A novel assessment model for teaching robot-assisted living donor nephrectomy in abdominal transplant surgery fellowship.

BACKGROUND: An increasing number of transplant centers have adopted robot-assisted living donor nephrectomy. Thus, a transplant fellow assessment tool is needed for promoting operative independence in an objective and safe manner. METHODS: In this pilot study, data was prospectively collected on both fellow performance with focus on technique, efficiency, and communication ("overall RO-SCORE"), and operative steps ("operative steps RO-SCORE"). Robotic user performance metrics were analyzed from the da Vinci Xi system, including fellow percent active control time (ACT) and handoff counts. RESULTS: From July 2020 to February 2021, twenty-one robot-assisted donor nephrectomies were performed. In regression analysis, fellow performance (based on both RO-SCOREs and robot % ACT) was significantly associated with both time and case number, with time-to-independence modelled at 8.4-14.2 months, and case number-to-independence estimated at 15-22 cases. Robot user metrics provided valid objective measures alongside RO-SCOREs. CONCLUSIONS: This pilot study provides an effective assessment tool for promoting operative competency in robot-assisted donor nephrectomy among transplant fellows.

Changing Education Paradigms: Training Transplant Fellows for High Stake Procedures.

OBJECTIVE: Living kidney donation is a unique operation, as healthy patients are placed at risks inherent with major surgery without physical benefit. The ethical implications associated with any morbidity make it a high-stakes procedure. Fellowships are faced with the dilemma of optimizing fellow training in this demanding procedure while providing safe outcomes to donors. The Laparoscopic Living Donor Nephrectomy (LDN) Workshop is a resource that can provide intense instruction to help bridge the training deficit. Our aim was to examine the course's effectiveness in improving fellows' skill and confidence related to implementing LDN into future practice. METHODS: From 2017 to 2018, 36 abdominal transplant surgery fellows participated in a 2-day workshop consisting of live surgery observation, cadaver lab, and didactic sessions. Surveys were completed precourse, postcourse, and at 3-month postcourse follow-up. RESULTS: Preworkshop, 61% of participants reported less than 50% confidence in independent performance of LDN. Following workshop completion, 95% reported improved confidence. At 3-month follow-up, there was a 30% (p < 0.05) increase in median confidence level. Immediately following the course, 67% reported improved ability to analyze kidneys prior to donation, 74% changed the way donor candidates were evaluated, and 67% reported enhanced ability to risk stratify donors. Eighty-five percent felt it strengthened operative techniques with 70% implementing new diagnostic treatments and surgical strategies. Seventy percent of participants felt it improved their communication with colleagues and 67% had enhanced communication with patients. These trends were maintained at 3-month follow-up. CONCLUSION: These results indicated that the LDN Workshop improves confidence and increases fellows' skillset in a high-stakes procedure. The LDN Workshop is a useful adjunct to fellowship training to optimize successful, efficient, and safe performance of a demanding procedure in a uniquely healthy donor population.

Liver Transplantation and Bariatric Surgery: Timing and Outcomes.

Nonalcoholic steatohepatitis (NASH) is projected to become the leading indication for liver transplantation (LT) in the next decade in the United States. Strategies to treat the underlying etiology of NASH, which is almost always obesity, are being pursued. One such strategy is the utilization of bariatric surgery (BS) in the peritransplant period. The use of BS prior to LT could prevent the progression of NASH and abrogate the need for LT. BS at the time of LT or postoperatively has the potential to not only improve obesity-associated conditions such as diabetes, but also the potential to influence the incidence of NASH in the post-LT setting. However, there continues to be no consensus on the use and timing of BS in this patient population. This review aims to discuss the current literature and possible future action.

Sphingosine rescues aged mice from pulmonary pseudomonas infection.

