Lipotoxicity and immunometabolism in ischemic acute kidney injury: current perspectives and future directions.

Dysregulated lipid metabolism is implicated in the pathophysiology of a range of kidney diseases. The specific mechanisms through which lipotoxicity contributes to acute kidney injury (AKI) remain poorly understood. Herein we review the cardinal features of lipotoxic injury in ischemic kidney injury; lipid accumulation and mitochondrial lipotoxicity. We then explore a new mechanism of lipotoxicity, what we define as "immunometabolic" lipotoxicity, and discuss the potential therapeutic implications of targeting this lipotoxicity using lipid lowering medications.

HLA matching between donors and recipients improves clinical liver transplant graft survival.

BACKGROUND AND AIMS: The importance of human leukocyte antigen (HLA) matching between liver transplant donors and recipients on graft survival remains unclear and is not a clinical consideration in liver transplantation. This study aimed to determine the relationship between HLA matching and liver graft survival using a large-scale multi-centre database (UNOS/OPTN) and multivariate logistic analysis. The secondary aim was to determine whether this relationship was influenced by transplant indication and donor status. METHODS: This retrospective observational analysis was performed using 22 702 liver transplant recipients from the UNOS/OPTN database. Patients were divided into two groups based on number of HLA mismatches (0-3 mismatches vs. 4-6 mismatches) and then subcategorized by indication and donor status. Risk-adjusted outcomes were assessed by multivariate Cox analysis adjusting for donor and recipient characteristics and visualized using Kaplan-Meier survival curves. RESULTS: Allograft survival and risk of acute rejection were associated with degree of HLA mismatch. This association between HLA mismatch and graft survival persisted in individuals who underwent transplant for hepatitis, metabolic, drug toxicity, and congenital indications. Donor status also influenced the relationship between HLA mismatch and graft survival. Graft survival in DBD recipients was longer than in DCD in the 4-6 HLA mismatch group, whereas no significant difference was found in the 0-3 HLA mismatch group. CONCLUSION: HLA mismatch significantly reduced graft survival and increased risk of acute rejection. This association was noted only in specific indications. These findings are of potential clinical relevance to organ allocation, allograft matching algorithms, immunosuppression protocols, and transplant surveillance.

Trends in the mortality, incidence and disability-adjusted life-years of appendicitis in EU15+ countries: an observational study of the Global Burden of Disease Database, 1990-2019.

BACKGROUND: Appendicitis places a substantial burden on healthcare systems, with acute appendicitis alone being the most common abdominal surgical emergency worldwide. Further characterisation of the disease burden in EU15+ countries may help optimise the distribution of healthcare resources. The aim of this observational study was to assess the trends in mortality, incidence and disability-adjusted life-years (DALYs) of appendicitis across European Union (EU) 15+ countries between the years 1990 and 2019, Supplemental Digital Content 3, http://links.lww.com/JS9/A589 . MATERIALS AND METHODS: Age-standardised mortality rates (ASMRs), age-standardised incidence rates (ASIRs) and DALYs data for appendicitis in males and females were extracted from the 2019 Global Burden of Disease (GBD) study. Temporal trends within the study period were analysed using Joinpoint regression analysis. RESULTS: The median ASMRs across EU15+ countries in 2019 were 0.08/100 000 and 0.13/100 000 for females and males, respectively. Between 1990 and 2019 the median percentage change in ASMR was -52.12% for females and -53.18% in males. The median ASIRs in 2019 for females and males were 251/100 000 and 278/100 000, respectively, with a median percentage change of +7.22% for females and +3.78% for males during the observation period. Decreasing trends in DALYs were observed over the 30-year study period, with median percentage changes of -23.57% and -33.81% for females and males, respectively, Supplemental Digital Content 3, http://links.lww.com/JS9/A589 . CONCLUSION: Overall, a general trend of decreasing appendicitis ASMRs and DALYs was observed across EU15+ countries, despite small overall increases in appendicitis ASIRs, Supplemental Digital Content 3, http://links.lww.com/JS9/A589 . Variations in both diagnostic and management strategies over the study period are likely contributory to the changing trends.

A Systematic Review of the Reported Complications Related to Facial and Upper Extremity Vascularized Composite Allotransplantation.

