Fulminant ectopic Cushing's syndrome caused by metastatic small intestine neuroendocrine tumour - a case report and review of the literature.

Cushing's syndrome (CS) secondary to adrenocorticotropic hormone (ACTH) producing tumours is a severe condition with a challenging diagnosis. Ectopic ACTH-secretion often involves neuroendocrine tumours (NET) in the respiratory tract. ACTH-secreting small intestine neuro-endocrine tumours (siNET) are extremely rare entities barely reported in literature. This review is illustrated by the case of a 75-year old woman with fulminant ectopic CS caused by a ACTH-secreting metastatic siNET. Severe hypokalemia, fluid retention and refractory hypertension were the presenting symptoms. Basal and dynamic laboratory studies were diagnostic for ACTH-dependent CS. Extensive imaging studies of the pituitary and thorax-abdomen areas were normal, while [68Ga]Ga-DOTATATE PET-CT revealed increased small intestine uptake in the left iliac fossa. The hypercortisolism was well controlled with somatostatin analogues, after which a debulking resection of the tumour was performed. Pathological investigation confirmed a well-differentiated NET with sporadic ACTH immunostaining and post-operative treatment with somatostatin analogues was continued with favourable disease control.

A single-centre analysis of post-colonoscopy colorectal cancer.

PATIENTS AND METHODS: A prospective registration of patients with colorectal cancer and a colonoscopy within the last 10 years. We tried to classify these post-colonoscopy colorectal cancers (PCCRCs) by most reasonable explanation and into subcategories suggested by the World Endoscopy Organization (WEO) and calculated the unadjusted PCCRC rate. RESULTS: 47 PCCRCs were identified. The average age at diagnosis of PCCRC was 73 years. PCCRCs were more located in the right colon with a higher percentage of MSI-positive and B-RAF mutated tumours. The average period between index colonoscopy and diagnosis of PCCRC was 4.2 years. Sixty-eight % of all PCCRCs could be explained by procedural factors. The mean PCCRC-3y of our department was 2.46%. CONCLUSIONS: The data of our centre are in line with the data of the literature from which can be concluded that most postcolonoscopy colorectal cancers are preventable. The PCCRC-3y is an important quality measure for screening colonoscopy. Ideally all centres involved in the population screening should measure the PCCRC-3 y annually, with cooperation of the cancer registry and reimbursement data provided by the Intermutualistic Agency (IMA).

Supernatants of irritable bowel syndrome mucosal biopsies impair human colonic smooth muscle contractility.

BACKGROUND: Changes in intestinal motility are likely to contribute to irritable bowel syndrome (IBS) pathophysiology. The aim of the study was to investigate the effects of IBS mucosal supernatants on human colonic muscle contractility. METHODS: Supernatants were obtained from biopsies of 18 IBS patients-nine with constipation (IBS-C) and nine with diarrhea-predominant IBS (IBS-D)-and nine asymptomatic subjects, used as controls. Colonic circular smooth muscle strips or isolated cells (SMC) were exposed to control or IBS supernatants. Spontaneous phasic contractions on strips and morphofunctional parameters on cells were evaluated in basal conditions and in response to acetylcholine (Ach). Incubation with IBS supernatants was also conducted in the presence of antagonists and inhibitors (namely histamine, protease and prostaglandin antagonists, nuclear factor-kappa B inhibitor, catalase, NADPH oxidase inhibitor, and the cAMP- and/or cGMP-cyclase inhibitors). KEY RESULTS: Exposure to IBS-C and IBS-D supernatants induced a significant reduction in basal tone and Ach-elicited contraction of muscle strips and a significant shortening and impairment of Ach contraction of SMCs. The NADPH oxidase inhibitor prevented the effect of supernatants, while the protease antagonist only IBS-C effect. No effect was observed with the other antagonists and inhibitors. Dilution of IBS-D supernatants partially restored the effects only on SMCs, whereas dilution of IBS-C supernatants significantly reverted the effects on muscle strips and Ach-elicited response on SMC. CONCLUSIONS & INFERENCES: Supernatants from mucosal biopsies of IBS patients reduce colonic contractility. The observed impairment was concentration dependent, likely occurring through intracellular oxidative stress damage, involving different neuromotor mechanisms depending on the IBS subtype.

