Lymphatic muscle cells are the innate pacemaker cells regulating mouse lymphatic collecting vessel contractions.

Collecting lymphatic vessels (cLVs) exhibit spontaneous contractions with a pressure-dependent frequency, but the identity of the lymphatic pacemaker cell is still debated. By analogy to pacemakers in the GI and lower urinary tracts, proposed cLV pacemaker cells include interstitial cells of Cajal like cells (ICLC), pericytes, as well as the lymphatic muscle (LMCs) cells themselves. Here we tested the extent to which these cell types are invested into the mouse cLV wall and if any cell type exhibited morphological and functional processes characteristic of pacemaker cells: a contiguous network; spontaneous Ca2+ transients; and depolarization-induced propagated contractions. We employed inducible Cre (iCre) mouse models routinely used to target these specific cell populations including: c-kitCreERT2 to target ICLC; PdgfrßCreERT2 to target pericytes; PdgfrαCreER™ to target CD34+ adventitial fibroblast-like cells or ICLC; and Myh11CreERT2 to target LMCs. These specific inducible Cre lines were crossed to the fluorescent reporter ROSA26mT/mG, the genetically encoded Ca2+ sensor GCaMP6f, and the light-activated cation channel rhodopsin2 (ChR2). c-KitCreERT2 labeled both a sparse population of LECs and round adventitial cells that responded to the mast cell activator compound 48-80. PdgfrßCreERT2 drove recombination in both adventitial cells and LMCs, limiting its power to discriminate a pericyte specific population. PdgfrαCreER™ labeled a large population of interconnected, oak leaf-shaped cells primarily along the adventitial surface of the vessel. Titrated induction of the smooth muscle-specific Myh11CreERT2 revealed a LMC population with heterogeneous morphology. Only LMCs consistently, but heterogeneously, displayed spontaneous Ca2+ events during the diastolic period of the contraction cycle, and whose frequency was modulated in a pressure-dependent manner. Optogenetic depolarization through the expression of ChR2 by Myh11CreERT2, but not PdgfrαCreER™ or c-KitCreERT2, resulted in a propagated contraction. These findings support the conclusion that LMCs, or a subset of LMCs, are responsible for mouse cLV pacemaking.

Excitatory cholinergic responses in mouse primary bronchial smooth muscle require both Ca2+ entry via l-type Ca2+ channels and store operated Ca2+ entry via Orai channels.

Malfunctions in airway smooth muscle Ca2+-signalling leads to airway hyperresponsiveness in asthma and chronic obstructive pulmonary disease. Ca2+-release from intracellular stores is important in mediating agonist-induced contractions, but the role of influx via l-type Ca2+ channels is controversial. We re-examined roles of the sarcoplasmic reticulum Ca2+ store, refilling of this store via store-operated Ca2+ entry (SOCE) and l-type Ca2+ channel pathways on carbachol (CCh, 0.1-10 µM)-induced contractions of mouse bronchial rings and intracellular Ca2+ signals of mouse bronchial myocytes. In tension experiments, the ryanodine receptor (RyR) blocker dantrolene (100 µM) reduced CCh-responses at all concentrations, with greater effects on sustained rather than initial components of contraction. 2-Aminoethoxydiphenyl borate (2-APB, 100 µM), in the presence of dantrolene, abolished CCh-responses, suggesting the sarcoplasmic reticulum Ca2+ store is essential for contraction. The SOCE blocker GSK-7975A (10 µM) reduced CCh-contractions, with greater effects at higher (e.g. 3 and 10 µM) CCh concentrations. Nifedipine (1 µM), abolished remaining contractions in GSK-7975A (10 µM). A similar pattern was observed on intracellular Ca2+-responses to 0.3 µM CCh, where GSK-7975A (10 µM) substantially reduced Ca2+ transients induced by CCh, and nifedipine (1 µM) abolished remaining responses. When nifedipine (1 µM) was applied alone it had less effect, reducing tension responses at all CCh concentrations by 25% - 50%, with greater effects at lower (e.g. 0.1 and 0.3 µM) CCh concentrations. When nifedipine (1 µM) was examined on the intracellular Ca2+-response to 0.3 µM CCh, it only modestly reduced Ca2+ signals, while GSK-7975A (10 µM) abolished remaining responses. In conclusion, Ca2+-influx from both SOCE and l-type Ca2+ channels contribute to excitatory cholinergic responses in mouse bronchi. The contribution of l-type Ca2+ channels was especially pronounced at lower doses of CCh, or when SOCE was blocked. This suggests l-type Ca2+ channels might be a potential target for bronchoconstriction under certain circumstances.

