NLRP1B allele 2 does not respond to Val-boro-Pro (VbP) in intestinal epithelial cells.

The intestinal mucosa must balance tolerance to commensal microbes and luminal antigens with rapid detection of enteric pathogens in order to maintain homeostasis. This balance is facilitated through the regulation of epithelial layer integrity by innate immune receptors. Certain NOD-like receptors (NLRs) expressed in intestinal epithelial cells, including NLRC4 and NLRP9B, form inflammasomes that protect against pathogens by activating caspase-1 to cause extrusion of infected cells. NLRP1B is a murine NLR encoded by five alleles of a highly polymorphic gene homologous to human NLRP1. NLRP1B forms inflammasomes in response to a variety of pathogens that cause intestinal infections, but it has almost exclusively been studied in immune cells and has not been characterized in cells of the intestinal epithelium. Here, we show that Nlrp1b allele 2 is expressed in ileal and colonic organoids derived for C57BL/6J mice, while the related gene Nlrp1a was not expressed. Nlrp1b was upregulated by interleukin-13 in organoids and by the protozoan Tritrichomonas muris in vivo, suggesting that NLRP1B may be involved in defense against enteric parasites. Surprisingly, while Val-boro-Pro (VbP) activated C57BL/6J-derived bone marrow-derived macrophages, which expressed both Nlrp1a and Nlrp1b, it did not activate intestinal organoids of the same genotype. We furthermore did not detect Nlrp1b in organoids derived from Balb/cJ mice, which express a different allele than the one expressed in C57BL/6J mice. Together, our results suggest that NLRP1B may have an allele-dependent function in murine IECs whose regulation is distinct from that of macrophages, and that the response to VbP might be exclusively driven by NLRP1A in C57BL/6J mice.

Factors Associated with Opting Out of an Unrelated Hematopoietic Stem Cell Donor Registry: Differences and Similarities across Five Key Groups of Young Race/Ethnically Diverse Potential Donors in the United States.

Young adults from underserved racial/ethnic groups are critically needed as unrelated hematopoietic stem cell (HSC) donors, yet they are more likely than other groups to opt out of donation after having matched a patient. Understanding which factors are most strongly associated with opting out among young underserved racial/ ethnic registered donors compared with their White counterparts will provide the basis for specific interventions to improve donor retention. We sought to determine the key, modifiable psychosocial, registry-related, and donation-related characteristics that are uniquely associated with opting out across 5 key racial/ethnic groups of young HSC donor registry members who had been contacted as a potential match for a patient. This study examines data from a large cross-sectional survey of young (age 18 to 30) registry members shortly after they preliminarily matched a patient (CT-stage) and continued toward or opted out of donation (CT-C and CT-NI), stratified by racial/ethnic group and sex. We assessed psychosocial, registry-related, and donation-related characteristics for all participants. We used chi-squared and F tests to assess differences between racial/ethnic groups. A separate logistic regression analysis for each racial/ethnic group was conducted to quantify adjusted associations between each variable and opting out. Then, we compared these associations across the racial/ethnic groups by evaluating the interaction effect between each variable and racial/ethnic group, with the same outcome (CT-C versus CT-NI) in question. Nine hundred thirty-five participants were surveyed, including 284 White, 165 Hispanic, 191 Black, 192 Asian/Pacific Islander, and 103 Multiracial/multiethnic participants. There were significant differences across racial/ethnic groups in values/goals, religious objections to donation, HSC-related medical mistrust, and parental involvement in donation decisions. Adjusted logistic regression subgroup analyses indicated that ambivalence was strongly associated with opting out across all racial/ethnic groups. Greater focus on intrinsic life goals (e.g., raising a family, becoming a community leader, influencing social values) was associated with opting out in the Multiracial/multiethnic, Hispanic, and Asian/Pacific Islander groups. Healthcare mistrust and insufficient registry contact was a significant factor for Hispanic participants. Protective factors against opting out included remembering joining the registry (Black participants), and parental support for donation decision (Asian/Pacific Islander participants). The performance of each logistic regression model was strong, with area-under-the curve ≥.88, CT-stage outcome classification accuracy ≥89%, and good fit between expected and observed opt-out probabilities. In the analysis across different racial/ethnic groups, the only significant interaction was race/ethnicity by whether more contact with the registry would have changed the decision at CT-stage; this variable was significant only for the Hispanic group. In the within-group analysis for Hispanic participants, the "more registry contact" variable was strongly associated with opting out (odds ratio 5.8, P = .03). Consistent with a growing body of HSC donor research, ambivalence was a key factor associated with opting-out for all racial/ethnic groups. Other key variables were differentially associated with opting-out depending on racial/ethnic group. Our study highlights key variables that registries should focus on as they develop targeted and tailored strategies to enhance commitment and reduce attrition of potential donors.