Atropisomerism in 3-arylthiazolidine-2-thiones. A combined dynamic NMR and dynamic HPLC study.

Sterically hindered 3-arylthiazolidine-2-thiones were prepared by a solvent-free reaction with arylisothiocyanates and 1,4-dithiane-2,5-diol. Atropisomerism was observed in two compounds (3 and 4, aryl = 1-naphthyl and 2-methylnaphth-1-yl), whose rotational energy barriers were measured using dynamic NMR and dynamic HPLC. The experimental analyses were supported by DFT calculations. Thermally stable atropisomers were obtained by dehydration of compounds 3 and 4 and the absolute configuration of the atropisomers of compound 6 was determined by theoretical simulation of the ECD and VCD spectra.

Patología ocular en niños: estudio de validación de un método de cribado en la etapa escolar: marzo–diciembre 2010, Montevideo, Uruguay / Ocular pathologies in children: validation study of a screening method during school years: march-december 2010, Montevideo, Uruguay / Patologia ocular em crianças: estudo de validação de um método de triagem na etapa escolar: março-dezembro 2010 Montevidéu, Uruguai

El Programa Nacional de Salud Ocular tiene por objetivo abordar las causas evitables o tratables de ceguera. En este marco, el Departamento de Medicina Familiar y Comunitaria ha desarrollado actividades de Promoción de salud y Prevención de enfermedades y problemas oculares en la infancia. Esta actividad se planificación se llevó a cabo en interdisciplina con la participación de estudiantes de grado de medicina. Este trabajo incorpora la evaluación de las competencias adquiridas por los estudiantes en la realización del cribado de alteraciones visuales (grado de concordancia inter-observador). El Objetivo fue evaluar validez y seguridad de la aplicación de un método de cribado de patología ocular en escolares y preescolares por estudiantes de medicina en tres instituciones del Zonal 12 de Montevideo (marzo-diciembre 2010). Conclusiones: El porcentaje de derivados es similar a trabajos realizados anteriormente. A partir de los resultados del trabajo sería oportuno incorporar la realización del cribado de alteraciones visuales por estudiantes de medicina capacitados, en las instituciones educativas, lo cual optimiza las intervenciones en el primer nivel de atención.

The objective of the National Program of Ocular Health is to tackle the avoidable and treatable causes of blindness. Within this framework, the Department of Community and Family Medicine has developed activities related to Health Promotion and Prevention of ocular illnesses and problems in childhood. This activity was planned and took place in an interdisciplinary way, with the participation of medical degree students, students of opthalmology, and ophthalmologists and with the participation of different sectors such as Hospital de Ojos, Ministry of Social Development and the Chamber of Opticians. This work includes the assessment of competences acquired by the students when screening visual alterations (inter-observer agreement degree). The objective was to evaluate the validity and safety of medical students applying a screening method of ocular pathology in kindergarten and school students at three institutions of Zonal 12 in Montevideo (MarchDecember 2010). Conclusions: The percentage of derivatives is similar to that of previous works. From the results of the study, it would be appropriate to include the screening of visual alterations by trained medical students at educational institutions, since it optimizes interventions at primary health care.

O Programa Nacional de Saúde Ocular tem como alvo a abordagem das causas evitáveis ou tratáveis da cegueira. Neste contexto, o Departamento de Medicina Familiar e Comunitária levou a cabo atividades de Promoção da saúde e Prevenção de doenças e problemas oculares na infância. Este programa foi planejado e realizado de maneira interdisciplinar com a participação de estudantes de medicina, de estudantes do bacharelado em oftalmologia, bacharéis em oftalmologia e oftalmologistas, e em parceria com de Ópticos. Este trabalho inclui a avaliação das competências adquiridas pelos estudantes na realização da triagem dos distúrbios visuais (grau de concordância inter-observador). O objetivo foi avaliar a validade e segurança de aplicação de um método de triagem para doenças oculares nos escolares e pré-escolares por estudantes de medicina em três instituições do Zonal Montevidéu 12 (de março a dezembro de 2010). Conclusões: A porcentagem de derivados é semelhante a trabalhos realizados previamente. A partir dos resultados do trabalho achase conveniente integrar a realização de triagem de distúrbios visuais por estudantes de medicina treinados em instituições educacionais, otimizando-se dessa maneira as intervenções nos cuidados primários.

Role of endoscopic sphincterotomy of the minor papilla in pancreas divisum.