BACKGROUND: Bacterial lung infection is a leading cause of death for those 65 y or older, often requiring intensive care unit admission and mechanical ventilation, which consumes considerable health care resources. Although administration of antibiotics is the standard of care for bacterial pneumonia, its overuse has led to the emergence of multidrug resistant organisms. Therefore, alternative strategies to help minimize the effects of bacterial pneumonia in the elderly are necessary. As studies have shown that sphingosine (SPH) has inherent bacterial killing properties, our goal was to assess whether it could act as a prophylactic treatment to protect aged mice from pulmonary infection by Pseudomonas aeruginosa. METHODS: Aged (51 wk) and young (8 wk) C57Bl/6 mice were used in this study. Pulmonary SPH levels were determined by histology. SPH content of microparticles was quantified using a SPH kinase assay. Pneumonia was induced by intranasally treating mice with 106 Colony Forming Unit (CFU) P aeruginosa. Microparticles were isolated from young mice, whereas some were further incubated with SPH. RESULTS: We observed that SPH levels are reduced in the bronchial epithelial cells as well as the bronchoalveolar lavage microparticles isolated from aged mice, which correlates with a susceptibility to infection. We demonstrate that SPH or microparticle treatment can protect aged mice from pulmonary P aeruginosa infection. Finally, we observed that enriching microparticles with SPH before treatment eliminated the bacterial load in P aeruginosa-infected aged mice. CONCLUSIONS: These data suggest that prophylactic treatment with SPH could reduce lung bacterial infections for the at-risk elderly population.

Balance Between the Proinflammatory and Anti-Inflammatory Immune Responses with Blood Transfusion in Sepsis.

Blood product transfusion may exacerbate the initial immunosuppressive response of sepsis. Nurses and other patient care providers must be diligent in recognizing and managing a worsening immune status, using flow cytometry to monitor patients' immune status. This type of monitoring may be instrumental in reducing morbidity and mortality in persons with sepsis. This article discusses the recent literature on the associated inflammatory responses that occur with blood transfusion and provides an analysis of alterations in key inflammatory pathways in response to transfusion in a sepsis population.

Bronchoalveolar Lavage Microvesicles Protect Burn-Injured Mice from Pulmonary Infection.

BACKGROUND: Pseudomonas aeruginosa is a major cause of morbidity and mortality among burn patients, despite antibiotic therapy. There is a need to identify innate immune defenses that prevent P aeruginosa infection in injured adults in an effort to find therapeutic alternatives to antibiotics. Here, we tested our hypothesis that microvesicles (MVs) in bronchoalveolar (BAL) fluid have a role in the immunity of the lung in response to pathogens. STUDY DESIGN: Microvesicles were isolated from murine BAL fluid, quantified using Nanoparticle Tracking Analysis, and injected into burn-injured mice before P aeruginosa infection. Survival was assessed and BAL bacterial loads enumerated. Neutrophil number and interleukin 6 activity were determined. Lungs were harvested and sphingosine (SPH) content analyzed via immunohistochemistry. Antimicrobial effects of MVs and SPH-enriched MVs were assessed in an in vitro assay. RESULTS: Burn-injured mice have reduced BAL MV number and SPH content compared with sham. When BAL MVs from healthy mice are administered to injured mice, survival and bacterial clearance are improved robustly. We also observed that intranasal administration of MVs restores SPH levels after burn injury, MVs kill bacteria directly, and this bacterial killing is increased when the MVs are supplemented with SPH. CONCLUSIONS: Using a preclinical model, BAL MVs are reduced after scald injury and BAL MV restoration to injured mice improves survival and bacterial clearance. The antimicrobial mechanisms leading to improved survival include the quantity and SPH content of BAL MVs.

Neutrophil derived microparticles increase mortality and the counter-inflammatory response in a murine model of sepsis.