INTRODUCTION: Twenty three years after the first successful upper extremity transplantation, the role of vascularized composite allotransplantation (VCA) in the world of transplantation remains controversial. Face and upper extremity reconstruction via transplantation have become successful options for highly selected patients with severe tissue and functional deficit when conventional reconstructive options are no longer available. Despite clear benefit in these situations, VCA has a significant potential for complications that are more frequent when compared to visceral organ transplantation. This study intended to perform an updated systematic review on such complications. MATERIALS AND METHODS: MEDLINE database via PubMed, Embase and Cochrane Library were searched. Face and upper extremity VCA performed between 1998 and 2021 were included in the study. Relevant media and press conferences reports were also included. Complications related to face and upper extremity VCA were recorded and reviewed including their clinical characteristics and complications. RESULTS: One hundred fifteen patients underwent facial (43%) or upper extremity (57%) transplantation. Overall, the surgical complication rate was 23%. Acute and chronic rejection was identified in 89% and 11% of patients, respectively. Fifty eight percent of patients experienced opportunistic infection. Impaired glucose metabolism was the most common immunosuppression-related complication other than infection. Nineteen percent of patients ultimately experienced partial or complete allograft loss. CONCLUSIONS: Complications related to VCA are a significant source of morbidity and potential mortality. Incidence of such complications is higher than previously reported and should be strongly emphasized in patient consent process. Strict patient selection criteria, complex preoperative evaluation, consideration of alternatives, and thorough disclosure to patients should be routinely performed prior to VCA indication.

Symptomatic atherosclerotic vascular disease and graft survival in primary kidney transplant recipients - Observational analysis of the united network of organ sharing database.

OBJECTIVES: The aim of this study was to investigate whether symptomatic atherosclerotic vascular disease (SAVD) was associated with graft survival in primary kidney transplant recipients. SUMMARY BACKGROUND DATA: Recipient atherosclerotic vascular disease is associated with increased mortality rates amongst renal transplant patients. However, its relationship with graft survival has not been well studied. METHODS: This retrospective observational analysis was performed using data for adult kidney transplant recipients between 11/09/2000 and 28/02/2020 extracted from the UNOS national organ transplantation database. Patients were divided into two groups based on recipient history of symptomatic atherosclerotic disease (angina or peripheral vascular disease). Risk-adjusted outcomes were assessed by multivariate Cox regression analysis adjusting for both donor and recipient characteristics. RESULTS: 11,771 adult kidney transplant recipients from the UNOS database were eligible for analysis (1543 had a history of SAVD, 10,228 did not have a history of SAVD). After adjusting for confounders, positive SAVD status was associated with an adverse effect on graft survival at both 1 year (HR 1.35, p < 0.001) and 10 years (HR 1.15, p < 0.001). CONCLUSIONS: SAVD should be considered an independent risk factor for poor prognosis in patients undergoing kidney transplant.

Improved Kasai Hepatoportoenterostomy Outcomes After Implementation of a Dedicated Biliary Atresia Team.

It can be difficult or impractical to refer all biliary atresia (BA) patients to high-volume centers. Our hypothesis was that a low volume center could improve outcomes with implementation of a dedicated multidisciplinary BA team. We conducted a retrospective study of patients with BA who underwent hepatic portoenterostomy at our institution from 2003 to 2020, before and after the development of a dedicated BA team. Ten consecutive patients with BA were identified following the establishment of a dedicated BA team. Since the establishment of the BA team, total bilirubin (TB) clearance (TB < 2 mg/dL) achieved by 3 and 6 months has been 60% and 60%, respectively, and survival of the native liver (SNL) at 1 and 2 years post HPE at 90% and 86%, respectively. Outcomes were markedly improved after the team was established. A dedicated BA team prioritizing communication and expeditious workup can improve outcomes at a low volume center.

Twenty Years of Abdominal Wall Allotransplantation: A Systematic Review of the Short- and Long-Term Outcomes.

