The Study On Duodenal Immune Activation And Alteration Of Neurotransmitters In Patients With Functional Dyspepsia

Background and ObjectivesFunctional dyspepsia (FD) is a functional syndrome thought to originate in the gastroduodenal region, the symptoms of FD is described as epigastric pain, epigastric burning, postprandial fullness and early satiation, in the absence of any organic, systemic, or metabolic disease that is likely to explain the symptoms. FD was divided into two subgroups, which were postprandial distress syndrome (PDS) and epigastric pain syndrome (EPS). As one of the most prevalent functional gastrointestinal disorders (FGIDs), the prevalence rate of FD has been noted to vary between11.0%-29.2%globally, thus induced a major impaired health-related quality of life and social and economic burden.So far, the etiology and pathogenesis of FD remain such obscure that there are some difficulties in prevention and cure of FD. Therefore, a further understanding of the nature of FD and a thorough study on its pathogenesis can guide clinical diagnosis and treatment, and has important social significance and scientific value.In FD, the duodenum is now implicated as a key area where symptoms originate. Post-infectious functional dyspepsia (PI-FD) is associated with persisting focal T-cell aggregates, decreased CD4+cells and increased macrophage counts surrounding the crypts of duodenum. As immune cells of gastrointestinal mucosa, eosinophils and mast cells play important role in regulating gastrointestinal function. Degranulated eosinophils can lead to nerve stimulation and smooth muscle contraction, resulting in gastrointestinal symptoms, such as abdominal pain and distension. Mast cells can release a variety of mediators such as5-HT (5-HT), nitric oxide (NO), vasoactive intestinal peptide (VIP) which can regulate gastrointestinal motility and visceral sensation. But up to now, only few foreign studies concerned the change of duodenal immune cells in patients with FD.Studies have shown that the incidence of food allergy was high in FD. Patients with FD were often accompanied by anxiety, depression, neuroticism, chronic tension, hostility, hypochondriasis psychology, pessimism and abnormal medical behavior. Existing opinion thinks that various stress factors can act on the stress-response system of brain, then act on the gastrointestinal target organ by the bidirectional regulation of brain-gut axis, finally induce the changes of the gastrointestinal motor, sensory, secretion and the immune function and lead to FD. This procedure may be associated with immune mechanism, but the role of duodenal eosinophils and mast cells was not clear.In the neuro-immuno-endocrine network of FGIDs, the enteric nervous system (ENS) is the important link. Various neurotransmitters released by enteric nerves play important role on regulating gastrointestinal motility and sensory. A large number of studies show that, nitric oxide (NO) and vasoactive intestinal peptide (VIP) is major inhibitory neurotransmitter in the enteric nervous system. In some disease, increased NO and VIP in muscle layer ENS can lead to attenuated gastric antral contractions and delayed gastric emptying. In human intestine, there are NO produced by enzymes and bacteria, two kinds of structural calcium-dependent NO synthase isoforms are responsible for a small amount of NO production:neuronal NOS (nNOS, Type Ⅰ) and endothelial NO synthase (eNOS and Type Ⅲ). The induction of inducible NOS (iNOS) tends to occur in the state of inflammation or immune activation. Inflammation, epithelial, vascular endothelial and nerve cells in the intestinal tract can express iNOS. Serotonin (5-HT) and substance P (SP) have exciting effect on gastric and duodenal motility. Neurotransmitter dysregulation of ENS may be a mechanism of FD, then, wether patients with FD have duodenal immune activation and abnormalities of duodenal iNOS and neurotransmitter?This study is aimed to understand the change of duodenal mucosa immune cells, explore the correlation of food allergy, psychosocial factors with duodenal immune cells, and find out the duodenal neurotransmitter dysregulation of ENS in patients with FD and its correlation with FD symptoms, duodenum immune cells, in order to clarify etiopathological role of duodenal neuro-immuno-endocrine network on FD.Methods1. Duodenal biopsies obtained by upper endoscopy in48patients with functional dyspepsia(23PDS and25EPS) and21healthy volunteers, eosinophils of the bulb (D1) and descending part (D2) of duodenum were identified and counted by H-E staining, major basic protein (MBP) immunostaining for eosinophil degranulation, mast cells and mast cells degranulation by toluidine blue staining.2. Peripheral venous blood was collected from48patients with functional dyspepsia (23PDS and25EPS) and21healthy volunteers, enzyme-linked immunosorbent assay was performed to detect14kinds of serum food allergen-specific IgG.3. Hospital anxiety depression scale (HADs) was used to evaluate the psychosocial status of48FD patients (23PDS and25EPS) and21healthy volunteers, the scores of anxiety (HADs-A) and depression (HADs-D) were given.4. The relationship among the species and positive scores of serum food allergen-specific IgG,HADs-A and HADs-D and the