Although advances in medical care have significantly improved sepsis survival, sepsis remains the leading cause of death in the ICU. This is likely due to a lack of complete understanding of the pathophysiologic mechanisms that lead to dysfunctional immunity. Neutrophil derived microparticles (NDMPs) have been shown to be the predominant microparticle present at infectious and inflamed foci in human models, however their effect on the immune response to inflammation and infection is sepsis has not been fully elucidated. As NDMPs may be a potential diagnostic and therapeutic target, we sought to determine the impact NDMPs on the immune response to a murine polymicrobial sepsis. We found that peritoneal neutrophil numbers, bacterial loads, and NDMPs were increased in our abdominal sepsis model. When NDMPs were injected into septic mice, we observed increased bacterial load, decreased neutrophil recruitment, increased expression of IL-10 and worsened mortality. Furthermore, the NDMPs express phosphatidylserine and are ingested by F4/80 macrophages via a Tim-4 and MFG-E8 dependent mechanism. Finally, upon treatment, NDMPs decrease macrophage activation, increase IL-10 release and decrease macrophage numbers. Altogether, these data suggest that NDMPs enhance immune dysfunction in sepsis by blunting the function of neutrophils and macrophages, two key cell populations involved in the early immune response to infection. This article is part of a Special Issue entitled: Immune and Metabolic Alterations in Trauma and Sepsis edited by Dr. Raghavan Raju.

Burn injury influences the T cell homeostasis in a butyrate-acid sphingomyelinase dependent manner.

Following burn injury, a key factor for patients susceptible to opportunistic infections is immune suppression. Butyrate levels are important in maintaining a functional immune system and these levels can be altered after injury. The acid sphingomyelinase (Asm) lipid signaling system has been implicated in a T cell actions with some evidence of being influenced by butyrate. Here, we hypothesized that burn-injury changes in butyrate levels would mediate Asm activity and, consequently, T cell homeostasis. We demonstrate that burn injury temporally decreases butyrate levels. We further determined that T cell Asm activity is increased by butyrate and decreased after burn injury. We additionally observed decreased T cell numbers in Asm-deficient, burn-injured, and microbiota-depleted mice. Finally, we demonstrate that butyrate reduced T cell death in an Asm-dependent manner. These data suggest that restoration of butyrate after burn injury may ameliorate the T cell lost observed in burn-injured patients by Asm regulation.

Dynamic Article: Percutaneous Nerve Evaluation Versus Staged Sacral Nerve Stimulation for Fecal Incontinence.

BACKGROUND: Sacral neuromodulation using a 2-staged approach is an established therapy for fecal incontinence. Office-based percutaneous nerve evaluation is a less-invasive alternative to the stage 1 procedure but is seldom used in the evaluation of patients with fecal incontinence. OBJECTIVE: The aim of this study was to determine the clinical success of percutaneous nerve evaluation versus a staged approach. DESIGN: This was a retrospective review of a prospectively maintained, single-institution database of patients treated with sacral neuromodulation for fecal incontinence. SETTINGS: This study was conducted at a single academic medical center. PATIENTS: Eighty-six consecutive patients were treated with sacral neuromodulation for fecal incontinence. INTERVENTIONS: Percutaneous nerve evaluation was compared with a staged approach. MAIN OUTCOME MEASURES: The primary outcome measured was the proportion of patients progressing to complete implantation based on >50% improvement in Wexner score during the testing phase. RESULTS: Percutaneous nerve evaluation was performed in 45 patients, whereas 41 underwent a staged approach. The mean baseline Wexner score did not differ between testing groups. Success was similar between the staged approach and percutaneous nerve evaluation (90.2% versus 82.2%; p = 0.36). The mean 3-month Wexner score was not significantly different between testing methods (4.4 versus 4.1; p = 0.74). However, infection was more likely to occur after the staged approach (10.5% versus 0.0%; p < 0.05). LIMITATIONS: This study was limited by its retrospective nature and potential for selection bias. CONCLUSIONS: Percutaneous nerve evaluation offers a viable alternative to a staged approach in the evaluation of patients for sacral neuromodulation in the setting of fecal incontinence. Not only are success rates similar, but percutaneous nerve evaluation also has the benefit of limiting patients to 1 operating room visit and has lower rates of infection as compared with the traditional staged approach for sacral neuromodulation.

Frontline Science: Sphingosine rescues burn-injured mice from pulmonary Pseudomonas aeruginosa infection.