TRAV26-2 T-Cell Receptor Expression Is Associated With Mucosal Lymphocyte Response to Wheat Proteins in Patients With Functional Dyspepsia.

INTRODUCTION: An association between functional dyspepsia (FD) and wheat-containing foods has been reported in observational studies; however, an adaptive response has not been demonstrated. We examined whether antigens present in wheat could provoke a response from FD duodenal lymphocytes. METHODS: Lamina propria mononuclear cells (LPMCs) were isolated from duodenal biopsies from 50 patients with FD and 23 controls. LPMCs were exposed to gluten (0.2 mg/mL) or gliadin (0.2 mg/mL) for 24 hours. Flow cytometry was performed to phenotype lymphocytes. Quantitative PCR was used to measure the expression of gliadin-associated T-cell receptor alpha variant ( TRAV ) 26-2. RESULTS: In response to gliadin (but not gluten) stimulation, the effector Th2-like population was increased in FD LPMCs compared with that in controls and unstimulated FD LPMCs. Duodenal gene expression of TRAV26- 2 was decreased in patients with FD compared with that in controls. We identified a positive association between gene expression of this T-cell receptor variant and LPMC effector Th17-like cell populations in patients with FD, but not controls after exposure to gluten, but not gliadin. DISCUSSION: Our findings suggest that gliadin exposure provokes a duodenal effector Th2-like response in patients with FD, supporting the notion that food antigens drive responses in some patients. Furthermore, these findings suggest that altered lymphocyte responses to wheat proteins play a role in FD pathogenesis.

Factors Associated with Opting Out of Donation among Registered Young Unrelated Hematopoietic Stem Cell Donors.

Young adults enrolled in hematopoietic stem cell (HSC) donation registries, including the Be The Match registry in the United States, often opt out of the registry when a potential recipient is identified. This results in a limited supply from the most desirable allogeneic source of HSCs used in transplantation to treat serious health conditions. The differences in demographic, psychosocial, registry-related, and donation-related characteristics between those who continue to donation and those who opt out may elucidate the modifiable risk factors for attrition, but these characteristics have not been extensively studied in young donors up to age 30 years. Our goal was to study demographic, psychosocial, registry-related, and donation-related characteristics in a group of young HSC donor registry members who had recently been contacted about a potential recipient, to determine the key characteristics that differ between those who continued toward donation and those who opted out and to examine the extent of these differences. We conducted a cross-sectional survey in a random sample of young (age 18 to 30 years) current and former registry members, stratified by race/ethnicity and sex. Demographic, psychosocial (eg, life goals, HSC allocation mistrust), registry-related (eg context and motive for joining the registry), and donation-related (eg, ambivalence, religious objections to donation, knowledge about donation) characteristics were assessed. Chi-square and 2-sample t tests were used to examine differences between those who continued (CT-C group) and those who opted out (CT-NI group). Hierarchical logistic regression was used to estimate adjusted covariate effects on the odds of opting out. A total of 935 participants were surveyed. Donation-related knowledge was higher in the CT-C group than in the CT-NI group. HSC allocation mistrust, religious objections, and concerns about donation were higher in the CT-NI group. After adjusting for covariates in a logistic regression model, we found that having more intrinsic life goals, having more ambivalence, and talking with registry staff only once/twice were significantly associated with opting out of the registry. Ambivalence had the strongest association with opting out. In contrast, remembering joining the registry, believing that parents would support donation, and having medical concerns were significantly associated with continuing toward donation. This effect of medical concerns on donation was discovered only after adjusting for the related but distinct ambivalence variable, with the remaining effect of medical concerns relating to engagement with the donation process and information-seeking. The model had strong discriminative ability (area under the receiver operating characteristic curve = .92) and classification accuracy (86.6%). Our data indicate that among young adult members of a national HSC donor registry, ambivalence and limited contact with registry staff were more strongly associated with opting out of donation. Medical concerns were associated with continuing toward donation. Further studies are needed to confirm a causal link between medical concerns and continuing to donation among young donors. Our study suggests that these concerns might not be directly related to attrition, whereas other factors (eg, ambivalence, low donation-related knowledge) are associated with attrition and thus should be targeted for attrition reduction strategies.

