Transcription factor Tcf21 modulates urinary bladder size and differentiation.

Urinary bladder organogenesis requires coordinated cell growth, specification, and patterning of both mesenchymal and epithelial compartments. Tcf21, a gene that encodes a helix-loop-helix transcription factor, is specifically expressed in the mesenchyme of the bladder during development. Here we show that Tcf21 is required for normal development of the bladder. We found that the bladders of mice lacking Tcf21 were notably hypoplastic and that the Tcf21 mutant mesenchyme showed increased apoptosis. There was also a marked delay in the formation of visceral smooth muscle, accompanied by a defect in myocardin (Myocd) expression. Interestingly, there was also a marked delay in the formation of the basal cell layer of the urothelium, distinguished by diminished expression of Krt5 and Krt14. Our findings suggest that Tcf21 regulates the survival and differentiation of mesenchyme cell-autonomously and the maturation of the adjacent urothelium non-cell-autonomously during bladder development.

Comorbidities increase COVID-19 hospitalization in young people with type 1 diabetes.

OBJECTIVES: We evaluated COVID-19 outcomes in children and young adults with type 1 diabetes (T1D) to determine if those with comorbidities are more likely to experience severe COVID-19 compared to those without. RESEARCH DESIGN AND METHODS: This cross-sectional study included questionnaire data on patients <25 years of age with established T1D and laboratory-confirmed COVID-19 from 52 sites across the US between April 2020 and October 2021. We examined patient factors and COVID-19 outcomes between those with and without comorbidities. Multivariate logistic regression analysis examined the odds of hospitalization among groups, adjusting for age, HbA1c, race and ethnicity, insurance type and duration of diabetes. RESULTS: Six hundred fifty-one individuals with T1D and COVID-19 were analyzed with mean age 15.8 (SD 4.1) years. At least one comorbidity was present in 31%, and more than one in 10%. Obesity and asthma were the most frequently reported comorbidities, present in 19% and 17%, respectively. Hospitalization occurred in 17% of patients and 52% of hospitalized patients required ICU level care. Patients with at least one comorbidity were almost twice as likely to be hospitalized with COVID-19 than patients with no comorbidities (Odds ratio 2.0, 95% CI: 1.3-3.1). This relationship persisted after adjusting for age, HbA1c, race and ethnicity (minority vs nonminority), insurance type (public vs. private), and duration of diabetes. CONCLUSIONS: Our findings show that comorbidities increase the risk for hospitalization with COVID-19 in children and young adults highlighting the need for tailored COVID-19 prevention and treatment strategies in T1D.

Expression and characterization of αvß5 integrin on intestinal macrophages.

Macrophages play a crucial role in maintaining homeostasis in the intestine, but the underlying mechanisms have not yet been elucidated fully. Here, we show for the first time that mature intestinal macrophages in mouse intestine express high levels of αvß5 integrin, which acts as a receptor for the uptake of apoptotic cells and can activate molecules involved in several aspects of tissue homeostasis such as angiogenesis and remodeling of the ECM. αvß5 is not expressed by other immune cells in the intestine, is already present on intestinal macrophages soon after birth, and its expression is not dependent on the microbiota. In adults, αvß5 is induced during the differentiation of monocytes in response to the local environment and it confers intestinal macrophages with the ability to promote engulfment of apoptotic cells via engagement of the bridging molecule milk fat globule EGF-like molecule 8. In the absence of αvß5, there are fewer monocytes in the mucosa and mature intestinal macrophages have decreased expression of metalloproteases and IL 10. Mice lacking αvß5 on haematopoietic cells show increased susceptibility to chemical colitis and we conclude that αvß5 contributes to the tissue repair by regulating the homeostatic properties of intestinal macrophages.

Screening for Nephropathy in Pediatric Type 2 Diabetes: Quality Improvement to Increase Nephropathy Screening.

