Unit:
Faculty of MedicineLibrary of the School of Health Sciences
Author:
Georgopoulos Ioannis
Dissertation committee:
Ιωάννης Παπακωνσταντίνου, Αναπληρωτής Καθηγητής, Ιατρική Σχολή, ΕΚΠΑ
Γεώργιος Κόλλιας, Καθηγητής, Ιατρική Σχολή, ΕΚΠΑ
Ιωάννης Βασιλείου, Καθηγητής, Ιατρική Σχολή, ΕΚΠΑ
Ντίνα Τηνιακού, Καθηγήτρια, Ιατρική Σχολή, ΕΚΠΑ
Γεώργιος Μπάμιας, Αναπληρωτής Καθηγητής, Ιατρικής Σχολή, ΕΚΠΑ
Αντώνιος Βεζάκης, Αναπληρωτής Καθηγητής, Ιατρικής Σχολή, ΕΚΠΑ
Γεώργιος Καραμανώλης, Αναπληρωτής Καθηγητής, Ιατρική Σχολή, ΕΚΠΑ
Original Title:
Η παρουσία στένωσης του εντέρου ως αναγκαίος συμπαράγοντας εκδήλωσης της νόσου Crohn σε έδαφος γενετικής προδιάθεσης. Πειραματική μελέτη σε μύες TnfΔare στην ηλικία της εφηβείας
Translated title:
Intestinal stenosis as an important co-factor on the expression of Crohn’s disease in subjects with genetic predisposition. Experimental study on TNFΔare mice in puberty
Summary:
Background
The pathophysiology of Crohn’s disease (CD) despite nearly a hundred years of modern research remains obscure. The widely accepted simplified concept of CD pathogenesis is that it results from abnormal responses of the intestinal immune system against the gut microflora in a genetically susceptible host. Clinical observations indicate that CD occurs or recurs proximal to narrow sites of the intestine, natural or iatrogenic. We advocate that site-specific factors, such as mechanical forces, contribute to the expression or recurrence of CD. To test that hypothesis, we investigated whether the creation of an intestinal stenosis could alter the severity of the expected Crohn-like ileitis, in a CD animal model, the TNFΔare/+ mouse.
Materials and Methods
For the evaluation of our hypothesis a reliable long term intestinal partial obstruction mouse model was needed in combination with a genetic mouse strain with genetic predisposition for Crohn-like inflammation.
Τhe mouse strain chosen for our study was TNFΔare/+ C57BL/6 due to the impressive resemblance of the Crohn-like disease presented on the animals, in pathologic, anatomic and molecular level, to the human CD.
Due to the absence of a robust long-term intestinal incomplete obstruction mouse model, we conducted a preliminary experiment to evaluate the best model of partial intestinal obstruction between the most widely used techniques in the literature and a novel technique that our team developed.
Sixty C57BL/6 mice aged 6 to 8 weeks were randomly divided into 5 major intervention groups: ligation, intestinal ring, partial ligation, microclips and the novel triple suture technique. The ring groups were subdivided into narrow, medium and wide ring and partial ligation groups were subdivided at 1/3, 1/2 and 2/3 of the lumen. The criteria of acceptance of a method were low mortality rates, long-term survival set at 4 weeks, macroscopic and microscopic evidence of obstruction, repeatability and reproducibility, low cost of materials and applicability to different animal sizes and species. Animals were euthanized after study period and studied macroscopically and the small bowel muscle layer thickness near the intervention was histopathologically evaluated. The “triple suture” group was the only group that met all criteria posed, with a survival rate of 90% at 4 weeks, the sutures were apparent in all cases, macroscopic evaluation showed small to mild proximal lumen dilatation in 6 out of 10 animals and histopathological evaluation of the specimens confirmed the partial obstruction. Thus, the triple suture technique was chosen for the protocol to test our core hypothesis.
For the evaluation of our core hypothesis, thirty-six, 6-weeks-old TNFΔare/+ C57BL/6 mice, were divided into 3 intervention groups: triple suture, single suture and sham. 3 cm from the ileocecal valve on the ileum, in the first group, a triple suture stenosis was created, whereas, in the second, a loose suture was placed. The third group was sham operated. Same triple-suture stenosis was performed on twelve C57BL/6 wild type (WT) mice. All animals were sacrificed at 6 weeks post-operatively and were biometrically and macroscopically studied. Subsequently, the ileum parts near the intervention were excised and evaluated histopathologically. A summative total ileitis score (TIS) was applied in each sample using a bespoke semiquantitative histological scoring system for the Crohn-like changes, constructed especially for our protocol.
Results
All mice gained weight and none developed signs of severe malaise or alterations in appetite and food consumption. The triple suture technique created an incomplete obstruction in all subjects as evaluated macroscopically and histopathologically by measuring muscle layer thickness of the specimens.
The triple suture stenosis induced significant muscular hypertrophy proximal to interventional site which was more prominent in TNFΔare/+ than wild type mice. In triple suture group, the total ileitis score (TIS) was significantly increased proximal to the intervention as compared to the single suture (P: 0.004) and the sham groups (P: 0.013). The TIS distally, was unaffected, regardless of the experimental intervention. Intestinal stenosis did not induce intestinal inflammation (CD-like or other) in WT mice.
Conclusions
Our research presents the first - to our knowledge - evidence that a surgically-induced long-term intestinal stenosis accelerates CD-like ileitis in inflammation-prone mice. Mechanical forces such as elevated intraluminal pressure prior to stenosis might be important co-factors to the pathophysiology of CD in genetically predisposed subjects.
Main subject category:
Health Sciences
Keywords:
Crohn's disease, Inflammatory bowel disease (IBD), Experimental animal model, Experimental intestinal stenosis, TNFΔare mouse model, Mechanical factors in Crohn's disease
Number of references:
251
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