The Impact of Pre-Apheresis Health Related Quality of Life on Peripheral Blood Progenitor Cell Yield and Donor's Health and Outcome: Secondary Analysis of Patient-Reported Outcome Data from the RDSafe and BMT CTN 0201 Clinical Trials.

There is a lack of evidence about how health-related quality of life (HRQoL), including psychosocial factors, might affect donation-related experiences and clinical markers in the context of hematopoietic stem cell donation. The broader literature suggests that psychological factors, including anxiety and depression, are associated with higher levels of inflammatory burden leading to poorer postprocedural outcomes including longer hospital stays and increased pain perception. In this study, we aimed to evaluate whether predonation HRQoL markers predict toxicity profile and stem cell yield after peripheral blood stem cell (PBSC) donation in healthy donors. The study population comprised adult granulocyte colony-stimulating factor mobilized PBSC-related donors (RD) (n = 157) and unrelated donors (URD) (n = 179) enrolled in the related donor safety study (RDSafe) and Blood and Marrow Transplant Clinical Trials Network (BMT CTN) 0201 clinical trials. Pre-donation HRQoL was assessed using the Short-Form-12 (SF-12) in RDSafe and SF-8 questionnaire in BMT CTN 0201 (higher score is better). The aims of this study were to (a) determine the impact of pre-donation HRQoL on peri-collection pain and acute toxicities experienced and (b) to investigate the pre-procedural HRQoL indicators on stem cells yield. URDs were younger than RDs (median age 35 versus 63). A higher proportion of RDs were female (50% versus 40%) and obese (41% versus 35%). A higher proportion of RD PBSC donations required 2 days or more of apheresis (44% versus 21%). More RD collections were lower volume procedures (<18L, 16% versus 28%), and required a central line (28% versus 11%). RDs were more likely to report pre-donation grade 1-2 pain (27% versus 8%) and other toxicities (16% versus 6%). Among RDs, a lower pre-donation physical component summary (PCS) score was associated with significantly more grade 2-4 pain at 1 month (P = .004) and at 1-year after donation (P = .0099) in univariable analyses. In multivariable analysis, pre-donation PCS remained significantly associated with grade 2-4 pain 1 month after donation (P = .0098). More specifically, RDs with predonation PCS scores in the highest quartile were less likely to report pain compared with donors with PCS scores in the lowest quartile (odds ratio 0.1; 95% confidence interval 0.01-0.83; P = .005). There was also a trend toward higher grade 2-4 pain at 1-year post-donation among RDs with lower predonation PCS score (P = .018). Among URDs, neither PCS nor mental component summary (MCS) scores were associated with pain or toxicities at any time point after donation based on the univariable analysis. Because of low rates of postdonation grade 2-4 pain and toxicities, multivariable analysis was not performed in the URD setting. Moreover, there was no correlation between preapheresis HRQoL score (PCS or MCS) and PBSC collection yield in either the RD or URD setting. Our study demonstrates that pre-donation HRQoL scores are significantly associated with the toxicity profile after PBSC donation in the RD setting, with adult RDs with lower predonation physical HRQoL experiencing higher levels of pain at 1 month and persisting up to 12 months after a PBSC collection procedure. There were no such associations found in URD. Our findings can help clinicians identify donors at higher risk of pain with donation, and lead to personalized information and interventions for specific donors. Lack of correlation between predonation HRQoL and stem cell yield may be due to a small sample size and warrants further evaluation.

Patient-Reported Experiences after Acute Kidney Injury across Multiple Health-Related Quality-of-Life Domains.