Long-term follow-up of the incidence of Helicobacter pylori.

OBJECTIVES: Helicobacter pylori causes peptic ulcer disease and gastric cancer. Understanding the incidence of H. pylori could help guide research on potential infection prevention strategies. Previous studies indicate infection occurs in young children, but the risk of infection in older children and adolescents is unclear. Our hypothesis was that H. pylori infection is rare in adolescence or adulthood. Our aim was to determine the incidence of H. pylori over a prolonged follow-up in a cohort of 626 noninfected individuals. METHODS: Participants, including index children, mothers, fathers and siblings, from a previous study (1997-2002) were traced, and 883 of 946 participated in this extended follow-up. We used the 13C urea breath test (13C-UBT) to determine the incidence of H. pylori among 626 family members not infected in 2002, including 75 younger siblings who were not born or too young for testing in 2002. RESULTS: Eight (3.8%) of 210 index participants (mean ± standard deviation age 17.92 ± 0.77 years) became infected during 11.07 ± 0.56 years of follow-up (incidence, 3.42 per 1000 person-years; 95% confidence interval (CI), 1.48-6.74). Only one (0.6%) of 165 older siblings became infected (incidence, 0.57 per 1000 person-years; 95% CI, 0.007-3.16) and one of 176 parents became infected (incidence, 0.63 per 1000 person-years; 95% CI, 0.01-3.5). Of 75 younger siblings (age 10.9 ± 2.85 years) who were too young for testing or not yet born in 2002, nine (12%) became infected (incidence, 11.32 per 1000 person-years; 95% CI, 5.27-21.49). The highest incidence of H. pylori infection was in those born after 2005. CONCLUSIONS: The incidence of H. pylori was extremely low in older children and adults in developed countries. Spontaneous clearance of infection was uncommon in our study population.

Characteristics and outcomes of older persons attending the emergency department: a retrospective cohort study.

BACKGROUND: The analysis of routinely collected hospital data informs the design of specialist services for at-risk older people. AIM: Describe the outcomes of a cohort of older emergency department (ED) attendees and identify predictors of these outcomes. DESIGN: retrospective cohort study. METHODS: All patients aged 65 years or older attending an urban university hospital ED in January 2012 were included (N = 550). Outcomes were retrospectively followed for 12 months. Statistical analyses were based on multivariate binary logistic regression models and classification trees. RESULTS: Of N = 550, 40.5% spent ≤6 h in the ED, but the proportion was 22.4% among those older than 81 years and not presenting with musculoskeletal problems/fractures. N = 349 (63.5%) were admitted from the ED. A significant multivariate predictor of in-hospital mortality was Charlson comorbidity index [CCI; odds ratio = 1.19, 95% confidence interval: 1.07, 1.34, P = 0.002]. Among patients who were discharged from ED without admission or after their first in-patient admission (N = 499), 232 (46.5%) re-attended ED within 1 year, with CCI being the best predictor of re-attendance (CCI ≤ 4: 25.8%, CCI > 5: 60.4%). Among N = 499, 34 (6.8%) had died after 1 year of initial ED presentation. The subgroup (N = 114) with the highest mortality (17.5%) was composed by those aged >77 years and brought in by ambulance on initial presentation. CONCLUSIONS: Advanced age and comorbidity are important drivers of outcomes among older ED attendees. There is a need to embed specialist geriatric services within frontline services to make them more gerontologically attuned. Our results predate the opening of an acute medical unit with specialist geriatric input.

Cyclical vomiting syndrome in children: a prospective study.