BACKGROUND: Pancreas divisum (PD) is a common anatomical variant of the pancreatic duct system. Only a little percentage of patients with this altered anatomy develop symptoms: acute recurrent pancreatitis (ARP), chronic pancreatitis (CP) and pancreatic-type pain alone. These have been supposed to arise from an obstruction to outflow of the pancreatic dorsal duct due to a stenosis of the minor papilla. Endoscopic sphincterotomy of the minor papilla (MiES) can be considered an effective treatment for patients with PD and ARP supported by stenosis or obstruction of the minor papilla. On the other hand, the access through the minor papilla is essential for therapeutic pancreatic endoscopy in patients with PD and CP, but the effectiveness of MiES in these patients is extremely controversial. MATERIALS AND METHODS: Aim of this brief review has been to evaluate the short and long-term effects of endoscopic pancreatic interventions in patients affected by symptomatic PD who underwent therapeutic endoscopic retrograde cholangiopancreatography (ERCP) with pancreatic sphincterotomy of the minor papilla (MiES). The main literature database was Medline (1966-2013). RESULTS: Data come from a personal overview of articles emerging from the same search strategy. CONCLUSIONS: Nowadays, when endoscopic pancreatic drainage is indicated in patient with symptomatic PD, access through the minor papilla (MiP) is required. Therapeutic endoscopic interventions provide less invasive alternatives to the surgical approach.

Contribution of intestinal smooth muscle to Crohn's disease fibrogenesis.

Mesenchymal cells transdifferentiation and extracellular matrix deposition are involved in the fibrotic process of Crohn's disease (CD). Mesenchymal smooth muscle cells (SMCs) de-differentiation, driven by Platelet-derived growth factor (PDGF) that counteracts Transforming growth factor (TGF-ß) has been studied in vascular muscle. The role of SMCs in intestinal fibrogenesis is still not clearly elucidated. Aim of the study was to evaluate the possible myogenic contribution to CD fibrotic process through the comparative analysis of histological, morphometric and molecular alterations occurring in human smooth muscle. Full thickness specimens were obtained from CD (non-involved and stenotic tracts) and healthy (control) ileum. Tissues were processed for histological and immunohistochemical (IHC) analyses and SMCs were isolated from the muscularis propria for morphofunctional and molecular (qPCR) analyses. CD stenotic ileum showed a significant increased thickness of all layers compared to CD non-involved and control ileum. IHC revealed an overexpression of α-smooth muscle actin and collagens I-III throughout all intestinal layers only in stenotic tracts. The two growth factors, PDGF and TGF-ß, showed a progressive increase in expression in the muscle layer from CD non-involved to stenotic tracts. Freshly isolated SMCs presented alterations in CD non-involved tracts that progressively increased in the stenotic tracts consisting in a statistical increase in mRNA encoding for PDGF-ß and collagen III, paralleled to a decrease in TGF-ß and Tribbles-like protein-3 mRNA, and altered morphofunctional parameters consisting in progressive decreases in cell length and contraction to acetylcholine. These findings indicate that intrinsic myogenic alterations occur in CD ileum, that they likely precede stricture formation, and might represent suitable new targets for anti-fibrotic interventions.

Lactobacillus rhamnosus protects human colonic muscle from pathogen lipopolysaccharide-induced damage.

BACKGROUND: Lactobacillus species might positively affect gastrointestinal motility. These Gram-positive bacteria bind Toll-like receptor 2 (TLR2) that elicits anti-inflammatory activity and exerts protective effects on damage induced by lipopolysaccharide (LPS). Whether such effect occurs in gastrointestinal smooth muscle has not been established yet. Aim of this study was to characterize the effects of Lactobacillus rhamnosus GG (LGG) and of supernatants harvested from LGG cultures on human colonic smooth muscle and to explore their protective activity against LPS-induced myogenic morpho-functional alterations. METHODS: The effects of LGG (ATCC 53103 strain) and of supernatants have been tested on both human colonic smooth muscle strips and isolated cells in the absence or presence of LPS obtained from a pathogenic strain of Escherichia coli. Their effects on myogenic morpho-functional properties, on LPS-induced NFκB activation, and on cytokine production have been evaluated. Toll-like receptor 2 expression has been analyzed by qPCR and flow cytometry. KEY RESULTS: Lactobacillus rhamnosus GG exerted negligible transient effects per se whereas it was capable of activating an intrinsic myogenic response counteracting LPS-induced alterations. In particular, both LGG and supernatants significantly reduced the LPS-induced morpho-functional alterations of muscle cells, i.e. cell shortening and inhibition of contractile response. They also hindered LPS-induced pro-inflammatory effects by decreasing pro-inflammatory transcription factor NFκB activation and pro-inflammatory cytokine IL-6 secretion, and restored the secretion levels of anti-inflammatory cytokine IL10. CONCLUSIONS & INFERENCES: Taken together these data demonstrate that LGG protects human colonic smooth muscle from LPS-induced myogenic damage and might be beneficial on intestinal motor disorders due to bacterial infection.