Background: Screening for early detection of microalbuminuria signaling kidney disease should begin as early as the time of diagnosis of youth-onset type 2 diabetes. This quality improvement initiative aimed to standardize urine nephropathy screening in pediatric patients with type 2 diabetes at a tertiary academic medical center and increase a baseline screening rate of 56%-75% over 6 months (September 2022-February 2023) and sustain that increase for 6 months (March through August 2023). Methods: A multi-disciplinary team used quality improvement methods and iterative Plan-Do-Study-Act cycles. Targeted interventions included previsit planning workflow, education, and a new-onset triage protocol. The team collected data at baseline and prospectively by reviewing electronic medical records. The primary outcome measure was pediatric type 2 diabetes clinic visits in diabetes clinic with urine nephropathy screening before or on the visit date. Results: A total of 121 youth were scheduled for T2D clinic visits between September 2021 and August 2023. The mean age was 14.5 years, and 60% were women, 40% were non-Hispanic Black, 28% were Hispanic/Latino, and 15% reported Spanish as their preferred language. Following the interventions of this project, urine nephropathy screening increased from 56% to 75%, and this change was sustained for 6 months. Conclusions: Interventions focused on efficient recognition of the population needing screening, coordinated internal processes around screening, a shared understanding between all stakeholders, and practical support in the healthcare system increased urine nephropathy screening with sustained improvement.

Guanylate cyclase C limits systemic dissemination of a murine enteric pathogen.

BACKGROUND: Guanylate Cyclase C (GC-C) is an apically-oriented transmembrane receptor that is expressed on epithelial cells of the intestine. Activation of GC-C by the endogenous ligands guanylin or uroguanylin elevates intracellular cGMP and is implicated in intestinal ion secretion, cell proliferation, apoptosis, intestinal barrier function, as well as the susceptibility of the intestine to inflammation. Our aim was to determine if GC-C is required for host defense during infection by the murine enteric pathogen Citrobacter rodentium of the family Enterobacteriacea. METHODS: GC-C+/+ control mice or those having GC-C genetically ablated (GC-C-/-) were administered C. rodentium by orogastric gavage and analyzed at multiple time points up to post-infection day 20. Commensal bacteria were characterized in uninfected GC-C+/+ and GC-C-/- mice using 16S rRNA PCR analysis. RESULTS: GC-C-/- mice had an increase in C. rodentium bacterial load in stool relative to GC-C+/+. C. rodentium infection strongly decreased guanylin expression in GC-C+/+ mice and, to an even greater degree, in GC-C-/- animals. Fluorescent tracer studies indicated that mice lacking GC-C, unlike GC-C+/+ animals, had a substantial loss of intestinal barrier function early in the course of infection. Epithelial cell apoptosis was significantly increased in GC-C-/- mice following 10 days of infection and this was associated with increased frequency and numbers of C. rodentium translocation out of the intestine. Infection led to significant liver histopathology in GC-C-/- mice as well as lymphocyte infiltration and elevated cytokine and chemokine expression. Relative to naïve GC-C+/+ mice, the commensal microflora load in uninfected GC-C-/- mice was decreased and bacterial composition was imbalanced and included outgrowth of the Enterobacteriacea family. CONCLUSIONS: This work demonstrates the novel finding that GC-C signaling is an essential component of host defense during murine enteric infection by reducing bacterial load and preventing systemic dissemination of attaching/effacing-lesion forming bacterial pathogens such as C. rodentium.

Murine guanylate cyclase C regulates colonic injury and inflammation.

Guanylate cyclase C (GUCY2C or GC-C) and its ligands, guanylin (GUCA2A or Gn) and uroguanylin (GUCA2B or Ugn), are expressed in intestinal epithelial cells and regulate ion secretion, intestinal barrier function, and epithelial monolayer homeostasis via cGMP-dependent signaling pathways. The aim of this study was to determine whether GC-C and its ligands direct the course of intestinal inflammation. In this article, we show that dextran sodium sulfate (DSS)-induced clinical disease and histological damage to the colonic mucosa were significantly less severe in GC-C(-/-) mice and moderately reduced in Gn(-/-) animals. Relative to wild-type controls, GC-C(-/-) and Gn(-/-) mice had reduced apoptosis and increased proliferation of intestinal epithelial cells during DSS colitis. Basal and DSS-induced production of resistin-like molecule ß (RELMß) was substantially diminished in GC-C(-/-) mice. RELMß is thought to stimulate cytokine production in macrophages in this disease model and, consistent with this, TNF-α and IFN-γ production was minimal in GC-C(-/-) animals. RELMß and cytokine levels were similar to wild-type in Gn(-/-) mice, however. Colonic instillation of recombinant RELMß by enema into GC-C(-/-) mice restores sensitivity to DSS-mediated mucosal injury. These findings demonstrate a novel role for GC-C signaling in facilitating mucosal wounding and inflammation, and further suggest that this may be mediated, in part, through control of RELMß production.

