Identification of a rhythmic firing pattern in the enteric nervous system that generates rhythmic electrical activity in smooth muscle

The enteric nervous system (ENS) contains millions of neurons essential for organization of motor behavior of the intestine. It is well established that the large intestine requires ENS activity to drive propulsive motor behaviors. However, the firing pattern of the ENS underlying propagating neurogenic contractions of the large intestine remains unknown. To identify this, we used high-resolution neuronal imaging with electrophysiology from neighboring smooth muscle. Myoelectric activity underlying propagating neurogenic contractions along murine large intestine [also referred to as colonic migrating motor complexes, (CMMCs)] consisted of prolonged bursts of rhythmic depolarizations at a frequency of ∼2 Hz. Temporal coordination of this activity in the smooth muscle over large spatial fields (∼7 mm, longitudinally) was dependent on the ENS. During quiescent periods between neurogenic contractions, recordings from large populations of enteric neurons, in mice of either sex, revealed ongoing activity. The onset of neurogenic contractions was characterized by the emergence of temporally synchronized activity across large populations of excitatory and inhibitory neurons. This neuronal firing pattern was rhythmic and temporally synchronized across large numbers of ganglia at ∼2 Hz. ENS activation preceded smooth muscle depolarization, indicating rhythmic depolarizations in smooth muscle were controlled by firing of enteric neurons. The cyclical emergence of temporally coordinated firing of large populations of enteric neurons represents a unique neural motor pattern outside the CNS. This is the first direct observation of rhythmic firing in the ENS underlying rhythmic electrical depolarizations in smooth muscle. The pattern of neuronal activity we identified underlies the generation of CMMCs

Use of severity classification systems in the surgical decision-making process in emergency laparotomy for perforated diverticulitis

Purpose: Hartman's procedure (HP) or primary anastomosis (PA) are the two surgical techniques used in patients undergoing emergency colectomy for perforated diverticulitis. There are no objective criteria to guide the surgeon's choice of procedure. This study assesses whether classification and scoring systems can be used in the decision-making process. Methods: One hundred eleven patients undergoing emergency laparotomy for perforated diverticulitis were analyzed. Logistic regression and interaction models were used to determine the predictive value in the two settings. Results: Sixty five patients underwent HP and 46 patients underwent PA. Patients with HP had significantly higher scores, median age, and were more often on immunosuppressive medication. Mortality and surgical morbidity did not differ between the groups. The clinical anastomotic leak rate was 28.3% in the PA group. In the univariate logistic regression for in-hospital death, all scores showed a significant influence. The multivariate logistic regression analysis showed that only Charlson comorbidity index (CCI) and American Society of Anesthesiologists score had a significant influence on mortality. Each score was analyzed for its predictive value regarding mortality and morbidity with respect to type of operative procedure. Only CCI revealed a trend towards statistical significance. The risk of death increases with increasing CCI when PA is performed compared to HP. Conclusion: None of the tested scores can be used to help the surgeon decide whether a PA or HP is appropriate in a specific patient. Comorbidity, represented as CCI in this study, might be more important than the locoregional situatio

Identification of unique release kinetics of serotonin from guinea-pig and human enterochromaffin cells

This is the accepted version of the following article: [Raghupathi, R., Duffield, M. D., Zelkas, L., Meedeniya, A., Brookes, S. J. H., Sia, T. C., Wattchow, D. A., Spencer, N. J. and Keating, D. J. (2013), Identification of unique release kinetics of serotonin from guinea-pig and human enterochromaffin cells. The Journal of Physiology, 591: 5959–5975. doi: 10.1113/jphysiol.2013.259796], which has been published in final form at [http://dx.doi.org/10.1113/jphysiol.2013.259796]. In addition, authors may also transmit, print and share copies with colleagues, provided that there is no systematic distribution of the submitted version, e.g. posting on a listserve, network or automated delivery