Chronic constipation in hypercalcemic patients with primary hyperparathyroidism.

BACKGROUND: Chronic constipation (C), among gastrointestinal symptoms, is commonly associated with primary hyperparathyroidism (PHPT) and probably attributable to hypercalcemia. OBJECTIVE OF THE STUDY: To evaluate in patients affected with PHPT the prevalence of C utilizing a validated questionnaire and the current prevalence of C compared to that observed in the past and to evaluate the relationship between C and the severity of PHPT. METHODS: 55 outpatients affected with PHPT, admitted to our Department of Internal Medicine and Medical Specialities in the years (2006-2009) were studied (group 1: 50 postmenopausal women and 5 men, mean age 61.9 +/- 9.4 years), together with 55 sex and age matched controls (group 2). Also considered were a group of PHPT patients observed, in the same ambulatory, during the years '70-'80 (group 3). A questionnaire, Rome II criteria, was administered and used to define C, whereas only anamneses were used to define C in group 3. RESULTS: The prevalence of C in patients with PHPT was 21.8% in group 1 vs 12.7% in group 2 (n.s.) and 32.7% in group 3. There is a decreasing trend in the prevalence of C in patients with PHPT as observed from 1970-89 to 2006-2009 (p < 0.05). The reduction of C was associated together with a significant reduction in the serum calcium level (p < 0.001). The presence of C vs its absence in patients with PHPT is characterized by higher values of calcemia (p < 0.001), ionized calcium (p < 0.001), and parathyroid hormone (p = 0.019). CONCLUSION: The actual prevalence of C in patients with PHPT is not significantly different from that found in the control group and is decreasing with respect to the past years. Moreover, C seems to be associated with the severity of the disease rather than with the diagnosis of PHPT per se.

Myogenic regional responsiveness to cholinergic and vipergic stimulation in human colon.

BACKGROUND: Differences in the actions of enteric neurotransmitters on colonic circular and longitudinal muscle layers have not been clearly determined, nor the possible existence of intrinsic myogenic phenotypes that might contribute to regional differences in human colon motor activity. The aim of this study was to analyze the direct pharmaco-mechanical coupling of carbachol (CCh) and vasoactive intestinal polypeptide (VIP) on human colonic smooth muscle strips and cells. METHODS: Circular and longitudinal muscle strips and cells were obtained from 15 human specimens of ascending and sigmoid colon. Both isometric tension on muscle strips and contraction and relaxation on cells were measured in response to increasing CCh and VIP concentrations. KEY RESULTS: Circular muscle strips of ascending colon were more sensitive to the effect of CCh than that of sigmoid colon, EC(50) values being, respectively, 4.15µmolL(-1) and 8.47µmolL(-1) (P<0.05), although there were no differences in maximal responses. No regional differences were observed in longitudinal muscle strips or in smooth muscle cells. Maximal responses to CCh were higher on circular than longitudinal muscle strips and cells throughout the colon. A greater sensitivity to VIP was observed in ascending colon compared with sigmoid colon, both in circular (EC(50:) 0.041 and 0.15µmolL(-1) , respectively, P<0.01) and longitudinal (EC(50:) 0.043 and 0.09µmolL(-1) , respectively, P<0.05) strips, and similar differences were observed in longitudinal smooth muscle cells (EC(50:) 44.85 and 75.24nmolL(-1) , respectively, P<0.05). CONCLUSIONS & INFERENCES: Regional myogenic differences in pharmaco-mechanical coupling between the enteric neurotransmitters and smooth muscle contribute to the complex regional motor patterns of human colon.