counts and the degranulation rates of MC or the counts of eosinophils in duodenum and FD symptoms scores, the species and positive scores of serum food allergen-specific IgG,HADs-A and HADs-D and the counts and the degranulation rates of MC or the counts of eosinophils in duodenum were analyzed by a linear correlation analysis.5. Expression of duodenal neurotransmitter including serotonin (5-HT), substance P (SP),nitric oxide (NO) and vasoactive intestinal peptide (VIP) were detected by immunohistochemistry. The relationship among neurotransmitter and FD symptoms scores, duodenal immune cells were analyzed. Results1. Compared to controls, eosinophils were significantly increased in FD and PDS and EPS in D2(23.23±5.18;22.65±5.81;23.76±4.58)(P<0.05), the difference of expression of MBP in D1and D2in FD between FD, PDS, EPS and controls were not significant (P>0.05), eosinophil degranulation were significantly increased in FD and PDS and EPS in D2(26/48;12/23;14/25)(P<0.05); There were not correlation between eosinophils counts in D1and D2and FD symptoms scores, D2MBP immunohistochemistry scores were positively correlated to epigastric burning symptom scores (r=0.528, P=0.024).2. Mast cells were significantly increased in FD and PDS and EPS in D1(120.94±13.31;121.00±13.75;120.88±13.13)(P<0.01) and D2(123.28±13.40;124.32±16.53;122.24±9.54)(P<0.01), mast cells degranulation rates (%) were significantly increased in FD and PDS and EPS in D1(60.23±5.10;59.84±4.50;60.58±5.66)(P<0.001) and D2(66.97±5.30;66.63±5.37;67.28±5.32)(P<0.01); Mast cell counts and degranulation rates were not correlated to FD symptoms scores (P>0.05).3. In PDS, positive rate of specific IgG of beef, chicken, crab, shrimp, wheat were significantly higher than control (beef:15/23, P<0.001; chicken:17/23, P <0.001; crab:20/23, P<0.001; shrimp:10/23, P=0.009; wheat:8/23, P=0.036); In EPS, positive rate of specific IgG of beef, chicken, soy were significantly higher than control (beef:14/25, P=0.003; chicken:14/25, P<0.001; soybean:18/25, P<0.001). Compared to controls, the scores of serum food allergen-specific IgG were significantly increased in FD, PDS and EPS (6.69±4.73,6.96±5.09,6.43±4.46; P<0.001); The species of serum food allergen-specific IgG were significantly increased in FD, PDS and EPS (3.89±2.62,4.19±2.83,3.60±2.42; P<0.01).4. The scores and species of serum food allergen-specific IgG were not positively correlated with FD symptoms scores(P>0.05); The scores of serum food allergen-specific IgG were positively correlated with the counts (r=0.247,P=0.038) and degranulation rates (r=0.307,P=0.011) of mast cells in D1; The species of serum food allergen-specific IgG were positively correlated with the counts (r=0.243, P=0.041) and degranulation rates (r=0.326, P=0.007) of mast cells in D1.5. Compared to controls, the scores of HADs-A (FD:6.92±5.51, P=0.001; PDS:7.00±5.46, P=0.032; EPS:7.35±5.41, P=0.018) and HADs-D (FD:7.19±5.12, P=0.005:PDS:7.31±5.28, P=0.023; EPS:7.08±5.05, P=0.035) in PDS and EPS patients were significantly higher. HADS-A was positively correlated to epigastric distention, early satiety, epigastric burning symptom scores (P<0.05), HADS-D was positively correlated to bloating, early satiety symptom scores (P<0.05).6. The scores of HADS-A were positively correlated with the counts of mast cells D1:r=0.843, P=0.000; D2:r=0.612, P=0.002) and degranulation rates (D1: r=0.714,P=0.000; D2:r=0.734, P=0.000) of mast cells in Dl and D2.7. Compared to controls, expression of iNOS was significantly enhanced in PDS in D1(P<0.05), in D2, the difference of expression of iNOS between PDS and controls was not significant (P>0.05), in D1and D2, the difference of expression of iNOS between EPS and controls was not significant (P>0.05); In Dl and D2, the difference of expression of5-HT between PDS or EPS and controls was not significant (P>0.05); In D1and D2, the difference of expression of SP between PDS or EPS and controls was not significant (P>0.05); In D1and D2, the difference of expression of VIP between PDS or EPS and controls was not significant (P>0.05).8. Expression of iNOS in D1in patients with FD was positively correlated to epigastric distention scores (r=0.496, P=0.003) and D1mast cell degranulation rates (r=0.321, P=0.008)Conclusions1. There was duodenal immune activition in FD, duodenal eosinophils and mast cells may be involved in the pathogenesis of FD.2. FD patients showed food allergy, serum food allergen-specific IgG may be involved in the pathogenesis of FD.3. FD patients showed the abnormal psychosocial status.4. Food allergy, anxiety and depression may be the initiating factors in the duodenal immune activation.5. iNOS of duodenum bulb may be involved in the pathogenesis of PDS. 6. Enhanced expression of iNOS in D1in FD patients may be associated with mast cell activation.The significance of studyThis study explored the change of duodenal immune cells, found out the correlation among food allergy, psychosocial factors and duodenal eosinophils and mast cells, approached duodenal neurotransmitter dysregulation of ENS in patients with FD, clarified the role of duodenal immune activation on FD. The results provide evidence for diagnose and new therapy of FD, it has important theoretical value and social significance.