BACKGROUND: Cyclical vomiting syndrome (CVS) is a disorder that affects all ages and is characterized by episodes of severe nausea and vomiting with symptom-free intervals between episodes. The incidence in children is 3.15/100 000 children per year. Our objective was to evaluate the natural history of CVS and examine factors that predict symptom resolution. METHODS: Thirty newly diagnosed children (mean 9.15 years, SD 3.31 range 3.5-15.7) were enrolled. All children had a follow-up interview at 3 months, 27/30 at 6 months, and 22/30 at 9 months. KEY RESULTS: Following diagnosis of CVS, only 5/22(22.7%) children had no further episodes of vomiting at 9 months, whereas 17/22 (77.3%) continued to vomit. In the year prior to diagnosis, 15/30 (50%) children were admitted to hospital. Of the 22 children with follow-up for 9 months, only one child required hospital admission. Children who continued to vomit had higher internalizing scores on CBCL compared with those who stopped vomiting (P = NS). The Pediatric Quality-of-Life Score suggested those who continued to vomit had a poorer quality of life at diagnosis compared with those who stopped vomiting (P < 0.05). CONCLUSIONS & INFERENCES: Making a positive diagnosis of CVS and providing families with information is very important in the management of CVS. Although 75% of children reported regular episodes of vomiting 9 months after diagnosis, there was a significant reduction in the frequency and severity of symptoms in addition to a marked reduction in the use of medical services.

Disease outcome for children who present with oral manifestations of Crohn's disease.

AIM: To describe the outcome for children with oral Crohn's disease (OCD) at diagnosis, and to determine if there was a difference in the Paediatric Crohn's Disease Activity Index (PCDAI) scores between those with and those without oral lesions at follow-up. METHODS: Thirty-one patients with OCD who had enrolled in two previous studies were invited to participate. Clinical and laboratory data were collected to calculate the PCDAI. Details of the management of Crohn's disease were also recorded. RESULTS: Twenty-four of 31 patients participated (77%), of whom 17 were boys (M:F = 2.4:1). Mean age at follow-up was 15.7 years (SD 1.98, range 11.9-19.7 years). Mean duration of follow-up was 55 months (SD 22, range 20-97 months). Oral manifestations were present at follow-up in 7 (29%) of 24 patients. There were no differences between patients with and without OCD at follow-up with regard to medical treatments received or intestinal disease location. There was no difference in median PCDAI scores between those who had and those who had not oral lesions at follow-up. CONCLUSIONS: OCD resolved in the majority of children treated for intestinal Crohn's disease. The occurrence of mouth lesions during follow-up of children who had oral manifestations at initial diagnosis was not a marker for Crohn's disease activity elsewhere in the intestinal tract.

Outcome for children with cyclical vomiting syndrome.

OBJECTIVE: Cyclical vomiting syndrome (CVS) is a disorder that carries a significant burden of disease for children and their families. The aim of this study was to examine the outcome of a group of children diagnosed with CVS from 1993 to 2003. METHODS: Children diagnosed with CVS over a 10-year period were identified and a review of the clinical records was carried out to define demographic features and the spectrum of disease at presentation. The patient's parent was contacted to establish the child's current well-being. Ethical approval for the study was obtained. RESULTS: Fifty one children were diagnosed with CVS and 41 agreed to participate in follow-up. Mean age was 5.8 (SD 3.3) years at onset of CVS, 8.2 (SD 3.5) years at diagnosis, and 12.8 (SD 4.8) years at follow-up. Vomiting had resolved at the time of follow-up in 25/41 (61%) children. Sixteen of 41 (39%) children reported resolution of symptoms either immediately or within weeks of diagnosis. However, a large number of children from the group whose vomiting resolved and the group that were still vomiting continued to have somatic symptoms, with 42% of children suffering regular headaches and 37% having abdominal pain. 32 (78%) parents felt that the provision of a positive diagnosis and information made a significant impact on the severity of vomiting. CONCLUSIONS: While 60% of children with CVS have resolution of symptoms, a significant proportion of both those in whom symptoms have resolved and those in whom vomiting persists continue to suffer from other somatic symptoms.