BACKGROUND: Abdominal wall allotransplantation following intestinal and multivisceral transplant procedures has proven to be successful in achieving adequate closure in patients in whom other techniques have proven inadequate. Thus far, the focus of these abdominal wall allotransplants has been on graft and overall patient survival following surgery and the implementation of immunosuppression. The purpose of this study was to review the outcomes of abdominal wall allotransplantation reported in the literature. METHODS: The PubMed database was queried, and 2595 articles were found. Search criteria used were "abdominal wall transplant" and "abdominal wall allotransplant." Of these, eight met inclusion/exclusion criteria. RESULTS: In the present study, eight publications were identified reporting abdominal wall allotransplants, for a total of 38 full-thickness abdominal wall allotransplantations performed worldwide. All studies reported abdominal wall allotransplantation in combination with visceral organ allotransplantation. Abdominal wall allotransplantations reported thus far have been nonneurotized. Abdominal wall allotransplantations have proven to be beneficial both in terms of abdominal wall closure and acting as a sentinel marker for rejection for underlying visceral organ allotransplantation. The success of abdominal wall allotransplants and their long-term survival has introduced the question of functionality and long-term durability. Cadaveric studies have shown that it is possible to neurotize abdominal wall allotransplants, and future direction toward neurotized abdominal wall allotransplantation requires tools to assess functional outcomes of these transplants. CONCLUSIONS: Abdominal wall allotransplantation is an important reconstructive option when abdominal wall closure is challenging and should be considered in combination with visceral organ allotransplantations. There may be potential benefit in neurotizing the abdominal wall allotransplant for functional use, and future studies should aim to include functional outcomes.

The Effect of Normothermic Machine Perfusion on the Immune Profile of Donor Liver.

Background: Normothermic machine perfusion (NMP) allows viability assessment and potential resuscitation of donor livers prior to transplantation. The immunological effect of NMP on liver allografts is undetermined, with potential implications on allograft function, rejection outcomes and overall survival. In this study we define the changes in immune profile of human livers during NMP. Methods: Six human livers were placed on a NMP device. Tissue and perfusate samples were obtained during cold storage prior to perfusion and at 1, 3, and 6 hours of perfusion. Flow cytometry, immunohistochemistry, and bead-based immunoassays were used to measure leukocyte composition and cytokines in the perfusate and within the liver tissue. Mean values between baseline and time points were compared by Student's t-test. Results: Within circulating perfusate, significantly increased frequencies of CD4 T cells, B cells and eosinophils were detectable by 1 hour of NMP and continued to increase at 6 hours of perfusion. On the other hand, NK cell frequency significantly decreased by 1 hour of NMP and remained decreased for the duration of perfusion. Within the liver tissue there was significantly increased B cell frequency but decreased neutrophils detectable at 6 hours of NMP. A transient decrease in intermediate monocyte frequency was detectable in liver tissue during the middle of the perfusion run. Overall, no significant differences were detectable in tissue resident T regulatory cells during NMP. Significantly increased levels of pro-inflammatory and anti-inflammatory cytokines were seen following initiation of NMP that continued to rise throughout duration of perfusion. Conclusions: Time-dependent dynamic changes are seen in individual leukocyte cell-types within both perfusate and tissue compartments of donor livers during NMP. This suggests a potential role of NMP in altering the immunogenicity of donor livers prior to transplant. These data also provide insights for future work to recondition the intrinsic immune profile of donor livers during NMP prior to transplantation.

Donor leukocyte trafficking during human ex vivo lung perfusion.

