Spatiotemporal immune atlas of a clinical-grade gene-edited pig-to-human kidney xenotransplant.

Pig-to-human xenotransplantation is rapidly approaching the clinical arena; however, it is unclear which immunomodulatory regimens will effectively control human immune responses to pig xenografts. Here, we transplant a gene-edited pig kidney into a brain-dead human recipient on pharmacologic immunosuppression and study the human immune response to the xenograft using spatial transcriptomics and single-cell RNA sequencing. Human immune cells are uncommon in the porcine kidney cortex early after xenotransplantation and consist of primarily myeloid cells. Both the porcine resident macrophages and human infiltrating macrophages express genes consistent with an alternatively activated, anti-inflammatory phenotype. No significant infiltration of human B or T cells into the porcine kidney xenograft is detectable. Altogether, these findings provide proof of concept that conventional pharmacologic immunosuppression may be able to restrict infiltration of human immune cells into the xenograft early after compatible pig-to-human kidney xenotransplantation.

Crohn's patients and healthy infants share immunodominant B cell response to commensal flagellin peptide epitopes.

About half of patients with Crohn's disease (CD) develop selective serum IgG response to flagellin proteins of the Lachnospiraceae family. Here, we identified a dominant B cell peptide epitope in CD, locating in the highly conserved "hinge region" between the D0 and D1 domains at the amino-terminus of Lachnospiraceae flagellins. Serum IgG reactive to this epitope is present at an elevated level in adult CD patients and in pediatric CD patients at diagnosis. Most importantly, high levels of serum IgG to the hinge epitope were found in most infants from 3 different geographic regions (Uganda, Sweden, and the USA) at one year of age. This vigorous homeostatic response decrements with age as it is not present in healthy adults. These data identify a distinct subset of CD patients, united by a shared reactivity to this dominant flagellin epitope that may represent failure of a homeostatic response beginning in infancy.

B cell and antibody targeting of transplant alloantigen epitopes is supported by both self and allo-recognition.

Donor-specific antibody (DSA) responses against human leukocyte antigen (HLA) proteins mismatched between kidney transplant donors and recipients cause allograft loss. Using single-cell, molecular, structural, and proteomic techniques, we profiled the HLA-specific (alloreactive) B cell response in kidney and blood of a transplant recipient with antibody-mediated rejection (AMR). We identified 14 distinct alloreactive B cell lineages, which spanned the rejected organ and blood and expressed high-affinity anti-donor HLA-specific B cell receptors, many of which were clonally linked to circulating DSA. The alloreactive B cell response was focused on exposed, solvent-accessible mismatched HLA residues, while also demonstrating extensive contacts with self-HLA residues. Consistent with structural evidence of self-recognition, measurable self-reactivity by donor-specific B cells was common and positively correlated with anti-donor affinity maturation. Thus, allo- and self-reactive signatures appeared to converge, suggesting that during AMR, the recognition of non-self and breaches of tolerance conspire to produce a pathogenic donor-specific adaptive response.

Spatiotemporal immune atlas of the first clinical-grade, gene-edited pig-to-human kidney xenotransplant.

Pig-to-human xenotransplantation is rapidly approaching the clinical arena; however, it is unclear which immunomodulatory regimens will effectively control human immune responses to pig xenografts. We transplanted a gene-edited pig kidney into a brain-dead human recipient on pharmacologic immunosuppression and studied the human immune response to the xenograft using spatial transcriptomics and single-cell RNA sequencing. Human immune cells were uncommon in the porcine kidney cortex early after xenotransplantation and consisted of primarily myeloid cells. Both the porcine resident macrophages and human infiltrating macrophages expressed genes consistent with an alternatively activated, anti-inflammatory phenotype. No significant infiltration of human B or T cells into the porcine kidney xenograft was detected. Altogether, these findings provide proof of concept that conventional pharmacologic immunosuppression is sufficient to restrict infiltration of human immune cells into the xenograft early after compatible pig-to-human kidney xenotransplantation.