Background: Investigations of health-related quality of life (HRQoL) in AKI have been limited in number, size, and domains assessed. We surveyed AKI survivors to describe the range of HRQoL AKI-related experiences and examined potential differences in AKI effects by sex and age at AKI episode. Methods: AKI survivors among American Association of Kidney Patients completed an anonymous online survey in September 2020. We assessed: (1) sociodemographic characteristics; (2) effects of AKI-physical, emotional, social; and (3) perceptions about interactions with health care providers using quantitative and qualitative items. Results: Respondents were 124 adult AKI survivors. Eighty-four percent reported that the AKI episode was very/extremely impactful on physical/emotional health. Fifty-seven percent reported being very/extremely concerned about AKI effects on work, and 67% were concerned about AKI effects on family. Only 52% of respondents rated medical team communication as very/extremely good. Individuals aged 22-65 years at AKI episode were more likely than younger/older counterparts to rate the AKI episode as highly impactful overall (90% versus 63% younger and 75% older individuals; P=0.04), more impactful on family (78% versus 50% and 46%; P=0.008), and more impactful on work (74% versus 38% and 10%; P<0.001). Limitations of this work include convenience sampling, retrospective data collection, and unknown AKI severity. Conclusions: These findings are a critical step forward in understanding the range of AKI experiences/consequences. Future research should incorporate more comprehensive HRQoL measures, and health care professionals should consider providing more information in their patient communication about AKI and follow-up.

Racial and Ethnic Differences in Attitudes, Perceptions, and Knowledge about Unrelated Hematopoietic Stem Cell Donation: A Study of Younger Newly Recruited Potential Donors.

Attrition of young adult registry members is a significant issue impacting hematopoietic stem cell (HSC) donation registries, including the Be The Match registry in the US. The resulting limited supply of allogeneic HSCs, used to treat serious health conditions, has a stronger impact on racial/ethnic minority groups in the US. Compared with young white adults, young adults identifying with these minority groups are more likely to drop out of the donor registry when called to donate. However, the underlying psychosocial factors that differ between white and nonwhite registrants have not been fully investigated. The central goal of this study was to examine demographic, registry-related, and donation-related characteristics in a young, newly registered group of potential donors and to determine whether these characteristics differed by, or were distributed differently among, racial/ethnic groups. We conducted a cross-sectional survey in a random sample of young (age 18 to 30 years) newly registered members, stratified by racial/ethnic group and sex. Demographic, registry-related (eg, context and motive for joining the registry), and donation-related (eg, ambivalence, religious objections to donation, knowledge about donation) characteristics were assessed. The chi-square test and analysis of variance were used to examine differences among racial/ethnic groups. Discriminant function analysis was used to assess whether patterns of the 3 classes of characteristics were associated with membership in particular racial/ethnic groups. A total of 524 participants were surveyed. Joining online was most common among white individuals, whereas joining at college was most common among black and Hispanic individuals. Ambivalence toward donation was higher among Asian/Pacific Islanders compared with white or multiracial/multiethnic individuals. Discriminant function analysis revealed 4 psychosocial/attitudinal functions predicting membership in certain racial/ethnic groups. The function accounting for the most variance in responses included mistrust of HSC allocation, religious objections to donation, low parental support, and low knowledge level. This function discriminated significantly between the white and nonwhite groups. Another function also identified ambivalence as a discriminating factor, which was most strongly associated with Asian/Pacific Islanders. Among young adult members of an HSC donor registry, such factors as ambivalence, family concerns about donation, mistrust of HSC allocation, religious objections, and less knowledge about donation were more strongly associated with membership in the nonwhite groups compared with the white group. These factors are known to be associated with a higher risk of opting out after having been preliminarily matched with a patient. The finding that these characteristics are associated with racial/ethnic minority group membership provides targets for recruitment strategies aimed at improving retention of young registry members.

Defects in NLRP6, autophagy and goblet cell homeostasis are associated with reduced duodenal CRH receptor 2 expression in patients with functional dyspepsia.