Exocrine gastric secretion and gastritis: pathophysiological and clinical relationships.

Gastric exocrine secretion, both acid and non-acid, is required for micronutrients absorption, such as iron, calcium and vitamin B12, drugs absorption, protein digestion. Clinical presentation of a gastric secretion impairment might be then characterized by the presence of both gastrointestinal and non-gastrointestinal specific symptoms (i.e. anemia) or to a non-response to therapies. The main factor that impairs gastric exocrine secretion homeostasis is mucosal chronic inflammation that principally occurs after colonization by Helicobacter pylori (Hp). The extent and distribution of gastritis ultimately determine the clinical outcome linked to differences in gastric acid secretion status, the involvement of gastric body leading to a decrease in gastric exocrine secretion with possible progression to mucosal atrophy towards cancer. A correct clinical strategy in the management of Hp infected patients should be then to early identify body involvement, a diagnosis generally missed in that body biopsies are not routinely performed. The use of gastric serological markers, gastrin and pepsinogens, are helpful in suspecting the presence of mucosal atrophy but their diagnostic accuracy for non-atrophic chronic gastritis topography is not adequate despite a good specificity due to the low sensitivity, of all the available biomarkers. Gastric serology associated to anemia/iron-deficiency screening might nevertheless been helpful in the framing of patients that undergo endoscopy in order to highlight the need of extensive mucosal biopsies sampling.

Microscopic colitis: a concise review.

Microscopic colitis is an increasingly common cause of chronic watery diarrhoea, and often a causes of abdominal pain of unknown origins. The increase of interest for this clinical entity is due to a misdiagnosis of any symptoms that have been frequently attributed to diarrhea-predominant irritable bowel syndrome, often for many years before diagnosis. Presumably, most estimates of incidence and prevalence understate the true frequency of microscopic colitis for this reason. The aim of this paper is to evaluate the importance of microscopic colitis as cause of chronic non bloody diarrhoea, on the basis of literature review. These kind of colitis are characterized by normal colonic mucosa at endoscopy or barium enema but with increased inflammation in colonic biopsies. Microscopic colitis consists of two main subtypes, collagenous colitis and lymphocytic colitis, distinguished by the presence of absence of a thickened subepithelial collagen band. Several models of pathogenesis has been proposed but no convincing mechanism has been identified, although is difficult to characterize this clinical entity as an independent phenomenon or a simple manifestation or related factors active to induce microscopic changing in the colonic mucosa. A rational approach to therapy does not exist and was conduct with several types of drugs after the exclusion of other causes, commonly characterized by this symptoms and the definitive histological assessment in the biopsies specimens. In the majority of cases this condition tends to follow a self-limited course but potentially can assume the characteristics of relapsing course with the necessity to a chronic therapy. Several long-term follow-up studies excluded a possible progression to neoplastic malignancies of microscopic colitis.

Correlation between faecal calprotectin and magnetic resonance imaging (MRI) in the evaluation of inflammatory pattern in Crohn's disease.

BACKGROUND: Calprotectin, a major cytosolic protein of neutrophils, is increased in inflammatory bowel disease (IBD) and may be considered a suitable marker of intestinal inflammation. Abdominal MRI is becoming more frequently used for the evaluation of IBD patients. Aim of this study was to investigate the role of MRI in IBD for the assessment of disease activity in comparison with faecal calprotectin levels. PATIENTS AND METHODS: Twenty-four consecutive hospitalized pts (12 F, 12 M, median age: 56; range: 22-77) with a proven diagnosis of CD were studied. At the time of the MRI examination, pts provided a single stool sample for calprotectin measurement. Calprotectin was measured by ELISA (Calprest(R)). Pathological values were considered more than 50 microg/g. All pts underwent MRI, performed at 1.5 T, with HASTE T2w with and without fat-saturation, FLASH T1w fat-saturated sequences pre and post iv injection of 0.1 ml/kg of Gadolinium. Presence, degree and length of wall inflammation were evaluated. The MRI degree of wall inflammation was graded with a 0-3 scoring system (0=absent 1=light 2=moderate 3=severe) by considering findings observed on T1 post Gd and T2 fat-suppressed images, as well as the degree of wall thickness. The length of extension was considered as less than 15 cm, between 15 cm and 30 cm, or more than 30 cm. Spearman's correlation coefficient was used to evaluated differences in calprotectin levels among the groups obtained by MRI findings. RESULTS: Grade 0 MRI was found in 1 pt with a faecal calprotectin measurement of 206.25 microg/g; Grade 1 MRI was found in 4 pts with a median faecal calprotectin of 100 microg/g (5-325); Grade 2 MRI was found in 10 pts with a median faecal calprotectin of 243.75 microg/g (7.5-606.25); Grade 3 MRI was found in 9 pts with a median faecal calprotectin of 1012.5 microg/g (30-1268.8). A trend of positive correlation was therefore found between MRI scores of activity and calprotectin levels (p less than 0.0001) and between MRI scores of thickening of intestinal involvement and calprotectin levels (p = 0.005). No apparent correlation was observed between faecal calprotectin concentration and length. CONCLUSIONS: Data presenting show that faecal calprotectin levels well correlate with the degree of mucosal inflammation are in agreement with previous studies. Considering the correlation obtained between calprotectin level and MRI findings, we believe that MRI is helpful in assessing and monitoring the degree of disease in Crohn's disease.