BACKGROUND: Ex vivo lung perfusion (EVLP) is used to assess and preserve lungs prior to transplantation. However, its inherent immunomodulatory effects are not completely understood. We examine perfusate and tissue compartments to determine the change in immune cell composition in human lungs maintained on EVLP. METHODS: Six human lungs unsuitable for transplantation underwent EVLP. Tissue and perfusate samples were obtained during cold storage and at 1-, 3- and 6-h during perfusion. Flow cytometry, immunohistochemistry, and bead-based immunoassays were used to measure leukocyte composition and cytokines. Mean values between baseline and time points were compared by Student's t test. RESULTS: During the 1st hour of perfusion, perfusate neutrophils increased (+22.2 ± 13.5%, p < 0.05), monocytes decreased (-77.5 ± 8.6%, p < 0.01) and NK cells decreased (-61.5 ± 22.6%, p < 0.01) compared to cold storage. In contrast, tissue neutrophils decreased (-22.1 ± 12.2%, p < 0.05) with no change in monocytes and NK cells. By 6 h, perfusate neutrophils, NK cells, and tissue neutrophils were similar to baseline. Perfusate monocytes remained decreased, while tissue monocytes remained unchanged. There was no significant change in B cells or T cell subsets. Pro-inflammatory cytokines (IL-1b, G-CSF, IFN-gamma, CXCL2, CXCL1 granzyme A, and granzyme B) and lymphocyte activating cytokines (IL-2, IL-4, IL-6, IL-8) increased during perfusion. CONCLUSIONS: Early mobilization of innate immune cells occurs in both perfusate and tissue compartments during EVLP, with neutrophils and NK cells returning to baseline and monocytes remaining depleted after 6 h. The immunomodulatory effect of EVLP may provide a therapeutic window to decrease the immunogenicity of lungs prior to transplantation.

Profiling B cell immunodominance after SARS-CoV-2 infection reveals antibody evolution to non-neutralizing viral targets.

Dissecting the evolution of memory B cells (MBCs) against SARS-CoV-2 is critical for understanding antibody recall upon secondary exposure. Here, we used single-cell sequencing to profile SARS-CoV-2-reactive B cells in 38 COVID-19 patients. Using oligo-tagged antigen baits, we isolated B cells specific to the SARS-CoV-2 spike, nucleoprotein (NP), open reading frame 8 (ORF8), and endemic human coronavirus (HCoV) spike proteins. SARS-CoV-2 spike-specific cells were enriched in the memory compartment of acutely infected and convalescent patients several months post symptom onset. With severe acute infection, substantial populations of endemic HCoV-reactive antibody-secreting cells were identified and possessed highly mutated variable genes, signifying preexisting immunity. Finally, MBCs exhibited pronounced maturation to NP and ORF8 over time, especially in older patients. Monoclonal antibodies against these targets were non-neutralizing and non-protective in vivo. These findings reveal antibody adaptation to non-neutralizing intracellular antigens during infection, emphasizing the importance of vaccination for inducing neutralizing spike-specific MBCs.

Nanotraps for the containment and clearance of SARS-CoV-2.

SARS-CoV-2 enters host cells through its viral spike protein binding to angiotensin-converting enzyme 2 (ACE2) receptors on the host cells. Here, we show that functionalized nanoparticles, termed "Nanotraps," completely inhibited SARS-CoV-2 infection by blocking the interaction between the spike protein of SARS-CoV-2 and the ACE2 of host cells. The liposomal-based Nanotrap surfaces were functionalized with either recombinant ACE2 proteins or anti-SARS-CoV-2 neutralizing antibodies and phagocytosis-specific phosphatidylserines. The Nanotraps effectively captured SARS-CoV-2 and completely blocked SARS-CoV-2 infection to ACE2-expressing human cell lines and primary lung cells; the phosphatidylserine triggered subsequent phagocytosis of the virus-bound, biodegradable Nanotraps by macrophages, leading to the clearance of pseudotyped and authentic virus in vitro. Furthermore, the Nanotraps demonstrated an excellent biosafety profile in vitro and in vivo. Finally, the Nanotraps inhibited pseudotyped SARS-CoV-2 infection in live human lungs in an ex vivo lung perfusion system. In summary, Nanotraps represent a new nanomedicine for the inhibition of SARS-CoV-2 infection.

SARS-CoV-2 Infection Severity Is Linked to Superior Humoral Immunity against the Spike.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is currently causing a global pandemic. The antigen specificity of the antibody response mounted against this novel virus is not understood in detail. Here, we report that subjects with a more severe SARS-CoV-2 infection exhibit a larger antibody response against the spike and nucleocapsid protein and epitope spreading to subdominant viral antigens, such as open reading frame 8 and nonstructural proteins. Subjects with a greater antibody response mounted a larger memory B cell response against the spike, but not the nucleocapsid protein. Additionally, we revealed that antibodies against the spike are still capable of binding the D614G spike mutant and cross-react with the SARS-CoV-1 receptor binding domain. Together, this study reveals that subjects with a more severe SARS-CoV-2 infection exhibit a greater overall antibody response to the spike and nucleocapsid protein and a larger memory B cell response against the spike.IMPORTANCE With the ongoing pandemic, it is critical to understand how natural immunity against SARS-CoV-2 and COVID-19 develops. We have identified that subjects with more severe COVID-19 disease mount a more robust and neutralizing antibody response against SARS-CoV-2 spike protein. Subjects who mounted a larger response against the spike also mounted antibody responses against other viral antigens, including the nucleocapsid protein and ORF8. Additionally, this study reveals that subjects with more severe disease mount a larger memory B cell response against the spike. These data suggest that subjects with more severe COVID-19 disease are likely better protected from reinfection with SARS-CoV-2.

