A humanized mouse that mounts mature class-switched, hypermutated and neutralizing antibody responses.

Humanized mice are limited in terms of modeling human immunity, particularly with regards to antibody responses. Here we constructed a humanized (THX) mouse by grafting non-γ-irradiated, genetically myeloablated KitW-41J mutant immunodeficient pups with human cord blood CD34+ cells, followed by 17ß-estradiol conditioning to promote immune cell differentiation. THX mice reconstitute a human lymphoid and myeloid immune system, including marginal zone B cells, germinal center B cells, follicular helper T cells and neutrophils, and develop well-formed lymph nodes and intestinal lymphoid tissue, including Peyer's patches, and human thymic epithelial cells. These mice have diverse human B cell and T cell antigen receptor repertoires and can mount mature T cell-dependent and T cell-independent antibody responses, entailing somatic hypermutation, class-switch recombination, and plasma cell and memory B cell differentiation. Upon flagellin or a Pfizer-BioNTech coronavirus disease 2019 (COVID-19) mRNA vaccination, THX mice mount neutralizing antibody responses to Salmonella or severe acute respiratory syndrome coronavirus 2 Spike S1 receptor-binding domain, with blood incretion of human cytokines, including APRIL, BAFF, TGF-ß, IL-4 and IFN-γ, all at physiological levels. These mice can also develop lupus autoimmunity after pristane injection. By leveraging estrogen activity to support human immune cell differentiation and maturation of antibody responses, THX mice provide a platform to study the human immune system and to develop human vaccines and therapeutics.

B Cell Endosomal RAB7 Promotes TRAF6 K63 Polyubiquitination and NF-κB Activation for Antibody Class-Switching.

Upon activation by CD40 or TLR signaling, B lymphocytes activate NF-κB to induce activation-induced cytidine deaminase and, therefore, Ig class switch DNA recombination, as central to the maturation of the Ab and autoantibody responses. In this study, we show that NF-κB activation is boosted by colocalization of engaged immune receptors, such as CD40, with RAB7 small GTPase on mature endosomes, in addition to signals emanating from the receptors localized on the plasma membrane, in mouse B cells. In mature endosomes, RAB7 directly interacts with TRAF6 E3 ubiquitin ligase, which catalyzes K63 polyubiquitination for NF-κB activation. RAB7 overexpression in Cd19+/creRosa26fl-STOP-fl-Rab7 mouse B cells upregulates K63 polyubiquitination activity of TRAF6, enhances NF-κB activation and activation-induced cytidine deaminase induction, and boosts IgG Ab and autoantibody levels. This, together with the extensive intracellular localization of CD40 and the strong correlation of RAB7 expression with NF-κB activation in mouse lupus B cells, shows that RAB7 is an integral component of the B cell NF-κB activation machinery, likely through interaction with TRAF6 for the assembly of "intracellular membrane signalosomes."

Small Molecule Inhibition of Rab7 Impairs B Cell Class Switching and Plasma Cell Survival To Dampen the Autoantibody Response in Murine Lupus.

IgG autoantibodies mediate pathology in systemic lupus patients and lupus-prone mice. In this study, we showed that the class-switched IgG autoantibody response in MRL/Faslpr/lpr and C57/Sle1Sle2Sle2 mice was blocked by the CID 1067700 compound, which specifically targeted Ras-related in brain 7 (Rab7), an endosome-localized small GTPase that was upregulated in activated human and mouse lupus B cells, leading to prevention of disease development and extension of lifespan. These were associated with decreased IgG-expressing B cells and plasma cells, but unchanged numbers and functions of myeloid cells and T cells. The Rab7 inhibitor suppressed T cell-dependent and T cell-independent Ab responses, but it did not affect T cell-mediated clearance of Chlamydia infection, consistent with a B cell-specific role of Rab7. Indeed, B cells and plasma cells were inherently sensitive to Rab7 gene knockout or Rab7 activity inhibition in class switching and survival, respectively, whereas proliferation/survival of B cells and generation of plasma cells were not affected. Impairment of NF-κB activation upon Rab7 inhibition, together with the rescue of B cell class switching and plasma cell survival by enforced NF-κB activation, indicated that Rab7 mediates these processes by promoting NF-κB activation, likely through signal transduction on intracellular membrane structures. Thus, a single Rab7-inhibiting small molecule can target two stages of B cell differentiation to dampen the pathogenic autoantibody response in lupus.

Intrinsic B cell TLR-BCR linked coengagement induces class-switched, hypermutated, neutralizing antibody responses in absence of T cells.