COVID-19 Vaccination and Remdesivir are Associated With Protection From New or Increased Levels of Donor-Specific Antibodies Among Kidney Transplant Recipients Hospitalized With COVID-19.

Alloimmune responses in kidney transplant (KT) patients previously hospitalized with COVID-19 are understudied. We analyzed a cohort of 112 kidney transplant recipients who were hospitalized following a positive SARS-CoV-2 test result during the first 20 months of the COVID-19 pandemic. We found a cumulative incidence of 17% for the development of new donor-specific antibodies (DSA) or increased levels of pre-existing DSA in hospitalized SARS-CoV-2-infected KT patients. This risk extended 8 months post-infection. These changes in DSA status were associated with late allograft dysfunction. Risk factors for new or increased DSA responses in this KT patient cohort included the presence of circulating DSA pre-COVID-19 diagnosis and time post-transplantation. COVID-19 vaccination prior to infection and remdesivir administration during infection were each associated with decreased likelihood of developing a new or increased DSA response. These data show that new or enhanced DSA responses frequently occur among KT patients requiring admission with COVID-19 and suggest that surveillance, vaccination, and antiviral therapies may be important tools to prevent alloimmunity in these individuals.

First clinical-grade porcine kidney xenotransplant using a human decedent model.

A radical solution is needed for the organ supply crisis, and the domestic pig is a promising organ source. In preparation for a clinical trial of xenotransplantation, we developed an in vivo pre-clinical human model to test safety and feasibility tenets established in animal models. After performance of a novel, prospective compatible crossmatch, we performed bilateral native nephrectomies in a human brain-dead decedent and subsequently transplanted two kidneys from a pig genetically engineered for human xenotransplantation. The decedent was hemodynamically stable through reperfusion, and vascular integrity was maintained despite the exposure of the xenografts to human blood pressure. No hyperacute rejection was observed, and the kidneys remained viable until termination 74 h later. No chimerism or transmission of porcine retroviruses was detected. Longitudinal biopsies revealed thrombotic microangiopathy that did not progress in severity, without evidence of cellular rejection or deposition of antibody or complement proteins. Although the xenografts produced variable amounts of urine, creatinine clearance did not recover. Whether renal recovery was impacted by the milieu of brain death and/or microvascular injury remains unknown. In summary, our study suggests that major barriers to human xenotransplantation have been surmounted and identifies where new knowledge is needed to optimize xenotransplantation outcomes in humans.

Single-Dose Intranasal Administration of AdCOVID Elicits Systemic and Mucosal Immunity against SARS-CoV-2 and Fully Protects Mice from Lethal Challenge.

The coronavirus disease 2019 (COVID-19) pandemic has highlighted the urgent need for effective prophylactic vaccination to prevent the spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Intranasal vaccination is an attractive strategy to prevent COVID-19 as the nasal mucosa represents the first-line barrier to SARS-CoV-2 entry. The current intramuscular vaccines elicit systemic immunity but not necessarily high-level mucosal immunity. Here, we tested a single intranasal dose of our candidate adenovirus type 5-vectored vaccine encoding the receptor-binding domain (RBD) of the SARS-CoV-2 spike protein (AdCOVID) in inbred, outbred, and transgenic mice. A single intranasal vaccination with AdCOVID elicited a strong and focused immune response against RBD through the induction of mucosal IgA in the respiratory tract, serum neutralizing antibodies, and CD4+ and CD8+ T cells with a Th1-like cytokine expression profile. A single AdCOVID dose resulted in immunity that was sustained for over six months. Moreover, a single intranasal dose completely protected K18-hACE2 mice from lethal SARS-CoV-2 challenge, preventing weight loss and mortality. These data show that AdCOVID promotes concomitant systemic and mucosal immunity and represents a promising vaccine candidate.

Memory B Cells in Pregnancy Sensitization.