Improved integration of single-cell transcriptome and surface protein expression by LinQ-View.

Multimodal advances in single-cell sequencing have enabled the simultaneous quantification of cell surface protein expression alongside unbiased transcriptional profiling. Here, we present LinQ-View, a toolkit designed for multimodal single-cell data visualization and analysis. LinQ-View integrates transcriptional and cell surface protein expression profiling data to reveal more accurate cell heterogeneity and proposes a quantitative metric for cluster purity assessment. Through comparison with existing multimodal methods on multiple public CITE-seq datasets, we demonstrate that LinQ-View efficiently generates accurate cell clusters, especially in CITE-seq data with routine numbers of surface protein features, by preventing variations in a single surface protein feature from affecting results. Finally, we utilized this method to integrate single-cell transcriptional and protein expression data from SARS-CoV-2-infected patients, revealing antigen-specific B cell subsets after infection. Our results suggest LinQ-View could be helpful for multimodal analysis and purity assessment of CITE-seq datasets that target specific cell populations (e.g., B cells).

Cutaneous leukocyte lineages in tolerant large animal and immunosuppressed clinical vascularized composite allograft recipients.

Vascularized composite allografts (VCAs) can restore fully functional anatomic units in patients with limb amputations or severe facial tissue loss. However, acute rejection of the skin is frequently observed and underscores the importance of developing tolerance induction protocols. In this study, we have characterized the skin immune system in VCAs. We demonstrate infiltration of recipient leukocytes, regardless of rejection status, and in tolerant mixed hematopoietic chimeras, the co-existence of these cells with donor leukocytes in the absence of rejection. Here we characterize the dermal T cell and epidermal Langerhans cell components of the skin immune system in our porcine model of VCA tolerance, and the kinetics of cutaneous chimerism in both of these populations in VCAs transplanted to tolerant and nontolerant recipients, as well as in host skin. Furthermore, in biopsies from the first patient to receive a hand transplant in our program, we demonstrate the presence of recipient T cells in the skin of the transplanted limb in the absence of clinical or histological evidence of rejection.

Distinct B cell subsets give rise to antigen-specific antibody responses against SARS-CoV-2.

Discovery of durable memory B cell (MBC) subsets against neutralizing viral epitopes is critical for determining immune correlates of protection from SARS-CoV-2 infection. Here, we identified functionally distinct SARS-CoV-2-reactive B cell subsets by profiling the repertoire of convalescent COVID-19 patients using a high-throughput B cell sorting and sequencing platform. Utilizing barcoded SARS-CoV-2 antigen baits, we isolated thousands of B cells that segregated into discrete functional subsets specific for the spike, nucleocapsid protein (NP), and open reading frame (ORF) proteins 7a and 8. Spike-specific B cells were enriched in canonical MBC clusters, and monoclonal antibodies (mAbs) from these cells were potently neutralizing. By contrast, B cells specific to ORF8 and NP were enriched in naïve and innate-like clusters, and mAbs against these targets were exclusively non-neutralizing. Finally, we identified that B cell specificity, subset distribution, and affinity maturation were impacted by clinical features such as age, sex, and symptom duration. Together, our data provide a comprehensive tool for evaluating B cell immunity to SARS-CoV-2 infection or vaccination and highlight the complexity of the human B cell response to SARS-CoV-2.