Maturation of antibody responses entails somatic hypermutation (SHM), class-switch DNA recombination (CSR), plasma cell differentiation, and generation of memory B cells, and it is thought to require T cell help. We showed that B cell Toll-like receptor 4 (TLR4)-B cell receptor (BCR) (receptor for antigen) coengagement by 4-hydroxy-3-nitrophenyl acetyl (NP)-lipopolysaccharide (LPS) (Escherichia coli lipid A polysaccharide O-antigen) or TLR5-BCR coengagement by Salmonella flagellin induces mature antibody responses to NP and flagellin in Tcrß-/-Tcrδ-/- and NSG/B mice. TLR-BCR coengagement required linkage of TLR and BCR ligands, "linked coengagement." This induced B cell CSR/SHM, germinal center-like differentiation, clonal expansion, intraconal diversification, plasma cell differentiation, and an anamnestic antibody response. In Tcrß-/-Tcrδ-/- mice, linked coengagement of TLR4-BCR by LPS or TLR5-BCR by flagellin induced protective antibodies against E. coli or Salmonella Typhimurium. Our findings unveiled a critical role of B cell TLRs in inducing neutralizing antibody responses, including those to microbial pathogens, without T cell help.

Maternal anti-Toxoplasma gondii antibodies IgG2, IgG3 and IgG1 are markers of vertical transmission and clinical evolution of toxoplasmosis in the offspring.

Toxoplasma gondii can be transmitted vertically during pregnancy and may cause neurological, ocular, and even systemic damage to the offspring. Congenital toxoplasmosis (CT) can be diagnosed during gestation and/or after birth in the postnatal period. The timely diagnosis is highly relevant for efficient clinical management. The most common laboratory methods for diagnosing CT are based on Toxoplasma-specific humoral immune responses. However, these methods are of low sensitivity or specificity. In a previous study with a small number of cases, the comparison of anti-T. gondii IgG subclasses between mothers and their offspring showed promising results for CT diagnosis and prognosis. Thus, in this work, we analyzed specific IgG subclasses and IgA in 40 T. gondii-infected mothers and their children, of which 27 were congenitally infected and 13 uninfected. A higher frequency of anti-Toxoplasma IgG2, IgG3, IgG4, and IgA antibodies was observed in mothers and congenitally infected offspring. Of these, IgG2 or IgG3 were statistically the most conspicuous. In the CT group, maternal IgG3 antibodies were significantly associated with severe disease of the infants and IgG1 and IgG3 with disseminated disease. The results support that maternal anti-T. gondii IgG3, IgG2 and IgG1 are markers of congenital transmission and severity/spread of disease in the offspring.

LUBAC Suppresses IL-21-Induced Apoptosis in CD40-Activated Murine B Cells and Promotes Germinal Center B Cell Survival and the T-Dependent Antibody Response.

B cell activation by Tfh cells, i.e., through CD154 engagement of CD40 and IL-21, and survival within GCs are crucial for the T-dependent Ab response. LUBAC, composed of HOIP, SHARPIN, and HOIL-1, catalyzes linear ubiquitination (Linear M1-Ub) to mediate NF-κB activation and cell survival induced by TNF receptor superfamily members, which include CD40. As shown in this study, B cells expressing the Sharpin null mutation cpdm (Sharpincpdm ) could undergo proliferation, CSR, and SHM in response to immunization by a T-dependent Ag, but were defective in survival within GCs, enrichment of a mutation enhancing the BCR affinity, and production of specific Abs. Sharpincpdm B cells stimulated in vitro with CD154 displayed normal proliferation and differentiation, marginally impaired NF-κB activation and survival, but markedly exacerbated death triggered by IL-21. While activating the mitochondria-dependent apoptosis pathway in both Sharpin+/+ and Sharpincpdm B cells, IL-21 induced Sharpincpdm B cells to undergo sustained activation of caspase 9 and caspase 8 of the mitochondria-dependent and independent pathway, respectively, and ultimately caspase 3 in effecting apoptosis. These were associated with loss of the caspase 8 inhibitor cFLIP and reduction in cFLIP Linear M1-Ub, which interferes with cFLIP poly-ubiquitination at Lys48 and degradation. Finally, the viability of Sharpincpdm B cells was rescued by caspase inhibitors but virtually abrogated - together with Linear M1-Ub and cFLIP levels - by a small molecule HOIP inhibitor. Thus, LUBAC controls the cFLIP expression and inhibits the effects of caspase 8 and IL-21-activated caspase 9, thereby suppressing apoptosis of CD40 and IL-21-activated B cells and promoting GC B cell survival.

Impact of bariatric surgery on the risk of hospitalization due to influenza virus infection.