Gastrointestinal infection as a trigger for inflammatory bowel disease.

PURPOSE OF REVIEW: There is accumulating evidence on the importance of microbes in the development and maintenance of both the intestinal and immune systems. This review focuses on the current findings on the role of gastrointestinal pathogens in the cause of chronic inflammatory bowel disease. RECENT FINDINGS: A number of intestinal pathogens including Mycobacterium avium subspecies paratuberculosis, adherent-invasive Escherichia coli, and Campylobacter species are associated at fairly high prevalence with Crohn's disease, while two recent studies found a low prevalence for cytomegalovirus. In a prospective study, M. avium subspecies paratuberculosis detection in early Crohn's disease was low and comparable to controls, while much higher in an established inflammatory bowel disease cohort. In the pediatric setting, a high prevalence of Clostridium difficile was seen in both active and inactive Crohn's disease and ulcerative colitis patients. Some studies have speculated that Salmonella or Campylobacter infection may increase the risk of inflammatory bowel disease on long-term follow-up, but detection bias was found to obscure the risk. Recent studies in mouse models have demonstrated that a combination of factors, including viral pathogens, genetic susceptibility, and commensal microflora, can lead to intestinal pathology. SUMMARY: No evidence for causation of inflammatory bowel disease by a single agent has been found, whereas a number of microbes have been strongly associated with the presence of disease. The majority of recent studies support a role for the ability of intestinal pathogens to promote chronic inflammation in individuals with genetic susceptibility and/or other environmental factors which remain to be identified. These factors may include subsets of commensal microflora.

Computerized decision support system improves fluid resuscitation following severe burns: an original study.

OBJECTIVE: Several formulas have been developed to guide resuscitation in severely burned patients during the initial 48 hrs after injury. These approaches require manual titration of fluid that may result in human error during this process and lead to suboptimal outcomes. The goal of this study was to analyze the efficacy of a computerized open-loop decision support system for burn resuscitation compared to historical controls. DESIGN: Fluid infusion rates and urinary output from 39 severely burned patients with >20% total body surface area burns were recorded upon admission (Model group). A fluid-response model based on these data was developed and incorporated into a computerized open-loop algorithm and computer decision support system. The computer decision support system was used to resuscitate 32 subsequent patients with severe burns (computer decision support system group) and compared with the Model group. SETTING: Burn intensive care unit of a metropolitan Level 1 Trauma center. PATIENTS: Acute burn patients with >20% total body surface area requiring active fluid resuscitation during the initial 24 to 48 hours after burn. MEASUREMENTS AND MAIN RESULTS: We found no significant difference between the Model and computer decision support system groups in age, total body surface area, or injury mechanism. Total crystalloid volume during the first 48 hrs post burn, total crystalloid intensive care unit volume, and initial 24-hr crystalloid intensive care unit volume were all lower in the computer decision support system group. Infused volume per kilogram body weight (mL/kg) and per percentage burn (mL/kg/total body surface area) were also lower for the computer decision support system group. The number of patients who met hourly urinary output goals was higher in the computer decision support system group. CONCLUSIONS: Implementation of a computer decision support system for burn resuscitation in the intensive care unit resulted in improved fluid management of severely burned patients. All measures of crystalloid fluid volume were reduced while patients were maintained within urinary output targets a higher percentage of the time. The addition of computer decision support system technology improved patient care.

Clostridium difficile infection and treatment in the pediatric inflammatory bowel disease population.