Functional dyspepsia (FD) affects up to 15% of the population and is characterised by recurring upper gastrointestinal (GI) symptoms occurring in the absence of clinically identifiable pathology. Psychological stress is a key factor associated with the onset of FD and locally acting hypothalamic-pituitary-adrenal (HPA) axis hormones have been implicated in GI motility and barrier dysfunction. Recent pre-clinical work has identified mechanistic pathways linking corticotropin-releasing hormone (CRH) with the innate epithelial immune protein NLRP6, an inflammasome that has been shown to regulate GI mucus secretion. We recruited twelve FD patients and twelve healthy individuals to examine whether dysregulation of hypothalamic-pituitary adrenal (HPA) axis hormones and altered NLRP6 pathways were evident in the duodenal mucosa. Protein expression was assessed by immunoblot and immunohistochemistry in D2 duodenal biopsies. Plasma HPA axis hormones were assayed by ELISA and enteroid and colorectal cancer cell line cultures were used to verify function. FD patients exhibited reduced duodenal CRH-receptor 2, compared to non-GI disease controls, indicating a dysregulation of duodenal HPA signalling. The loss of CRH-receptor 2 correlated with reduced NLRP6 expression and autophagy function, processes critical for maintaining goblet cell homeostasis. In accordance, duodenal goblet cell numbers and mucin exocytosis was reduced in FD patients compared to controls. In vitro studies demonstrated that CRH could reduce NLRP6 in duodenal spheroids and promote mucus secretion in the HT29-MTX-E12 cell line. In conclusion, FD patients exhibit defects in the NLRP6-autophagy axis with decreased goblet cell function that may drive symptoms of disease. These features correlated with loss of CRH receptor 2 and may be driven by dysregulation of HPA signalling in the duodenum of FD patients.

Type 2 and type 17 effector cells are increased in the duodenal mucosa but not peripheral blood of patients with functional dyspepsia.

Background: Functional dyspepsia is characterised by chronic symptoms of post-prandial distress or epigastric pain not associated with defined structural pathology. Increased peripheral gut-homing T cells have been previously identified in patients. To date, it is unknown if these T cells were antigen-experienced, or if a specific phenotype was associated with FD. Objective: This study aimed to characterise T cell populations in the blood and duodenal mucosa of FD patients that may be implicated in disease pathophysiology. Methods: We identified duodenal T cell populations from 23 controls and 49 Rome III FD patients by flow cytometry using a surface marker antibody panel. We also analysed T cell populations in peripheral blood from 37 controls and 61 patients. Where available, we examined the number of duodenal eosinophils in patients and controls. Results: There was a shift in the duodenal T helper cell balance in FD patients compared to controls. For example, patients had increased duodenal mucosal Th2 populations in the effector (13.03 ± 16.11, 19.84 ± 15.51, p=0.038), central memory (23.75 ± 18.97, 37.52 ± 17.51, p=0.007) and effector memory (9.80±10.50 vs 20.53±14.15, p=0.001) populations. Th17 populations were also increased in the effector (31.74±24.73 vs 45.57±23.75, p=0.03) and effector memory (11.95±8.42 vs 18.44±15.63, p=0.027) subsets. Peripheral T cell populations were unchanged between FD and control. Conclusion: Our findings identify an association between lymphocyte populations and FD, specifically a Th2 and Th17 signature in the duodenal mucosa. The presence of effector and memory cells suggest that the microinflammation in FD is antigen driven.

Bioavailability of arsenic, cadmium, lead and mercury as measured by intestinal permeability.

In this study, the intestinal permeability of metal(loid)s (MLs) such as arsenic (As), cadmium (Cd), lead (Pb) and mercury (Hg) was examined, as influenced by gut microbes and chelating agents using an in vitro gastrointestinal/Caco-2 cell intestinal epithelium model. The results showed that in the presence of gut microbes or chelating agents, there was a significant decrease in the permeability of MLs (As-7.5%, Cd-6.3%, Pb-7.9% and Hg-8.2%) as measured by apparent permeability coefficient value (Papp), with differences in ML retention and complexation amongst the chelants and the gut microbes. The decrease in ML permeability varied amongst the MLs. Chelating agents reduce intestinal absorption of MLs by forming complexes thereby making them less permeable. In the case of gut bacteria, the decrease in the intestinal permeability of MLs may be associated to a direct protection of the intestinal barrier against the MLs or indirect intestinal ML sequestration by the gut bacteria through adsorption on bacterial surface. Thus, both gut microbes and chelating agents can be used to decrease the intestinal permeability of MLs, thereby mitigating their toxicity.