High efficacy of bismuth subcitrate for Helicobacter pylori eradication in pangastritis.

BACKGROUND: The influence of gastritis patterns in Helicobacter pylori eradication rates has been poorly investigated. AIMS: To compare the efficacy of bismuth or proton pump inhibitors triple therapy for H. pylori eradication in pangastritis. PATIENTS AND METHODS: One hundred and eight patients with pangastritis were assigned to either lansoprazole 30 mg once a day (n=54) or bismuth 240 mg bis in die (n=54) for 14 days combined, for the first week, with amoxicillin 1g plus metronidazole 250 mg tris in die. Eradication was confirmed by (13)C-urea breath test. RESULTS: With bismuth, successful eradication was observed in 75.9% (41/54) in the intention-to-treat analysis and 78.8% (41/52) in the per-protocol analysis. With lansoprazole, the eradication rates were respectively 46.3% (25/54) and 51.0% (25/49). Bismuth had a significant higher efficacy according to both intention-to-treat analysis (p=0.0029) and per-protocol analysis (p=0.0038) with OR of 3.66 (95% CI: 1.61-8.32) and 3.58 (95% CI: 1.50-8.54) respectively. At regression analysis, the only independent variable affecting eradication was the type of regimen (p=0.026) with an OR of 3.31 (95% CI: 1.16-9.44). CONCLUSIONS: In pangastritis patients, bismuth is more effective than PPI in first-line eradication. For improving the overall eradication rates, an evaluation of gastritis extent might need to be considered.

Vasoactive intestinal peptide receptor subtypes and signalling pathways involved in relaxation of human stomach.

Vasoactive intestinal peptide (VIP) relaxes smooth muscle by interacting with receptors coupled to cAMP- or cGMP-signalling pathways. Their relative contribution to human gastric relaxation is unknown. This study aimed at investigating, in terms of biological activity, receptor expression and related signalling pathways, the action of VIP separately on the human fundus and the antrum. VIP caused greater relaxation of smooth muscle cells (SMC) and strips of the antrum presenting on the former a higher efficacy and potency (ED(50): 0.53 +/- 0.17 nmol L(-1)) than on the fundus (ED(50): 3.4 +/- 1.4 nmol L(-1)). On both fundus and antrum strips, its effect was tetrodotoxin insentitive. Reverse transcriptase-polymerase chain reaction analysis showed the sole expression of VPAC2 and natriuretic peptide clearance receptors, with VPAC2 being more abundant in the antrum. Functional regional differences in receptor-related signalling pathways were found. Activation of the cAMP-pathway by forskolin or its inhibition by adenylate cyclase (2'5'-dideoxyadenosine) or kinase (Rp-cAMPs) inhibitors had more pronounced effects on antrum SMC. Activation of the cGMP-pathway by sodium nitroprusside or its inhibition by guanylate cyclase (LY83583) or kinase (KT5823) inhibitors had more effects on fundus SMC, on which a higher expression of endothelial nitric oxide synthase was found. In conclusion, regional differences in VIP action on human stomach are related to distinct myogenic properties of SMC of the antrum and the fundus.

Somatostatin receptor subtypes mediate contractility on human colonic smooth muscle cells.