OBJECTIVE: Recent changes in the epidemiology of Clostridium difficile infection include an increase in the incidence of C difficile-associated disease (CDAD) and the identification of patients with inflammatory bowel disease (IBD) as a group at risk. In addition, the effectiveness of antimicrobial therapies has been questioned. Our aim was to estimate the incidence of CDAD in a pediatric IBD population and review treatment efficacy. PATIENTS AND METHODS: We identified patients ages 18 years or younger from our center's IBD database who tested positive for C difficile toxin A and/or B between August 1, 2007 and December 31, 2008. Demographic information and treatment details were recorded. Chi-square and Fisher exact tests were used to compare categorical variables and the Student t test was used for continuous variables. RESULTS: From 372 pediatric patients with IBD, we identified 29 patients who experienced a total of 40 cases of CDAD. The annualized incidence rate of CDAD was 7.2%. Initial treatment was successful in 17 cases (43%). Eventual success was documented with metronidazole in 15 cases (41%), with vancomycin in 16 cases (43%), and with other agents or a combination of agents in 6 cases (16%). Age, sex, and IBD type were not associated with initial treatment outcome or recurrence. The choice of initial antimicrobial treatment was not associated with treatment outcome. The type of IBD therapy medication was not associated with the likelihood of CDAD recurrence, although the use of anti-inflammatory therapy was positively associated with initial antimicrobial treatment success. CONCLUSIONS: CDAD occurred frequently in our cohort of pediatric patients with IBD. Antimicrobial treatment success was achieved equally with either metronidazole or vancomycin. Initial treatment failed more than half of the time, regardless of medication choice. Apparent lack of antimicrobial efficacy in resolving symptoms may reflect resistant C difficile infection or increased IBD severity in a subset of patients who are C difficile carriers. Awareness of the potential for a high incidence of CDAD and frequent failure rate of initial therapy is important in the management of children with IBD.

Anemia causes hypoglycemia in intensive care unit patients due to error in single-channel glucometers: methods of reducing patient risk.

OBJECTIVE: Intensive insulin therapy in the critically ill reduces mortality but carries the risk of increased hypoglycemia. Point-of-care blood glucose analysis is standard; however, anemia causes falsely high values and potentially masks hypoglycemia. Permissive anemia is practiced routinely in most intensive care units. We hypothesized that point-of-care glucometer error due to anemia is prevalent, can be corrected mathematically, and correction uncovers occult hypoglycemia during intensive insulin therapy. DESIGN: The study has both retrospective and prospective phases. We reviewed data to verify the presence of systematic error, determine the source of error, and establish the prevalence of anemia. We confirmed our findings by reproducing the error in an in vitro model. Prospective data were used to develop a correction formula validated by the Monte Carlo method. Correction was implemented in a burn intensive care unit and results were evaluated after 9 mos. SETTING: Burn and trauma intensive care units at a single research institution. PATIENTS/SUBJECTS: Samples for in vitro studies were taken from healthy volunteers. Samples for formula development were from critically ill patients who received intensive insulin therapy. INTERVENTIONS: Insulin doses were calculated based on predicted serum glucose values from corrected point-of-care glucometer measurements. MEASUREMENTS AND MAIN RESULTS: Time-matched point-of-care glucose, laboratory glucose, and hematocrit values. We previously found that anemia (hematocrit <34%) produces systematic error in glucometer measurements. The error was correctible with a mathematical formula developed and validated, using prospectively collected data. Error of uncorrected point-of-care glucose ranged from 19% to 29% (p < .001), improving to < or = 5% after mathematical correction of prospective data. Comparison of data pairs before and after correction formula implementation demonstrated a 78% decrease in the prevalence of hypoglycemia in critically ill and anemic patients treated with insulin and tight glucose control (p < .001). CONCLUSIONS: A mathematical formula that corrects erroneous point-of-care glucose values due to anemia in intensive care unit patients reduces the prevalence of hypoglycemia during intensive insulin therapy.

Lack of guanylate cyclase C results in increased mortality in mice following liver injury.