Pharmacological HIF-1 stabilization promotes intestinal epithelial healing through regulation of α-integrin expression and function.

Intestinal epithelia are critical for maintaining gastrointestinal homeostasis. Epithelial barrier injury, causing inflammation and vascular damage, results in inflammatory hypoxia, and thus, healing occurs in an oxygen-restricted environment. The transcription factor hypoxia-inducible factor (HIF)-1 regulates genes important for cell survival and repair, including the cell adhesion protein ß1-integrin. Integrins function as αß-dimers, and α-integrin-matrix binding is critical for cell migration. We hypothesized that HIF-1 stabilization accelerates epithelial migration through integrin-dependent pathways. We aimed to examine functional and posttranslational activity of α-integrins during HIF-1-mediated intestinal epithelial healing. Wound healing was assessed in T84 monolayers over 24 h with/without prolyl-hydroxylase inhibitor (PHDi) (GB-004), which stabilizes HIF-1. Gene and protein expression were measured by RT-PCR and immunoblot, and α-integrin localization was assessed by immunofluorescence. α-integrin function was assessed by antibody-mediated blockade, and integrin α6 regulation was determined by HIF-1α chromatin immunoprecipitation. Models of mucosal wounding and 2,4,6-trinitrobenzenesulfonic acid (TNBS)-induced colitis were used to examine integrin expression and localization in vivo. PHDi treatment accelerated wound closure and migration within 12 h, associated with increased integrin α2 and α6 protein, but not α3. Functional blockade of integrins α2 and α6 inhibited PHDi-mediated accelerated wound closure. HIF-1 bound directly to the integrin α6 promoter. PHDi treatment accelerated mucosal healing, which was associated with increased α6 immunohistochemical staining in wound-associated epithelium and wound-adjacent tissue. PHDi treatment increased α6 protein levels in colonocytes of TNBS mice and induced α6 staining in regenerating crypts and reepithelialized inflammatory lesions. Together, these data demonstrate a role for HIF-1 in regulating both integrin α2 and α6 responses during intestinal epithelial healing.NEW & NOTEWORTHY HIF-1 plays an important role in epithelial restitution, selectively inducing integrins α6 and α2 to promote migration and proliferation, respectively. HIF-stabilizing prolyl-hydroxylase inhibitors accelerate intestinal mucosal healing by inducing epithelial integrin expression.

Health-Related Quality-of-Life Comparison of Adult Related and Unrelated HSC Donors: An RDSafe Study.

Multiple investigations have documented the health-related quality-of-life (HRQoL) and donation-related experiences of unrelated donors (URDs), but similar investigations of the related donor (RD) experience have been less common. The central goal of this study was to longitudinally examine and compare HRQoL of RD and URD hematopoietic stem cell (HSC) donors from predonation through 1 year postdonation. This prospective investigation included adult HSC donors ages 18 to 60 years who donated bone marrow or peripheral blood stem cells at one of 48 geographically diverse US transplant/donor centers and completed HRQoL interviews at predonation and 4 weeks and 1 year postdonation. At predonation, related donors were less ambivalent about donation (t = -3.30; P = .001), more satisfied with their decision to donate (t = 2.65; P = .009), and more likely to define themselves as donors (t = 2.94; P = .004) than were URDs. However, related donors were more concerned about the use of needles (odds ratio [OR] = 2.19; P = .012), about who would pay for the procedure (OR = 2.80; P = .011), and the possibility that they would feel responsible if the transplant failed (t = 2.31; P = .022). Shortly postdonation, related donors were more likely to report donation-related pain (t = 2.50; P = .013) and lightheadedness (OR = 3.63; P = .028). At 1 year postdonation, related donors were less likely to be fully recovered from donation (OR = 0.10; P = .010) and more likely to report a longer recovery period following donation (t = 2.57; P = .011), although this latter finding was primarily due to the percentage of related versus unrelated donors not fully recovered at 1 year postdonation (10% versus 1%). Taken together, these findings suggest that current related donor management practices may be sufficient in preparing related donors for the psychological aspects of donation but that there may be more to do in terms of calibrating the description of donation-related experiences and recovery time to the related donor group (i.e., descriptions of donation experiences based on unrelated donation may not provide best estimates of experience for this group).