Memory B cells play an important role in immunity to pathogens as these cells are poised to rapidly differentiate into antibody-secreting cells upon antigen re-encounter. Memory B cells also develop over the course of HLA-sensitization during pregnancy and transplantation. In this review, we discuss the potential contribution of memory B cells to pregnancy sensitization as well as the impact of these cells on transplant candidacy and outcomes. We start by summarizing how B cell subsets are altered in pregnancy and discuss what is known about HLA-specific B cell responses given our current understanding of fetal antigen availability in maternal secondary lymphoid tissues. We then review the molecular mechanisms governing the generation and maintenance of memory B cells during infection - including the role of T follicular helper cells - and discuss the experimental evidence for the development of these cells during pregnancy. Finally, we discuss how memory B cells impact access to transplantation and transplant outcomes for a range of transplant recipients.

Single-dose intranasal administration of AdCOVID elicits systemic and mucosal immunity against SARS-CoV-2 in mice.

The coronavirus disease 2019 (COVID-19) pandemic has highlighted the urgent need for effective preventive vaccination to reduce burden and spread of severe acute respiratory syndrome (SARS) coronavirus 2 (SARS-CoV-2) in humans. Intranasal vaccination is an attractive strategy to prevent COVID-19 as the nasal mucosa represents the first-line barrier to SARS-CoV-2 entry before viral spread to the lung. Although SARS-CoV-2 vaccine development is rapidly progressing, the current intramuscular vaccines are designed to elicit systemic immunity without conferring mucosal immunity. Here, we show that AdCOVID, an intranasal adenovirus type 5 (Ad5)-vectored vaccine encoding the receptor binding domain (RBD) of the SARS-CoV-2 spike protein, elicits a strong and focused immune response against RBD through the induction of mucosal IgA, serum neutralizing antibodies and CD4+ and CD8+ T cells with a Th1-like cytokine expression profile. Therefore, AdCOVID, which promotes concomitant systemic and local mucosal immunity, represents a promising COVID-19 vaccine candidate.

A Predictive Model for a Reputation-Based General Surgery Residency Match and a Novel Online Calculator.

OBJECTIVE: This study aimed to identify medical student characteristics that predict a successful categorical match into a general surgery residency and a match based upon Doximity program rankings. DESIGN: This was a retrospective study that analyzed academic and personal predictors of a successful general surgery residency match. SETTING: This study was set at the University of Alabama at Birmingham School of Medicine, a public medical school. PARTICIPANTS: This study included 173 fourth-year medical students at a public medical school who matched into general surgery residency programs. METHODS: Our cohort comprised students graduating from our institution between 2004 and 2015 that matched into preliminary or categorical general surgery positions. We collected academic variables and performed univariate analyses and logistic regression to examine the likelihood of specific match outcomes. RESULTS: Of 173 students, 132 (76%) matched into a categorical position and 41 (24%) matched into a preliminary position. Of all variables, clinical ranking quartile was most effective in predicting a categorical match (R2 = 0.35). Models for a match based upon Doximity ranking lacked the same predictive power. CONCLUSIONS: This research identifies students that are at risk for not matching into a categorical position and predicts competitiveness for certain programs. It provides a novel calculator to give applicants easily interpretable match probabilities.

Scoliosis in Rett Syndrome: Progression, Comorbidities, and Predictors.

BACKGROUND: Scoliosis is prominent in Rett syndrome (RTT). Following the prior report from the US Natural History Study, the onset and progression of severe scoliosis (≥40° Cobb angle) and surgery were examined regarding functional capabilities and specific genotypes, addressing the hypothesis that abnormal muscle tone, poor oral feeding, puberty, and delays or absence of sitting balance and ambulation may be responsible for greater risk in RTT. METHODS: The multicenter RTT Natural History Study gathered longitudinal data for classic RTT, including mutation type, scoliosis, muscle tone, sitting, ambulation, hand function, and feeding. Cox regression models were used to examine the association between scoliosis and functional characteristics. All analyses utilized SAS 9.4; two-sided P values of <0.05 were considered significant. RESULTS: A total of 913 females with classic RTT were included. Scoliosis frequency and severity increased with age. Severe scoliosis was found in 251 participants (27%), 113 of whom developed severe scoliosis during the follow-up assessments; 168 (18%) had surgical correction. Severe MECP2 mutations (R106W, R168X, R255X, R270X, and large deletions) showed a higher proportion of scoliosis. Individuals developing severe scoliosis or requiring surgery were less likely to sit, ambulate, or use their hands and were more likely to have begun puberty. Significant differences were absent for epilepsy rates, sleep problems, or constipation. DISCUSSION: Scoliosis requires vigilance regarding the risk factors noted, particularly specific mutations and the role of puberty and motor abilities. Bracing is recommended for moderate curves and surgery for severe curves in accordance with published guidelines for scoliosis management.