BACKGROUND: Obesity independently increases the risk of hospitalization due to viral respiratory infections, including influenza virus and, more recently, severe acute respiratory syndrome coronavirus 2. As an independent risk factor, obesity impairs the immune response to viral infections and decreases the effectiveness of immunizations. OBJECTIVES: Using influenza as a proxy, we aimed to determine the impact of bariatric surgery (BaS) on the risk of hospitalization due to viral respiratory infections. SETTING: Academic hospital, United States. METHODS: National (Nationwide) Inpatient Sample data collected from 2010 to 2015 were examined. Patients were classified as treatment and control groups. Treatment subjects were defined as patients with a history of BaS and control subjects as patients with a body mass index ≥35 kg/m2 and without a history of BaS. Any hospitalization with influenza as a primary diagnosis was identified. Univariate analysis and multivariate regression models were performed to assess the differences between groups. RESULTS: A total of 2,300,845 subjects were reviewed, of which 2,004,804 were control subjects and 296,041 were treated patients. Univariate analysis showed that the hospitalization rate in the treatment group was significantly lower than in the control group (.007% versus .019%, P < .0001), which was confirmed after adjusting for covariables (control versus treatment: odds ratio = 2.21, P = .0010). CONCLUSIONS: BaS may decrease the risk of hospitalization due to influenza, but further prospective studies are needed to confirm these results. We also suggest that these results should be translated into the development of similar studies to determine the impact of BaS on the incidence and severity of the coronavirus disease 2019.

Trends in early postoperative major adverse cardiovascular and cerebrovascular events associated with bariatric surgery: an analysis of the Metabolic and Bariatric Surgery Accreditation and Quality Improvement Program data registry.

BACKGROUND: The population undergoing bariatric surgery (BaS) has many cardiovascular risk factors that can lead to significant perioperative cardiovascular morbidity. OBJECTIVES: We aimed to examine trends in the incidence of major adverse cardiovascular and cerebrovascular events (MACCE) after BaS. SETTING: Academic Hospital, United States METHODS: We performed a retrospective analysis of the Metabolic and Bariatric Surgery Accreditation and Quality Improvement Program (MBSAQIP) data registry for patients aged ≥18 years undergoing laparoscopic sleeve gastrectomy (LSG) and laparoscopic Roux-en-Y gastric bypass (LRYGB) from 2015 to 2019. Data on demographics, co-morbidities, and type of procedure were collected. MACCE was defined as a composite variable including perioperative acute myocardial infarction (AMI), cardiac arrest requiring cardiopulmonary resuscitation, acute stroke, and all-cause mortality. We utilized the Cochrane-Armitage and Jonckheere-Terpstra tests to assess for significant trend changes throughout the years. RESULTS: A total of 752,722 patients were included in our analysis (LSG = 73.2%, LRYGB = 26.8%). Postoperative MACCE occurred in 1058 patients (.14%), and was more frequent in patients undergoing LRYGB (.20%). The frequency of MACCE declined from .17% to .14% (P = .053), driven by a decline in the frequency of AMI (.04% to .02%, P = .002), cardiac arrest (.05% to .04%, P = .897), and all-cause death (.11% to .08%, P = .040), but with an increase in perioperative stroke (.01% to .02%, P = .057). CONCLUSION: The overall risk of MACCE after BaS is .14% and has been declining in the last 5 years. This trend is likely multifactorial and further analysis is necessary to provide a detailed explanation.

Improvement of glucose metabolism following rapid weight loss after bariatric surgery and its impact on reduction of visceral abdominal fat versus free fat muscle.

BACKGROUND: Body fat distribution is highly associated with metabolic disturbances. Skeletal muscle plays an important role in glucose metabolism, as it serves as an important organ for glucose storage in the form of glycogen. In fact, low muscle mass has been associated with metabolic syndrome, type 2 diabetes (T2D), systemic inflammation, and decreased survival. OBJECTIVES: To compare the relationship between visceral abdominal fat (VAF) and fat free mass (FFM) with the improved glucose metabolism after bariatric surgery. SETTING: University hospital, United States. METHODS: A retrospective review was performed of all patients who underwent bariatric surgery between 2011 and 2017 at a university hospital in the United States. In severely obese patients with T2D, we measured the VAF via abdominal computed tomography scan and we calculated the FFM preoperatively and at a 12-month follow-up. Data collected included baseline demographic characteristics and perioperative parameters, such as treatment for hypertension (HTN) and T2D, body mass index (BMI), glycated hemoglobin (HbA1C), glucose, and lipid profile. RESULTS: A total of 25 patients met the inclusion criteria. The average age was 52.5 ± 11.6 years. The initial BMI was 41.41 ± 5.7 kg/m2 and the postoperative BMI was 31.7 ± 6.9 kg/m2 (P < .0001). The preoperative VAF volume was 184.6 ± 90.2 cm3 and the postoperative VAF volume was 93.8 ± 46.8 cm3 at the 12-month follow-up (P < .0001). The preoperative FFM was 55.2 ± 11.4 kg and the postoperative FFM was 49.1 ± 12 kg (P < .072). The preoperative HbA1C was 5.8% ± .9%, which decreased postoperatively to 5.3% ± .4% at the 12-month follow-up (P < .013). CONCLUSION: Bariatric surgery has been demonstrated to be an effective treatment modality for severe obesity and T2D. Our results suggest that at 12 months, there is a reduction in VAF and HbA1C without a significant loss of FFM. Further prospective studies are needed to better understand these findings.

