain rescue. Methods: For this scoping review, we searched the PubMed and Cochrane libraries, updated the recommendations, and obtained consensus approval within the writing group and the ICAR MedCom. Results: We screened a total of 9,102 articles, of which 120 articles met the inclusion criteria. We developed 17 recommendations graded according to the strength of recommendation and level of evidence. Conclusions: Most of the recommendations from 2012 are still valid. We made minor changes regarding the safety of rescuers and responses to primary or traumatic cardiac arrest. The criteria for termination of CPR remain unchanged. The principal changes include updated recommendations for mechanical chest compression, point of care ultrasound (POCUS), extracorporeal life support (ECLS) for hypothermia, the effects of water temperature in drowning, and the use of burial times in avalanche rescue.

Ion permeation through the Na+,K+-ATPase.

P-type ATPase pumps generate concentration gradients of cations across membranes in nearly all cells. They provide a polar transmembrane pathway, to which access is strictly controlled by coupled gates that are constrained to open alternately, thereby enabling thermodynamically uphill ion transport (for example, see ref. 1). Here we examine the ion pathway through the Na+,K+-ATPase, a representative P-type pump, after uncoupling its extra- and intracellular gates with the marine toxin palytoxin. We use small hydrophilic thiol-specific reagents as extracellular probes and we monitor their reactions, and the consequences, with cysteine residues introduced along the anticipated cation pathway through the pump. The distinct effects of differently charged reagents indicate that a wide outer vestibule penetrates deep into the Na+,K+-ATPase, where the pathway narrows and leads to a charge-selectivity filter. Acidic residues in this region, which are conserved to coordinate pumped ions, allow the approach of cations but exclude anions. Reversing the charge at just one of those positions converts the pathway from cation selective to anion selective. Close structural homology among the catalytic subunits of Ca2+-, Na+,K+- and H+,K+-ATPases argues that their extracytosolic cation exchange pathways all share these physical characteristics.

Structural insights into the lysophospholipid brain uptake mechanism and its inhibition by syncytin-2.

Brain development and function require uptake of essential omega-3 fatty acids in the form of lysophosphatidylcholine via major-facilitator superfamily transporter MFSD2A, a potential pharmaceutical target to modulate blood-brain barrier (BBB) permeability. MFSD2A is also the receptor of endogenous retroviral envelope syncytin-2 (SYNC2) in human placenta, where it mediates cell-cell fusion and formation of the maternal-fetal interface. Here, we report a cryo-electron microscopy structure of the human MFSD2A-SYNC2 complex that reveals a large hydrophobic cavity in the transporter C-terminal domain to occlude long aliphatic chains. The transporter architecture suggests an alternating-access transport mechanism for lipid substrates in mammalian MFS transporters. SYNC2 establishes an extensive binding interface with MFSD2A, and a SYNC2-soluble fragment acts as a long-sought-after inhibitor of MFSD2A transport. Our work uncovers molecular mechanisms important to brain and placenta development and function, and SYNC2-mediated inhibition of MFSD2A transport suggests strategies to aid delivery of therapeutic macromolecules across the BBB.

Fibrin(ogen) Is Constitutively Expressed by Differentiated Intestinal Epithelial Cells and Mediates Wound Healing.

Fibrinogen is a large molecule synthesized in the liver and released in the blood. Circulating levels of fibrinogen are upregulated after bleeding or clotting events and support wound healing. In the context of an injury, thrombin activation drives conversion of fibrinogen to fibrin. Fibrin deposition contains tissue damage, stops blood loss, and prevents microbial infection. In most circumstances, fibrin needs to be removed to allow the resolution of inflammation and tissue repair, whereas failure of this may lead to the development of various disorders. However, the contribution of fibrinogen to tissue inflammation and repair is likely to be context-dependent. In this study, the concept that fibrin needs to be removed to allow tissue repair and to reduce inflammation is challenged by our observations that, in the intestine, fibrinogen is constitutively produced by a subset of intestinal epithelial cells and deposited at the basement membrane as fibrin where it serves as a substrate for wound healing under physiological conditions such as epithelial shedding at the tip of the small intestinal villus and surface epithelium of the colon as well as under pathological conditions that require rapid epithelial repair. The functional integrity of the intestine is ensured by the constant renewal of its simple epithelium. Superficial denuding of the epithelial cell layer occurs regularly and is rapidly corrected by a process called restitution that can be influenced by various soluble and insoluble factors. Epithelial cell interaction with the extracellular matrix greatly influences the healing process by acting on cell morphology, adhesion, and migration. The functional contribution of a fibrin(ogen) matrix in the intestine was studied under physiological and pathological contexts. Our results (immunofluorescence, immunoelectron microscopy, and quantitative PCR) show that fibrin(ogen) is a novel component of the basement membrane associated with the differentiated epithelial cell population in both the small intestine and colon. Fibrin(ogen) alone is a weak ligand for epithelial cells and behaves as an anti-adhesive molecule in the presence of type I collagen. Furthermore, the presence of fibrin(ogen) significantly shortens the time required to achieve closure of wounded epithelial cell monolayers and co-cultures in a PI3K-dependent manner. In human specimens with Crohn's disease, we observed a major accumulation of fibrin(ogen) throughout the tissue and at denuded sites. In mice in which fibrin formation was inhibited with dabigatran treatment, dextran sulfate sodium administration provoked a significant increase in the disease activity index and pathological features such as mucosal ulceration and crypt abscess formation. Taken together, these results suggest that fibrin(ogen) contributes to epithelial healing under both normal and pathological conditions.

Robot-Assisted Unicompartmental Knee Arthroplasty: Increasing Surgical Accuracy? A Cadaveric Study.