Burn patients with concomitant pulmonary Pseudomonas aeruginosa (PA) infection have mortality rates as high as 50%, despite antibiotic therapy. Sphingosine is generated from ceramide via ceramidase and has been reported to have antimicrobial properties. We observed a reduction in sphingosine and a concurrent increase in ceramide in bronchial epithelial cells after burn injury. After PA inoculation, these mice had a significant decrease in survival compared to noninjured mice. However, when injured mice were pretreated with sphingosine or neutral ceramidase and subsequently infected, mortality and bacterial levels were robustly reduced. We further observed that sphingosine directly kills PA. Together, these results demonstrate that reduction in sphingosine is associated with an increased susceptibility to pulmonary infection after burn injury. Restoration of sphingosine levels through direct sphingosine administration or conversion of the increased ceramide to sphingosine by neutral ceramidase reduces mortality and mitigates pulmonary infection after burn injury.

Amitriptyline Usage Exacerbates the Immune Suppression Following Burn Injury.

Currently, over 10% of the US population is taking antidepressants. Numerous antidepressants such as amitriptyline are known to inhibit acid sphingomyelinase (Asm), an enzyme that is known to mediate leukocyte function and homeostasis. Severe burn injury can lead to an immunosuppressive state that is characterized by decreased leukocyte function and numbers as well as increased susceptibility to infection. Based upon the intersection of these facts, we hypothesized that amitriptyline-treated, scald-injured mice would have an altered immune response to injury as compared with untreated scald mice. Prior to burn, mice were pretreated with amitriptyline. Drug- or saline-treated mice were subjected full thickness dorsal scald- or sham-injury. Immune cells from spleen, thymus, and bone marrow were subsequently harvested and characterized. We first observed that amitriptyline prior to burn injury increased body mass loss and spleen contraction. Both amitriptylinetreatment and burn injury resulted in a 40% decrease of leukocyte Asm activity. Following scald injury, we demonstrate increased reduction of lymphocyte precursors in the bone marrow and thymus, as well as mature leukocytes in the spleen in mice that were treated with amitriptyline. We also demonstrate that amitriptyline treatment prior to injury reduced neutrophil accumulation following peptidoglycan stimulus in scald-injured mice. These data show that Asm alterations can play a significant role in mediating alterations to the immune system after injury. The data further suggest that those taking antidepressants may be at a higher risk for complications following burn injury.

Fecal Microbiota Transplant Restores Mucosal Integrity in a Murine Model of Burn Injury.

The gut microbiome is a community of commensal organisms that are known to play a role in nutrient production as well as gut homeostasis. The composition of the gut flora can be affected by many factors; however, the impact of burn injury on the microbiome is not fully known. Here, we hypothesized that burn-induced changes to the microbiome would impact overall colon health. After scald-burn injury, cecal samples were analyzed for aerobic and anaerobic colony forming units, bacterial community, and butyrate levels. In addition, colon and total intestinal permeabilities were determined. These parameters were further determined in a germ-reduced murine model. Following both burn injury and germ reduction, we observed decreases in aerobic and anaerobic bacteria, increased colon permeability and no change to small intestinal permeability. After burn injury, we further observed a significant decrease in the butyrate producing bacteria R. Gnavus, C. Eutactus, and Roseburia species as well as decreases in colonic butyrate. Finally, in mice that underwent burn followed by fecal microbiota transplant, bacteria levels and mucosal integrity were restored. Altogether our data demonstrate that burn injury can alter the microbiome leading to decreased butyrate levels and increased colon permeability. Of interest, fecal microbiota transplant treatment was able to ameliorate the burn-induced changes in colon permeability. Thus, fecal transplantation may represent a novel therapy in restoring colon health after burn injury.

Impact of Platelets and Platelet-Derived Microparticles on Hypercoagulability Following Burn Injury.