Role of Race/Ethnicity in Donor Decisions about Unrelated Hematopoietic Progenitor Cell Donation: Exploring Reasons for Higher Attrition among Racial/Ethnic Minorities.

There are more than 30 million potential unrelated hematopoietic progenitor cell (HPC) donors listed on international registries, but 30% to 50% are unavailable after matching a patient. In the United States racial/ethnic minorities opt out of donation at higher rates, and a previous study identified factors associated both with attrition and ethnic group membership. Attrition among minorities is also higher in the Anthony Nolan UK registry (35% in white British [WB] and 56% in nonwhite British [NWB]), but it is not clear what factors produce higher attrition in the United Kingdom and whether they are similar to those found in the United States. Three hundred fifty-seven UK potential donors who matched a patient completed a questionnaire. Key factors were compared by donation decision (continue or opt out) and by race/ethnicity (WB versus NWB). The pattern of UK results was compared with that of the previous US study for variables assessed in both studies. Across WB and NWB donors, higher attrition was associated with poorer physical/mental health, greater ambivalence, and more concerns about donation. Donors who opted out also reported less interaction with the registry, and 16% indicated that more interaction with the registry would have changed their decision. Those opting out of the registry and minorities were both more likely to report religious objections to donation and to mistrust the fairness of HPC allocation. The pattern of findings was similar in UK and US samples. Registries should maintain contact with potential donors after recruitment, aiming to educate members about the donation procedure and to address potential misconceptions associated with religious beliefs and HPC allocation.

Canonical and noncanonical inflammasomes in intestinal epithelial cells.

Inflammasomes are cytosolic, multimeric protein complexes capable of activating pro-inflammatory cytokines such as IL-1ß and IL-18, which play a key role in host defence. Inflammasome components are highly expressed in the intestinal epithelium. In recent years, studies have begun to demonstrate that epithelial-intrinsic inflammasomes play a critical role in regulating epithelial homeostasis, both by defending the epithelium from pathogenic insult and through the regulation of the mucosal environment. However, the majority of research regarding inflammasome activation has focused on professional immune cells, such as macrophages. Here, we present an overview of the current understanding of inflammasome function in epithelial cells and at mucosal surfaces and, in particular, in the intestine.

Isolation and In Vitro Culture of Human Gut Progenitor Cells.

The gastrointestinal epithelium is a highly regenerative organ, where each cell is replaced in a cycle of 4-6 days, depending on the mammalian species. This highly proliferative state is driven by gastrointestinal stem and progenitor cells, located at the base of crypts. These cells give rise to at least six types of differentiated epithelial cells, each with a distinct function in maintaining homeostasis between the intestinal interface and the luminal environment. The isolation and culture of these cells from mammalian gastrointestinal tissue is a novel technique, which allows for the generation and maintenance of an in vitro culture system for adult epithelial cells. There are two predominant methods for isolation and propagation of gastrointestinal epithelial cells, the first is the organoid system developed in 2009, and the second is a later version known as the L-WRN spheroid system. In this chapter, we describe the method to isolate and culture human gastrointestinal stem and progenitor cells and culture them as three-dimensional spheroids using L-WRN cell conditioned media.

Platelet activating factor receptor regulates colitis-induced pulmonary inflammation through the NLRP3 inflammasome.

Extra-intestinal manifestations (EIM) are common in inflammatory bowel disease (IBD). One such EIM is sub-clinical pulmonary inflammation, which occurs in up to 50% of IBD patients. In animal models of colitis, pulmonary inflammation is driven by neutrophilic infiltrations, primarily in response to the systemic bacteraemia and increased bacterial load in the lungs. Platelet activating factor receptor (PAFR) plays a critical role in regulating pulmonary responses to infection in conditions, such as chronic obstructive pulmonary disease and asthma. We investigated the role of PAFR in pulmonary EIMs of IBD, using dextran sulfate sodium (DSS) and anti-CD40 murine models of colitis. Both models induced neutrophilic inflammation, with increased TNF and IL-1ß levels, bacterial load and PAFR protein expression in mouse lungs. Antagonism of PAFR decreased lung neutrophilia, TNF, and IL-1ß in an NLRP3 inflammasome-dependent manner. Lipopolysaccharide from phosphorylcholine (ChoP)-positive bacteria induced NLRP3 and caspase-1 proteins in human alveolar epithelial cells, however antagonism of PAFR prevented NLRP3 activation by ChoP. Amoxicillin reduced bacterial populations in the lungs and reduced NLRP3 inflammasome protein levels, but did not reduce PAFR. These data suggest a role for PAFR in microbial pattern recognition and NLRP3 inflammasome signaling in the lung.