Lumbar Epidural Steroid Injections.

Lumbar epidural steroid injections under fluoroscopic guidance are used very commonly for the treatment of low back and lower extremity radicular pain. These procedures have been shown to be effective for pain relief in the short term and are relatively safe. The indications, evidence, and safety considerations for 2 different techniques-namely, interlaminar and transforaminal-are discussed.

Clinical, biomechanical and morphological assessment of anterior cruciate ligament Kevlar®-based artificial prosthesis in rabbit model.

BACKGROUND: The aim of this study was to evaluate the clinical, biomechanical and morphological characteristics of a Kevlar®-based prosthetic ligament as a synthetic graft of the anterior cruciate ligament (ACL) in an experimental animal model in rabbits. METHODS: A total of 27 knees of rabbits randomly divided into 3 groups (control, ACL excision and ACL replacement with a Kevlar® prosthesis) were analyzed using clinical, biomechanical and morphological tests at 6, 12 and 18 weeks postprocedure. RESULTS: The mean displacement in mechanical testing was 0.73 ± 0.06 mm, 1.58 ± 0.19 mm and 0.94 ± 0.20 mm for the control, ACL excision and ACL replacement with synthetic prosthesis groups, respectively. The results showed an improvement in the stability of the knee with the use of the Kevlar® synthetic prosthesis in the biomechanical testing (p<0.05) compared with rabbits that underwent ACL excision, in addition to displacements that were larger but comparable to that in the control group (p>0.05), between the replacement group and the control group. The histological study revealed a good morphological adaptation of the synthetic material to the knee. CONCLUSIONS: This study proposes a new animal model for the placement and evaluation of Kevlar®-based synthetic ACL implants. The studied prosthesis showed promising behavior in the clinical and biomechanical tests and in the histological analysis. This study lays the foundation for further basic and clinical studies of artificial ACL prostheses using this material.

Core and Lumbopelvic Stabilization in Runners.

Core muscles provide stability that allows generation of force and motion in the lower extremities, as well as distributing impact forces and allowing controlled and efficient body movements. Imbalances or deficiencies in the core muscles can result in increased fatigue, decreased endurance, and injury in runners. Core strengthening should incorporate the intrinsic needs of the core for flexibility, strength, balance, and endurance, and the function of the core in relation to its role in extremity function and dysfunction. Specific exercises are effective in strengthening the core muscles.

Aplicaciones clínicas de la colangiopancreatografía por resonancia magnética / Clinical usefulness of magnetic resonance cholangiopancreatography

La colangiopancreatografía por resonancia magnética (CPRM) es la alternativa diagnóstica más importante que ha surgido en los últimos años para la evaluación de las vías biliar y pancreática. Las ventajas de este método son: no utiliza medio de contraste ni radiación ionizante, no es invasivo, está exento de complicaciones y el tiempo de estudio es relativamente corto (aproximadamente 20 a 30 minutos). Tiene altas sensibilidad y especificidad para diagnosticar la dilatación biliar y para demostrar el sitio y la causa de la estenosis. Para los cálculos biliares y pancreáticos su exactitud diagnóstica es similar a la de la colangiopancreatografía endoscópica retrógrada (CPER). En variantes anatómicas biliopancreáticas ha reemplazado a la CPER como método diagnóstico. En la CPER fallida, la CPRM es casi la única modalidad diagnóstica para la evaluación de los conductos biliares. Otra aplicación más reciente es la colangitis esclerosante primaria para cuyo diagnóstico tiene algunas ventajas sobre la CPER. Este artículo es una revisión de las aplicaciones clínicas de la CPRM en la evaluación de las enfermedades biliopancreáticas.