Focally enhanced gastritis in children with Crohn's disease and ulcerative colitis.

OBJECTIVES: Focally enhanced gastritis (FEG) has been suggested as a specific diagnostic marker for patients with Crohn's disease. However, the utility of FEG for distinguishing Crohn's disease from ulcerative colitis is uncertain in adults, and the occurrence of this lesion in children has not been defined. The aim of this study was to evaluate the occurrence of FEG and other gastric histological abnormalities in children with inflammatory bowel disease (IBD) and to examine the utility of FEG in discriminating between ulcerative colitis and Crohn's disease. METHODS: This is a retrospective, case-controlled study of upper GI histopathological findings in children with IBD. Gastric histopathology was defined and graded according to the Updated Sydney System. RESULTS: FEG was present in 28 of 43 (65.1%) children with Crohn's disease and five of 24 (20.8%) children with ulcerative colitis, compared to three of 132 (2.3%) children without IBD or one of 39 (2.6%) children with Helicobacter pylori infection. There were no differences between those with and without FEG with regard to upper GI symptoms or previous anti-inflammatory drug ingestion (5-aminosalicylic acid compounds or steroids). All patients with H. pylori infection had chronic antral gastritis, but only one child with H. pylori had FEG. In addition, mild to moderate chronic gastritis was present in 15 of 43 (34.9%) children with Crohn's disease and in 12 of 24 (50%) patients with ulcerative colitis. CONCLUSIONS: The presence of FEG suggests underlying IBD. Although FEG is particularly common in children with Crohn's disease, it does not reliably differentiate between Crohn's disease and ulcerative colitis.

Limitations to carbon 13-labeled urea breath testing for Helicobacter pylori in infants.

We determined the validity of the carbon 13-labeled urea breath test in young children. We found that although the 13C-labeled urea breath test had a specificity greater than 90%, borderline or false positive results occurred more frequently in children younger than 2 years compared with older children. False positive results may be caused by oral-urease-producing organisms because direct intragastric administration of 13C urea reduced the excess delta 13CO2. Care is urged in interpreting one positive 13C-labeled urea breath test in children younger than 2 years.

The oral cavity in Crohn's disease.

We assessed the utility of expert oral examination as a part of the diagnostic evaluation of patients with suspected Crohn's disease. Of 45 patients with newly diagnosed CD, 25 had been examined by a dentist. Twelve (48%) of these had oral CD lesions. Mucosal tags constituted the most frequent form of oral lesion (8/12). Of 8 oral biopsy specimens, 6 (75%) contained non-caseating granulomas. Patients with oral CD had more oral symptoms, presented for diagnosis sooner, and were more likely to have other upper gastrointestinal inflammation than those without oral lesions. Oral manifestations of CD are common in children; therefore, expert oral examination may be useful during diagnostic evaluation of children with suspected inflammatory bowel disease.

Antigastric autoantibodies in ferrets naturally infected with Helicobacter mustelae.

Infection with Helicobacter pylori has been associated with induction of autoantibodies that cross-react with the gastric mucosa. There have been discordant reports as to whether or not these autoantibodies arise due to molecular mimicry between H. pylori and host cell antigens on parietal cells. In this study, we investigated whether molecular mimicry by H. mustelae causes autoantibodies in infected ferrets. Serum from H. mustelae-infected ferrets reacted with parietal cells in the ferret gastric mucosa but not with duodenal or colonic mucosa. These sera did not react with the blood group A epitope on erythrocytes or H. mustelae lipopolysaccharide, and absorption with H. mustelae whole cells or red blood cells did not remove autoantibodies. In conclusion, ferrets naturally infected with H. mustelae generate antibodies that react with parietal cells, but these autoantibodies are not due to molecular mimicry.

Is Helicobacter pylori infection in childhood a risk factor for gastric cancer?