BACKGROUND: Guanylate Cyclase C (GC-C) expression in the intestine plays a role in the regulation of fluid and ion transport, as well as epithelial cell apoptosis and proliferation. In the adult rat liver, GC-C expression is increased in response to injury. We hypothesized that GC-C is required for repair/recovery from liver injury. METHODS: We subjected wild type (WT) and GC-C deficient mice to acute liver injury with a single injection of the hepatotoxin carbon tetrachloride. Changes in the level of expression of GC-C and its ligands uroguanylin and guanylin were quantified by real-time PCR. Liver morphology, and hepatocyte necrosis, apoptosis and proliferation, were examined at 1-3 days post-injury in mice on a mixed genetic background. Survival was followed for 14 days after carbon tetrachloride injection in wild type and GC-C deficient mice on both a mixed genetic background and on an inbred C57BL6/J background. RESULTS: GC-C deficient mice on the mixed genetic background nearly all died (median survival of 5 days) following carbon tetrachloride injection while WT littermates experienced only 35% mortality. Elevated levels of TUNEL-positive hepatocyte death on post-injury day 1, increased apoptosis on day 2, and increased areas of centrilobular necrosis on days 2 and 3, were evident in livers from GC-C null mice compared to WT. Collectively these data suggest increased hepatocyte death in the GC-C null mice in the early time period after injury. This corresponds temporally with increased expression of GC-C and its ligands guanylin and uroguanylin in post-injury WT mouse liver. The hepatocyte proliferative response to injury was the same in both genotypes. In contrast, there was no difference in survival between GC-C null and WT mice on the inbred C57BL/6 J background in response to acute liver injury. CONCLUSIONS: Signalling via GC-C promotes hepatocyte survival in vivo and is required for effective recovery from acute toxic injury to the liver in a strain-specific manner.

Factors associated with health psychology use in pediatric type 1 diabetes.

AIMS: Identify the prevalence of health psychology use in children with type 1 diabetes (T1D) and evaluate how individual and contextual characteristics are associated with use. METHODS: Children ages 8-16 years with T1D and their parents were recruited from two tertiary diabetes clinics. Cross-sectional data included parent and adolescent surveys and hemoglobin A1c. Parents self-reported the child's use of health psychology in the last year along with individual factors (e.g., predisposing factors including demographics, enabling factors including health insurance type, evaluated need including mental health diagnoses and perceived need including self-management barriers). Association of health psychology use with individual (e.g., demographics, enabling factors, evaluated and perceived need) and contextual (e.g., clinical site) characteristics was evaluated using logistic regression. RESULTS: Of 363 eligible participants, 267 (74%) participated. Health psychology use was reported by only 8.2% (n = 22) of participants and was significantly associated with evaluated need factor of mental health diagnosis (OR 5.8; p < 0.001) and perceived need factor of parent-reported self-management barriers. Use was not associated with other individual or contextual factors. CONCLUSIONS: Though infrequent, health psychology use was positively associated with mental health diagnoses and self-management barriers.

Mouse knockout of guanylyl cyclase C: Recognition memory deficits in the absence of activity changes.

Guanylyl cyclase C (GC-C) is found in brain regions where dopamine is expressed. We characterized a mouse in which GC-C was knocked out (KO) that was reported to be a model of attention deficit hyperactivity disorder (ADHD). We re-examined this model and controlled for litter effects, used 16 to 23 mice per genotype per sex and assessed an array of behavioral and neurochemical outcomes. GC-C KO mice showed no phenotypic differences from wild-type mice on most behavioral tests, or on striatal or hippocampal monoamines, and notably no evidence of an ADHD-like phenotype. KO mice were impaired on novel object recognition, had decreased tactile startle but not acoustic startle, and females had increased latency on cued training trials in the Morris water maze, but not hidden platform spatial learning trials. Open-field activity showed small differences in females but not males. The data indicate that the GC-C KO mouse with proper controls and sample sizes has a moderate cognitive and startle phenotype but has no ADHD-like phenotype.

Establishing the incidence and timing of hypoglycemia at a residential diabetes camp.

AIMS: To establish the incidence and timing of hypoglycemia at a week-long residential diabetes camp for children. We hypothesized that hypoglycemia would occur more frequently during the first two days of camp and following evening all-camp games. METHODS: 225 children (mean age 12.0 ±â€¯2.3 years, 56% female, mean hemoglobin A1c 8.4% [71.6 mmol/mol]) had blood glucose (BG) levels obtained before meals, at bedtime, and as needed to detect hypoglycemia. Insulin adjustments were made by medical staff according to camp protocol and at the discretion of medical staff during camper check-in. RESULTS: Mild hypoglycemia (BG 50-69 mg/dL [3.9 mmol/L]) occurred ≥ 1 time in 90% of campers while 43% had ≥ 1 episode of BG < 50 mg/dL (2.8 mmol/L). No episodes of hypoglycemia requiring glucagon occurred. More campers experienced ≥ 1 overnight hypoglycemia event during the first 48 hours of camp compared to later in the week (p = 0.01). Evening all-camp games did not impact hypoglycemia rates overnight. CONCLUSIONS: Nocturnal hypoglycemia occurred more frequently during the first two nights, establishing this period as high risk and supporting implementation of a standard protocol to lower insulin doses. Rates of hypoglycemia were unaffected by all-camp games, indicating current practices are effective at minimizing hypoglycemia.