A panel of genes methylated with high frequency in colorectal cancer

Background: The development of colorectal cancer (CRC) is accompanied by extensive epigenetic changes, including frequent regional hypermethylation particularly of gene promoter regions. Specific genes, including SEPT9, VIM1 and TMEFF2 become methylated in a high fraction of cancers and diagnostic assays for detection of cancer-derived methylated DNA sequences in blood and/or fecal samples are being developed. There is considerable potential for the development of new DNA methylation biomarkers or panels to improve the sensitivity and specificity of current cancer detection tests. Methods: Combined epigenomic methods - activation of gene expression in CRC cell lines following DNA demethylating treatment, and two novel methods of genome-wide methylation assessment - were used to identify candidate genes methylated in a high fraction of CRCs. Multiplexed amplicon sequencing of PCR products from bisulfite-treated DNA of matched CRC and non-neoplastic tissue as well as healthy donor peripheral blood was performed using Roche 454 sequencing. Levels of DNA methylation in colorectal tissues and blood were determined by quantitative methylation specific PCR (qMSP). Results: Combined analyses identified 42 candidate genes for evaluation as DNA methylation biomarkers. DNA methylation profiles of 24 of these genes were characterised by multiplexed bisulfite-sequencing in ten matched tumor/normal tissue samples; differential methylation in CRC was confirmed for 23 of these genes. qMSP assays were developed for 32 genes, including 15 of the sequenced genes, and used to quantify methylation in tumor, adenoma and non-neoplastic colorectal tissue and from healthy donor peripheral blood. 24 of the 32 genes were methylated in \u3e50% of neoplastic samples, including 11 genes that were methylated in 80% or more CRCs and a similar fraction of adenomas. Conclusions: This study has characterised a panel of 23 genes that show elevated DNA methylation in \u3e50% of CRC tissue relative to non-neoplastic tissue. Six of these genes (SOX21, SLC6A15, NPY, GRASP, ST8SIA1 and ZSCAN18) show very low methylation in non-neoplastic colorectal tissue and are candidate biomarkers for stool-based assays, while 11 genes (BCAT1, COL4A2, DLX5, FGF5, FOXF1, FOXI2, GRASP, IKZF1, IRF4, SDC2 and SOX21) have very low methylation in peripheral blood DNA and are suitable for further evaluation as blood-based diagnostic markers

Discovery and Validation of Molecular Biomarkers for Colorectal Adenomas and Cancer with Application to Blood Testing

BACKGROUND & AIMS: Colorectal cancer incidence and deaths are reduced by the detection and removal of early-stage, treatable neoplasia but we lack proven biomarkers sensitive for both cancer and pre-invasive adenomas. The aims of this study were to determine if adenomas and cancers exhibit characteristic patterns of biomarker expression and to explore whether a tissue-discovered (and validated) biomarker is differentially expressed in the plasma of patients with colorectal adenomas or cancer. METHODS: Candidate RNA biomarkers were identified by oligonucleotide microarray analysis of colorectal specimens (222 normal, 29 adenoma, 161 adenocarcinoma and 50 colitis) and validated in a previously untested cohort of 68 colorectal specimens using a custom-designed oligonucleotide microarray. One validated biomarker, KIAA1199, was assayed using qRT-PCR on plasma extracted RNA from 20 colonoscopy-confirmed healthy controls, 20 patients with adenoma, and 20 with cancer. RESULTS: Genome-wide analysis uncovered reproducible gene expression signatures for both adenomas and cancers compared to controls. 386/489 (79%) of the adenoma and 439/529 (83%) of the adenocarcinoma biomarkers were validated in independent tissues. We also identified genes differentially expressed in adenomas compared to cancer. KIAA1199 was selected for further analysis based on consistent up-regulation in neoplasia, previous studies and its interest as an uncharacterized gene. Plasma KIAA1199 RNA levels were significantly higher in patients with either cancer or adenoma (31/40) compared to neoplasia-free controls (6/20). CONCLUSIONS: Colorectal neoplasia exhibits characteristic patterns of gene expression. KIAA1199 is differentially expressed in neoplastic tissues and KIAA1199 transcripts are more abundant in the plasma of patients with either cancer or adenoma compared to controls