An acute burn induced coagulopathy develops after scald injury, which evolves into a subacute, hypercoagulable state. Microparticles, specifically platelet-derived MPs (PMPs), have been suggested as possible contributors. We first developed a model of burn-induced coagulopathy and then sought to investigate the role of platelets and PMPs in coagulation after burn. We hypothesized that changes in circulating platelet and PMP populations after injury would contribute to the post-burn, hypercoagulable state. A murine scald model with 28% TBSA full thickness burn injury was utilized and blood samples were collected at intervals after injury. Circulating MP populations, platelet counts, overall coagulation, and platelet function were determined. Burn injury led to hypercoagulability on post-burn day one (PBD1), which persisted 6 days after injury (PBD6). On PBD1, there was a significant decrease in platelet numbers and a decline in platelet contribution to clot formation with a concomitant increase in circulating procoagulant PMPs. On PBD6, there was a significant increase in platelet numbers and in platelet activation with no change in PMPs compared with sham. Further, on PBD1 decreased ADP-induced platelet activation was observed with a contrasting increase in ADP-induced platelet activation on PBD6. We therefore concluded that there was a temporal change in the mechanisms leading to a hypercoagulable state after scald injury, that PMPs are responsible for changes seen on PBD1, and finally that ADP-induced platelet activation was key to the augmented clotting mechanisms 6 days after burn.

Impact of caspase-8 and PKA in regulating neutrophil-derived microparticle generation.

The morbidity and mortality from sepsis continues to remain high despite extensive research into understanding this complex immunologic process. Further, while source control and antibiotic therapy have improved patient outcomes, many immunologically based therapies have fallen short. Microparticles (MPs) are intact vesicles that serve as mediators of intercellular communication as well as markers of inflammation in various disease processes. We have previously demonstrated that MPs can be produced at the infected foci during sepsis, are predominantly of neutrophil derivation (NDMPs) and can modulate immune cells. In this study, we sought to elucidate the molecular mechanisms underlying NDMP generation. Using thioglycolate (TGA) to recruit and activate neutrophils, we first determined that intra-peritoneal TGA increase NDMP accumulation. We next utilized TGA-elicited neutrophils in vitro to investigate signaling intermediates involved in NDMP production, including the intrinsic and extrinsic caspase pathways, cAMP dependent PKA and Epac activation as well as the role myosin light chain kinase (MLCK) as a final mediator of NDMP release. We observed that NDMP generation was dependent on the extrinsic caspase apoptotic pathway (caspase 3 and caspase 8), cAMP activation of PKA but not of Epac, and on activation of MLCK. Altogether, these data contribute to an overall framework depicting the molecular mechanisms that regulate NDMP generation.

Peritoneal wash contents used to predict mortality in a murine sepsis model.

BACKGROUND: Cecal ligation and puncture (CLP) is considered the gold standard for inducing abdominal sepsis in mice. However, the model lacks source control, a component of sepsis management in humans. Using a CLP-excision model, we characterized peritoneal cytokines and cells and hypothesized these analyses would allow us to predict survival. METHODS: Fifty-eight mice were first subjected to CLP. Twenty hours later, the necrotic cecums were debrided, abdominal cavity lavaged, and intraperitoneal antibiotics administered. Peritoneal cytokines and leukocytes collected from the peritoneal lavage were analyzed. These immune parameters were used to generate receiver operator characteristic curves. In separate experiments, the accuracy of the model was verified with a survival cohort. Finally, we collected the peritoneal lavage and analyzed both serum and peritoneal cytokines, bacterial load, and leukocyte functionality. RESULTS: Peritoneal interleukin (IL)-6 levels and neutrophil CD11b intensity were observed to be significantly different in mice that lived versus those who died. In separate experiments, mice predicted to live (P-LIVE) had decreased bacterial loads, systemic IL-10, and neutrophil oxidative burst and increased peritoneal inflammatory monocyte numbers and phagocytosis. CONCLUSIONS: This study couples a clinically relevant sepsis model with methodology to limit pathogen spread. Using surgical waste, stratification of the mice into groups P-LIVE and predicted to die was possible with a high degree of accuracy and specificity. In mice P-LIVE, increased inflammatory monocyte recruitment and phagocytosis were associated with decreased systemic IL-10 and bacterial loads.