Error rates resulting from anemia can be corrected in multiple commonly used point-of-care glucometers.

BACKGROUND: A point-of-care (POC) glucometer (G1) used for critical care at our institution is inaccurate in the presence of low hematocrit (HCT) values. The purpose of this study was to analyze error rates of three additional POC glucometer brands and determine mathematical correction formulas for each. METHODS: Blood samples (n = 196) from a cohort of surgical, trauma, medical, cardiothoracic, and burn intensive care unit patients were tested on three commonly used POC glucometer brands (G2-G4). Results were compared with reference laboratory values, and correction compared with the validated formula for G1. A mathematical formula specific to each glucometer type was derived from glucose measurements, associated HCT values, and the degree of difference relative to laboratory results. RESULTS: POC glucometer results were consistently elevated compared with reference laboratory values. Glucometer error rates for HCT

Prevalence of mutations in hepatitis C virus core protein associated with alteration of NF-kappaB activation.

The hepatitis C virus (HCV) core protein is a key structural element of the virion but also affects a number of cellular pathways, including nuclear factor kappaB (NF-kappaB) signaling. NF-kappaB is a transcription factor that regulates both anti-apoptotic and pro-inflammatory genes and its activation may contribute to HCV-mediated pathogenesis. Amino acid sequence divergence in core is seen at the genotype level as well as within patient isolates. Recent work has implicated amino acids 9-11 of core in the modulation of NF-kappaB activation. We report that the sequence RKT is highly conserved (93%) at this position across all HCV genotypes, based on sequences collected in the Los Alamos HCV database. Of the 13 types of variants present in the database, the two most prevalent substitutions are RQT and RKP. We further show that core encoding RKP fails to activate NF-kappaB signaling in vitro while NF-kappaB activation by core encoding RQT does not differ from control RKT core. The effect of RKP core is specific to NF-kappaB signaling as activator protein 1 (AP-1) activity is not altered. Further studies are needed to assess potential associations between specific amino acid substitutions at positions 9-11 and liver disease progression and/or response to treatment in individual patients.

Chronic social defeat, but not restraint stress, alters bladder function in mice.

BACKGROUND: Voiding disorders in humans, particularly in children are associated with increased incidence of behavioral issues as well as past history of childhood abuse. We hypothesized that creating stress in mice, utilizing either a chronic social defeat model (SD) or restraint stress in shallow water model (RSSW) would engender changes in bladder function, morphology, and behavior, thereby enabling us to study the resultant voiding dysfunction. METHODS: For SD stress (14 days), C57BL/6 male mice were exposed daily to a larger aggressive CD-1 male for 10 min, followed by sensory exposure in a barrier cage for 24h. Control mice were similarly housed with no exposure. For RSSW (21 days), C57BL/6 mice were put in a perforated conical tube with feet immersed in water daily for 4h, then returned to single housing cages. Control mice were also in single housing. After the stress period, voiding patterns were obtained on filter paper, followed by behavioral tests. At necropsy, blood was taken for corticosterone analysis, and bladder and body weights measured. Bladder cryosections were stained with hematoxylin and eosin (H&E) for morphological assessment. Sequential sections were immunostained with antibodies to Ki-67 as a proliferation marker, CD31 (endothelial cell marker), and uroplakin-II. ImageJ software was used to measure bladder wall thickness on blinded H&E photomicrographs as well as quantitate CD31 staining. Both Ki-67-positive and -negative nuclei were counted with Imaris software to obtain a proliferation index. RESULTS: Only SD mice had a single large void pattern. Bladder-to-body weight ratios increased in SD mice (p≤0.02) but not in RSSW mice. Plasma corticosterone levels were elevated in all stressed mice. SD mice exhibited lower levels of locomotor activity compared with controls; RSSW mice were hyperactive. In SD mice, bladder wall thickness was increased (p ≤ 0.003) but no change was seen in Ki-67 proliferation index, consistent with hypertrophy. No difference with control mice was seen in vascularity as visualized by CD31 staining. Uniform uroplakin-II staining lined the urothelium of both SD and control mice. CONCLUSIONS: Mice exposed to repeated SD (14 days) respond with altered voiding indicative of urine retention, and exhibit bladder wall changes consistent with hypertrophy while the urothelial barrier is maintained. These changes were not observed with repeated RSSW. SD, in contrast to RSSW, provides a model of psychological stress to further study the interplay of behavior and bladder dysfunction, enabling an improved understanding of voiding dysfunction, and the ability to create innovative and more effective management pathways for children who present with voiding dysfunction.