Transplant center practices for psychosocial assessment and management of pediatric hematopoietic stem cell donors.

Understanding the potential emotional and psychological risks of pediatric sibling HSC donation is an area of research that remains in its infancy. A cross-sectional survey was distributed electronically to directors at all CIBMTR and EBMT centers to describe current transplant center practices for obtaining assent, preparation for the physical/emotional experiences of donation, and monitoring the post-donation well-being of pediatric donors (<18 years of age). Respondents were 45/91 (49%) and 66/144 (46%) of CIBMTR and EBMT centers, respectively. Although 78% of centers reported having a mechanism in place to ensure donor free assent, centers also reported only limited assessment of psychosocial suitability to manage the emotional risks of donation. More than half of centers reported no psychosocial follow-up assessment post-donation. Few centers have policies in place to address donor psychological needs. Future investigations should include medical and psychosocial outcomes following full integration of comprehensive psychosocial screening and surveillance of pediatric donors.

Motives for joining an unrelated HSC donor registry: description, categorization, and association with donor availability.

A challenge for registries of unrelated hematopoietic stem cell (HSC) donors is to recruit and retain sufficient donors for patients needing transplants. Many registered HSC donors opt-out when called as a potential match for a patient. Anecdotal evidence suggests that motives for joining a registry may be linked to the donation decision. The primary goals of this investigation were to describe the range of motives for joining a registry and to examine donor availability by motive type. A diverse sample of 357 potential HSC donors from the Anthony Nolan registry in the UK was asked about their motives for joining and their decision to continue or opt-out after matching a patient was recorded. Motives for joining (N = 557) were first coded and categorized into 17 specific motive types and then arranged along a spectrum from internal to external. Internal values-based motives were most prevalent and availability was highest among potential donors expressing these motives (92%) and lowest among those expressing external motives (pressure, incentives; 0%). Although further research is needed to confirm these findings, they suggest that registries should assess donors' motives for joining at recruitment in order to conduct follow-up to ensure commitment among those potential members expressing external motives.

A Rodent Model of Anxiety: The Effect of Perinatal Immune Challenges on Gastrointestinal Inflammation and Integrity.

OBJECTIVES: Gastrointestinal (GI) inflammation and GI integrity deficits are common comorbidities of neuropsychiatric disorders. Ongoing research suggests that these aberrations may be contributing to heightened immune signals that have the potential to disrupt neuronal homeostasis and exacerbate behavioural deficits. The current study aimed to determine whether the well-characterized animal model of neuropsychopathology, the maternal immune activation (MIA) model, produced GI inflammation and integrity disruptions in association with anxiety-like behaviour. METHODS: Pregnant Wistar rats were exposed to the viral mimetic polyriboinosinic:polyribocytidilic acid (polyI:C) on gestational days (GD) 10 and 19. Evidence of ANS activation, GI inflammation, and GI barrier integrity was assessed in both neonatal (postnatal day, P7) and adult (P84) offspring. Anxiety-like behaviour was assessed at P100. RESULTS: Neonatal MIA offspring exhibited an altered intestinal inflammatory profile and evidence of an increase in lymphoid aggregates. MIA neonates also displayed disruptions to GI barrier tight junction protein mRNA. In addition, adult MIA offspring exhibited an increase in anxiety-like behaviours. CONCLUSION: These results indicate that the MIA rat model, which is well documented to produce behavioural, neurochemical, and neuroanatomical abnormalities, also produces GI inflammation and integrity disruptions. We suggest that this model may be a useful tool to elucidate biological pathways associated with neuropsychiatric disorders.