SARS-CoV-2 infection severity is linked to superior humoral immunity against the spike.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is currently causing a global pandemic. The antigen specificity and kinetics of the antibody response mounted against this novel virus are not understood in detail. Here, we report that subjects with a more severe SARS-CoV-2 infection exhibit a larger antibody response against the spike and nucleocapsid protein and epitope spreading to subdominant viral antigens, such as open reading frame 8 and non-structural proteins. Subjects with a greater antibody response mounted a larger memory B cell response against the spike, but not the nucleocapsid protein. Additionally, we revealed that antibodies against the spike are still capable of binding the D614G spike mutant and cross-react with the SARS-CoV-1 receptor binding domain. Together, this study reveals that subjects with a more severe SARS-CoV-2 infection exhibit a greater overall antibody response to the spike and nucleocapsid protein and a larger memory B cell response against the spike.

Local Immunosuppression for Vascularized Composite Allografts: Application of Topical FK506-TyroSpheres in a Nonhuman Primate Model.

Transplantation of vascularized composite allografts (VCAs) provides a means of restoring complex anatomical and functional units following burns and other disfigurement otherwise not amenable to conventional autologous reconstructive surgery. While short- to intermediate-term VCA survival is largely dependent on patient compliance with medication, the myriad of side effects resulting from lifelong systemic immunosuppression continue to pose a significant challenge. Topical immunosuppression is therefore a logical and attractive alternative for VCA. Current formulations are limited though, by poor skin penetration but this may be mitigated by conjugation of immunosuppressive drugs to TyroSpheres for enhanced delivery. Therefore, we investigated the topical application of FK506-TyroSpheres (in the form of a gel dressing) in a clinically relevant nonhuman primate VCA model to determine if allograft survival could be prolonged at reduced levels of maintenance systemic immunosuppression. Six Major Histocompatibility Complex (MHC)-mismatched cynomolgus macaques (Macaca fascicularis) served as reciprocal donors and recipients of radial forearm fasciocutaneous flaps. Standard Bacitracin ointment and FK506-TyroSpheres were applied every other day to the VCAs of animals in groups 1 (controls, n = 2) and 2 (experimental, n = 4), respectively, before gradual taper of systemic FK506. Clinical features of VCA rejection still developed when systemic FK506 fell below 10 ng/ml despite application of FK506-TyroSpheres and prolonged VCA survival was not achieved. However, unwanted systemic FK506 absorption was avoided with TyroSphere technology. Further refinement to optimize local drug delivery profiles to achieve and maintain therapeutic delivery of FK506 with TyroSpheres is underway, leveraging significant experience in controlled drug delivery to mitigate acute rejection of VCAs.

Topical Delivery of Immunosuppression to Prolong Xenogeneic and Allogeneic Split-Thickness Skin Graft Survival.

Cadaveric skin allograft is the current standard of treatment for temporary coverage of large burn wounds. Porcine xenografts are viable alternatives but undergo α-1,3-galactose (Gal)-mediated hyperacute rejection and are lost by post-operative day (POD) 3 because of naturally occurring antibodies to Gal in primate recipients. Using baboons, we previously demonstrated that xenografts from GalT-KO swine (lacking Gal) provided wound coverage comparable with allografts with systemic immunosuppression. In this study, we investigate topical immunosuppression as an alternative to prolong xenograft survival. Full-thickness wounds in baboons were created and covered with xenogeneic and allogeneic split-thickness skin grafts (STSGs). Animals were treated with slow-release (TyroSphere-encapsulated) topical formulations (cyclosporine-A [CSA] or Tacrolimus) applied 1) directly to the STSGs only, or 2) additionally to the wound bed before STSG and 1). Topical CSA did not improve either xenograft or allograft survival (median: treated grafts = 12.5 days, control = 14 days; P = 0.27) with similar results when topical Tacrolimus was used. Pretreatment of wound beds resulted in a significant reduction of xenograft survival compared with controls (10 vs 14 days; P = 0.0002), with comparable results observed in allografts. This observation was associated with marked reduction of inflammation on histology with Tacrolimus and not CSA. Prolongation of allograft and xenograft survival after application to full-thickness wound beds was not achieved with the current formulation of topical immunosuppressants. Modulation of inflammation within the wound bed was effective with Tacrolimus pretreatment before STSG application and may serve as a treatment strategy in related fields.