E. coli heat-stable enterotoxin and guanylyl cyclase C: new functions and unsuspected actions.

Some E. coli cause diarrhea by elaborating heat-labile and heat-stable (ST) enterotoxins which stimulate intestinal secretion. E. coli ST's are small peptides which bind to intestinal luminal epithelial cell receptors. The ST receptor, one of a family of receptor-cyclases called guanylyl cyclase C (GC-C), is a membrane spanning protein containing an extracellular binding domain and intracellular protein kinase and catalytic domains. The intestine synthesizes and secretes homologous peptides, guanylin and uroguanylin. The kidney also synthesizes uroguanylin. ST, guanylin or uroguanylin binding to GC-C results in increased cGMP, phosphorylation of the CFTR Cl- channel and secretion. Proguanylin and prouroguanylin circulate in blood and bind to receptors in intestine, kidney, liver, brain etc. In the kidney, they stimulate the excretion of Na+ and K+. Study of GC-C "knock-out" mice reveal that GC-C is important to intestinal salt and water secretion, duodenal bicarbonate secretion, recovery from CCl4-induced liver injury, and to intestinal polyp formation in Min mice lacking GC-C.

Guanylate cyclase C deficiency causes severe inflammation in a murine model of spontaneous colitis.

BACKGROUND: Guanylate Cyclase C (GC-C; Gucy2c) is a transmembrane receptor expressed in intestinal epithelial cells. Activation of GC-C by its secreted ligand guanylin stimulates intestinal fluid secretion. Familial mutations in GC-C cause chronic diarrheal disease or constipation and are associated with intestinal inflammation and infection. Here, we investigated the impact of GC-C activity on mucosal immune responses. METHODS: We utilized intraperitoneal injection of lipopolysaccharide to elicit a systemic cytokine challenge and then measured pro-inflammatory gene expression in colonic mucosa. GC-C(+/+) and GC-C(-/-) mice were bred with interleukin (IL)-10 deficient animals and colonic inflammation were assessed. Immune cell influx and cytokine/chemokine expression was measured in the colon of wildtype, IL-10(-/-), GC-C(+/+)IL-10(-/-) and GC-C(-/-)IL-10(-/-) mice. GC-C and guanylin production were examined in the colon of these animals and in a cytokine-treated colon epithelial cell line. RESULTS: Relative to GC-C(+/+) animals, intraperitoneal lipopolysaccharide injection into GC-C(-/-) mice increased proinflammatory gene expression in both whole colon tissue and in partially purified colonocyte isolations. Spontaneous colitis in GC-C(-/-)IL-10(-/-) animals was significantly more severe relative to GC-C(+/+)IL-10(-/-) mice. Unlike GC-C(+/+)IL-10(-/-) controls, colon pathology in GC-C(-/-)IL-10(-/-) animals was apparent at an early age and was characterized by severely altered mucosal architecture, crypt abscesses, and hyperplastic subepithelial lesions. F4/80 and myeloperoxidase positive cells as well as proinflammatory gene expression were elevated in GC-C(-/-)IL-10(-/-) mucosa relative to control animals. Guanylin was diminished early in colitis in vivo and tumor necrosis factor α suppressed guanylin mRNA and protein in intestinal goblet cell-like HT29-18-N2 cells. CONCLUSIONS: The GC-C signaling pathway blunts colonic mucosal inflammation that is initiated by systemic cytokine burst or loss of mucosal immune cell immunosuppression. These data as well as the apparent intestinal inflammation in human GC-C mutant kindred underscore the importance of GC-C in regulating the response to injury and inflammation within the gut.