Caretaker Quality of Life in Rett Syndrome: Disorder Features and Psychological Predictors.

OBJECTIVE: Rett syndrome is a severe neurodevelopmental disorder affecting approximately one in 10,000 female births. The clinical features of Rett syndrome are known to impact both patients' and caretakers' quality of life in Rett syndrome. We hypothesized that more severe clinical features would negatively impact caretaker physical quality of life but would positively impact caretaker mental quality of life. METHODS: Participants were individuals enrolled in the Rett Natural History Study with a diagnosis of classic Rett syndrome. Demographic data, clinical disease features, caretaker quality of life, and measures of family function were assessed during clinic visits. The Optum SF-36v2 Health Survey was used to assess caretaker physical and mental quality of life (higher scores indicate better quality of life). Descriptive, univariate, and multivariate analyses were used to characterize relationships between child and caretaker characteristics and caretaker quality of life. RESULTS: Caretaker physical component scores (PCS) were higher than mental component scores (MCS): 52.8 (9.7) vs 44.5 (12.1). No differences were demonstrated between the baseline and 5-year follow-up. In univariate analyses, disease severity was associated with poorer PCS (P = 0.006) and improved MCS (P = 0.003). Feeding problems were associated with poorer PCS (P = 0.007) and poorer MCS (P = 0.018). In multivariate analyses, limitations in caretaker personal time and home conflict adversely affected PCS. Feeding problems adversely impacted MCS. CONCLUSIONS: Caretaker quality of life in Rett syndrome is similar to that for caretakers in other chronic diseases. Disease characteristics significantly impact quality of life, and feeding difficulties may represent an important clinical target for improving both child and caretaker quality of life. The stability of quality-of-life scores between baseline and five years adds important value.

Pubertal development in Rett syndrome deviates from typical females.

BACKGROUND: Rett syndrome is a unique neurodevelopmental disorder, affecting approximately one in 10,000 live female births, most experiencing reduced growth. We characterized pubertal trajectories in females with Rett syndrome. We hypothesized that pubertal trajectory deviates from the general female population with early pubertal onset and delayed menarche. METHODS: Participants were individuals enrolled in the Rett Syndrome Natural History Study with clinical diagnosis of Rett syndrome or mutations in MECP2. Intervals to thelarche, adrenarche, and menarche were assessed by survival analysis; body mass index, mutation type, clinical severity, and pubertal milestone relationships were assessed by log-likelihood test; pathway synchrony (relationship between thelarche, adrenarche, and menarche) was assessed by chi-squared analysis. RESULTS: Compared with the general female population, more than 25% initiated puberty early, yet entered menarche later (median age 13.0 years). A total of 19% experienced delayed menarche. Median length of puberty, from thelarche to menarche, was 3.9 years. Higher body mass index correlated with earlier thelarche and adrenarche but not menarche; milder mutations correlated with earlier menarche; and milder clinical presentation correlated with earlier thelarche and menarche. Fifty-two percent entered puberty in synchrony, but different from the general population, 15% led with thelarche and 32% with adrenarche. CONCLUSIONS: Pubertal trajectories in Rett syndrome differ from general population, entering puberty early and reaching menarche later. Body mass index affects pubertal timing, but the relationship between specific mutations, clinical presentation, and underlying neuroendocrine pathology is less clear.