This study demonstrates the expression of functional somatostatin receptor (sstr) subtypes in human circular and longitudinal colonic smooth muscle cells (SMC). Native somatostatin (SS) and sstr subtype-specific analogues were used to characterize the sstr subtypes present in both cell types by contraction/relaxation studies. Qualitative and quantitative mRNA analysis and immunohistochemistry of sstr subtypes were also carried out. sstr subtype 2 mRNA was expressed in circular SMC, and various levels of subtypes 1, 2 and 3 mRNA were expressed in longitudinal colonic SMC. Native SS and each subtype-specific analogue exerted a modest, but significant, contraction, although inhibition of carbachol-induced contraction (relaxation) was the main effect on SMC from both layers. CH-288, a sstr subtype 1-specific analogue, and octreotide, a sstr subtype 2-specific analogue, were the most effective relaxant analogues on longitudinal and circular SMC, respectively. sstr subtypes display a distinct expression pattern on human colonic SMC; on circular SMC, subtype 2 is the only sstr, whereas sstr subtypes 1, 2 and 3 are expressed on human SMC isolated from the longitudinal layer. The contractile effects of SS are mediated through sstr subtype 2 and sstr subtype 1 on circular and longitudinal human colonic SMC, respectively.

Optimisation of isolation of richly pure and homogeneous primary human colonic smooth muscle cells.

BACKGROUND: Inherent properties of gastrointestinal smooth muscle can be assessed using isolated cell suspensions. Currently available isolation techniques, based on short 2-h enzymatic digestion, however, present the disadvantage of low cellular yield with brief viability. These features are an important limiting factor especially in studies in humans in which tissue may not be available daily and mixing of samples is not recommended. AIMS: To optimise the isolation procedure of cells from human colon to obtain a richly pure primary smooth muscle cell preparation. METHODS: Slices of circular muscle layer, obtained from surgical specimens of human colon, were incubated overnight in Dulbecco's modified eagle's medium supplemented with antibiotics, foetal bovine serum, an ATP-regenerating system and collagenase. On the following day, digested muscle strips were suspended in HEPES buffer, and spontaneously dissociated smooth muscle cells were harvested and used either immediately or maintained in suspension for up to 72 h. Cell yield, purity, viability, contractile responses, associated intracellular calcium signals and RNA and protein extraction were evaluated and compared to cell suspensions obtained with the current short digestion protocol. RESULTS AND CONCLUSION: The overnight isolation protocol offers the advantage of obtaining a pure, homogeneous, long-life viable cell suspension that maintains a fully differentiated smooth muscle phenotype unchanged for at least 72 h and that allows multiple functional/biochemical studies and efficient RNA extraction from a single human specimen.

Utility of combined use of plasma levels of chromogranin A and pancreatic polypeptide in the diagnosis of gastrointestinal and pancreatic endocrine tumors.

BACKGROUND: Chromogranin A (CgA) is considered the most accurate marker in the diagnosis of gastro-entero-pancreatic (GEP) endocrine tumors. Pancreatic polypeptide (PP) has also been proposed to play this role, but then not used due to its low sensitivity. The aim of the present study was to determine whether the assessment of PP would improve the diagnostic reliability of CgA in patients with GEP tumors. PATIENTS AND METHODS: Both markers were assessed in 68 patients [28 functioning (F), 40 non functioning (NF)]. Twenty-seven patients disease-free (DF) after surgery, and 24 with non-endocrine tumors (non-ETs) were used as control groups. RESULTS: CgA sensitivity was: 96% in F, 75% in NF, 74% in pancreatic, and 91% in gastrointestinal (GI) tumors. Specificity was 89% vs DF, and 63% vs non-ETs. PP sensitivity was: 54% in F, 57% in NF, 63% in pancreatic, and 53% in GI tumors. Specificity was 81% vs DF, and 67% vs non-ETs. By combining the two markers a significant gain in sensitivity vs CgA alone was obtained: overall in GEP tumors (96% vs 84%, p = 0.04), in NF (95% vs 75%, p = 0.02), and in pancreatic (94% vs 74%, p = 0.04). More specifically, a 25% gain of sensitivity was obtained in the subgroup of NF pancreatic tumors (93% vs 68%, p = 0.04). CONCLUSION: The combined assessment of PP and CgA leads to a significant increase in sensitivity in the diagnosis of GEP tumors, particularly in pancreatic NF.