Unicompartmental knee arthroplasty (UKA) represents 10% of knee arthroplasties. Advantages are better functional results, quicker recovery, shorter hospitalization time, and lower blood loss, among others. However, revision rates are larger than total knee arthroplasty. Among the most important factors that explain this are the implant position and alignment, and the correct surgical indication. Greater accuracy in the implant placement may improve clinical results and increase the rate of implant survival. The objective of this study is to evaluate the precision of the Navio robot-assisted system in the position and alignment of medial UKA compared with the conventional technique. This is an experimental pilot study. Twenty-six cadaveric models were randomized into 2 groups: Robot-Assisted surgery (R) and Conventional Surgery (C). Radiological study was performed pre- and post-surgery, evaluating the medial distal femoral angle (MDFA), medial proximal tibial angle (MPTA), tibial slope, tibiofemoral angle (TFA), sagittal femoral angle (SFA), and size of the femoral and tibial components. The main result measurement was the change in postoperative angulation. The results of this study are MDFA median of 1.07° (0.19-4.5) for group R and 0.12° (0.03-10.4) with a significant difference in variances; a Welch t-test of p = 0.013; and an MPTA of 1.28° (0.05-5.87) for R and 1.3°(0.08-14.1) for C with significantly different variances (p = 0.0064). Size of the femoral component has a difference of p < 0.05 between groups. No differences for dispersion of TFA nor for the size of the tibial component were observed. In conclusion, using robot-assisted UKA allows for greater accuracy in the positioning of the implants and in the prediction of the size of the femoral component.

Robotic-assisted vs conventional surgery in medial unicompartmental knee arthroplasty: a clinical and radiological study.

BACKGROUND: The use of unicompartmental knee arthroplasty (UKA) has increased and new technologies have been developed to improve patient survival and satisfaction, soft tissue balance, alignment, and component size. Robot-assisted systems offer an increase in surgical precision and accuracy. The purpose of this study is to evaluate the precision of component position using five radiological parameters in conventional and robotic-assisted medial UKA using the NAVIO system. METHODS: A cohort study was designed for patients who underwent medial UKA between April 2017 and March 2019 in a single center. Patients were allocated in the conventional (UKA-C) or robotic-assisted (UKA-R) group. The variables analyzed were age, gender, affected knee side, length of hospital stay, surgical time, and radiological measurements such as anatomical medial distal femoral angle (aMDFA), anatomical medial proximal tibial angle (aMPTA), tibial slope, the sagittal femoral angle, and the component size. A target was defined for each measurement, and a successful UKA was defined if at least four radiological measures were on target after surgery. Also, patients' reported outcomes were evaluated using the Oxford Knee Score (OKS) and a numeric rating scale (NRS) for pain. RESULTS: Thirty-four patients were included, 18 of them underwent UKA-R. The success rate for UKA in the UKA-R group was 87%; meanwhile, in the UKA-C group this was 28%, this difference was significant and powered (Fisher's exact test, p = 0.001; 1 - ß = 0.95). Also, a 5-point difference in favor of the UKA-R group in the median OKS (p = 0.01), and a significantly lower median NRS for pain (p < 0.000) were found after surgery. CONCLUSIONS: UKA-R achieved more precision in the radiological parameters' measure in this study. Also, UKA-R has a trend towards a better OKS and a lower NRS for pain at short-term follow-up.

Altered Mucus Barrier Integrity and Increased Susceptibility to Colitis in Mice upon Loss of Telocyte Bone Morphogenetic Protein Signalling.

FoxL1+-Telocytes (TCFoxL1+) are subepithelial cells that form a network underneath the epithelium. We have shown that without inflammatory stress, mice with loss of function in the BMP signalling pathway in TCFoxL1+ (BmpR1aΔFoxL1+) initiated colonic neoplasia. Although TCFoxL1+ are modulated in IBD patients, their specific role in this pathogenesis remains unclear. Thus, we investigated how the loss of BMP signalling in TCFoxL1+ influences the severity of inflammation and fosters epithelial recovery after inflammatory stress. BmpR1a was genetically ablated in mouse colonic TCFoxL1+. Experimental colitis was performed using a DSS challenge followed by recovery steps to assess wound healing. Physical barrier properties, including mucus composition and glycosylation, were assessed by alcian blue staining, immunofluorescences and RT-qPCR. We found that BmpR1aΔFoxL1+ mice had impaired mucus quality, and upon exposure to inflammatory challenges, they had increased susceptibility to experimental colitis and delayed healing. In addition, defective BMP signalling in TCFoxL1+ altered the functionality of goblet cells, thereby affecting mucosal structure and promoting bacterial invasion. Following inflammatory stress, TCFoxL1+ with impaired BMP signalling lose their homing signal for optimal distribution along the epithelium, which is critical in tissue regeneration after injury. Overall, our findings revealed key roles of BMP signalling in TCFoxL1+ in IBD pathogenesis.

Binding mechanisms of therapeutic antibodies to human CD20.

Monoclonal antibodies (mAbs) targeting human antigen CD20 (cluster of differentiation 20) constitute important immunotherapies for the treatment of B cell malignancies and autoimmune diseases. Type I and II therapeutic mAbs differ in B cell binding properties and cytotoxic effects, reflecting differential interaction mechanisms with CD20. Here we present 3.7- to 4.7-angstrom cryo-electron microscopy structures of full-length CD20 in complexes with prototypical type I rituximab and ofatumumab and type II obinutuzumab. The structures and binding thermodynamics demonstrate that upon binding to CD20, type II mAbs form terminal complexes that preclude recruitment of additional mAbs and complement components, whereas type I complexes act as molecular seeds to increase mAb local concentration for efficient complement activation. Among type I mAbs, ofatumumab complexes display optimal geometry for complement recruitment. The uncovered mechanisms should aid rational design of next-generation immunotherapies targeting CD20.