Helicobacter pylori infection is associated with chronic gastritis and peptic ulcer disease. Furthermore, the World Health Organization has classified this organism as a carcinogen for gastric cancer. H pylori infection is mainly acquired in childhood. Children with H pylori infection are asymptomatic except for a very small number that develop peptic ulcer disease. However, if H pylori gastritis is associated with gastric cancer, do pediatricians need to screen children for this infection and treat those who are infected? In an attempt to determine the significance of the association between H pylori and gastric cancer, we have reviewed all of the English language literature on this topic. H pylori infection seems to be associated with an increased risk of developing gastric cancer. However, only a small number of infected individuals (~1%) will develop gastric cancer. Furthermore, there are potential cofactors other than H pylori that could be equally important. The effect of the eradication of H pylori alone on the development of gastric cancer is unknown. Based on our knowledge to date, we suggest that it is not indicated to treat all children with H pylori infection because of the risk of developing gastric cancer or to institute a screening and treatment program.

A new polymorphism for the RI22H mutation in hereditary pancreatitis.

BACKGROUND AND AIMS: Hereditary pancreatitis (HP) is a rare form of recurrent acute and chronic pancreatitis. Mutations in the cationic trypsinogen (protease serine 1, PRSS1) gene have been identified as causing HP. The R122H (previously known as R117H) mutation is the commonest and can be detected by a single and rapid polymerase chain reaction/restriction fragment length polymorphism (PCR/RFLP) based technique using the AflIII enzyme. This test however may give a false negative result in the presence of a neutral polymorphism within the enzyme recognition site. The frequency of this event was examined by sequencing studies in patients with HP and in healthy controls. METHODS: Of 60 families identified by the UK and Ireland consortium of EUROPAC (European Registry for Hereditary Pancreatitis and Familial Pancreatic Cancer), 51 were screened for R122H, N29I, and A16V mutations using standard techniques, and by sequencing of all five exons of cationic trypsinogen. RESULTS: Twelve families had the N29I mutation, one family had A16V and, on standard testing, 15 families had the R122H mutation. An additional family with the R122H mutation was found on direct sequencing. The false negative result was due to a neutral polymorphism C-->T at the third base of the codon, not affecting the amino acid coded for, destroying the AflIII restriction site. This polymorphism was not observed in 50 DNA specimens (100 chromosomes) from controls nor from 50 individuals from PRSS1 mutation negative HP families. A novel mutation specific PCR was developed to avoid this pitfall. CONCLUSIONS: One of the 16 families with HP and an R122H mutation contained a polymorphism affecting the AflIII restriction site. Adoption of an alternative R122H assay is important for genetic studies in individuals with apparent HP.

Adherence of isogenic flagellum-negative mutants of Helicobacter pylori and Helicobacter mustelae to human and ferret gastric epithelial cells.

Isogenic flagellum-negative mutants of Helicobacter pylori and Helicobacter mustelae were screened for their ability to adhere to primary human and ferret gastric epithelial cells, respectively. We also evaluated the adherence of an H. pylori strain with a mutation in the flbA gene, a homologue of the flbF/lcrD family of genes known to be involved in the regulation of H. pylori flagellar biosynthesis. H. pylori and H. mustelae mutants deficient in production of FlaA or FlaB and mutants deficient in the production of both FlaA and FlaB showed no reduction in adherence to primary human or ferret gastric epithelial cells compared with the wild-type parental strains. However, adherence of the H. pylori flbA mutant to human gastric cells was significantly reduced compared to the adherence of the wild-type strain. These results show that flagella do not play a direct role in promoting adherence of H. pylori or H. mustelae to gastric epithelial cells. However, genes involved in the regulation of H. pylori flagellar biosynthesis may also regulate the production of an adhesin.

Pancreatic duct stenting as a treatment for hereditary pancreatitis.

Hereditary pancreatitis is a genetically transmitted condition usually presenting in childhood or adolescence. The natural history of the condition is that recurrent episodes of pancreatitis may be followed by the development of pancreatic exocrine and endocrine failure. Treatment options are limited, usually consisting of surgical drainage procedures whose efficacy is uncertain and whose effect on disease progression is unknown. We report a child with hereditary pancreatitis treated by means of a pancreatic duct stent placed via endoscopic retrograde cholangiopanctreatography resulting in long-term control of symptoms and speculate that earlier intervention may alter the disease course.