Confocal Endomicroscopy Utility (p-CLE) in Irritable Bowel Syndrome
Condition: Irritable Bowel Syndrome (IBS)
MeSH terms: Irritable Bowel Syndrome; Syndrome
Study type: Observational

Summary: 
      Irritable bowel syndrome (IBS) is one of the most common gastrointestinal disorders. It has
      prevalence in general population of 5-20% and is more common in women and young adults.

      Despite being one of the most frequent reasons for consultation many patients are
      undiagnosed. There are no reliable biomarkers. The diagnosis is clinical, based on the Rome
      III criteria. IBS is characterized by chronic or recurrent abdominal pain associated with
      changes in bowel frequency and consistency, when other etiologies are excluded. The
      combination of the Rome III criteria with the absence of alarm symptoms have a sensitivity of
      65%, specificity of 100%, 100% positive predictive value and negative predictive value of
      76%.

      Current tests commonly fail to obtain an objective diagnosis, and effective therapies are
      lacking. There are no specific endoscopic findings that can discriminate IBS patients from
      healthy patients. Most colonoscopies are performed to rule out other etiologies and in more
      than 50% of the cases are normal.
    

Detailed description: 
      The pathophysiology is complex, multifactorial and not completely known. Impaired intestinal
      barrier function and mucosal inflammation in the small and large intestine have been
      reported. Evidence of inflammation and neuronal degeneration in the myenteric plexus has been
      found in full-thickness intestinal biopsy samples from IBS patients.

      It has been found that IBS patients had a significantly higher density of intestinal
      epithelial gaps. The elevated epithelial gaps in the intestine of IBS patients, a surrogate
      marker for increased epithelial cell extrusion, may be a cause of altered intestinal
      permeability observed and may lead to mucosal inflammation.

      Confocal laser endomicroscopy (CLE) is a new endoscopic imaging tool enabling visualization
      of changes in the gut mucosa. The use of intravenous fluorescein enables high-resolution
      real-time imaging in vivo at a micron scale, allowing visualization of cellular details
      during ongoing endoscopy. This technology can be used to identify and quantify epithelial
      gaps and other signs of inflammation.

      The purpose of this study is to evaluate the presence of microscopic inflammation previously
      demonstrated in patients with IBS, using pCLE. The determination of the exact areas of
      inflammation allows a direct biopsy in a more accurate way. This type of systematic may allow
      in the future to classify these patients not only symptomatically (constipation, diarrhea or
      mixed predominant IBS) but also according to their degree of inflammation. There is no Gold
      Standard test in these patients to assess response to treatment. The possibility of using
      pCLE to identify the areas of microscopic inflammation allows that the degree of inflammation
      can be used as a monitoring parameter. Probably the CLE can be used to assess response to
      treatment and different types of treatments, determining differences between subtypes of
      patients with IBS, evaluate remission, recurrence and prognosis. Also it may clarify a
      potential mechanism of the pathogenesis and be endoscopic criteria for the diagnosis of the
      disease.

      The present study is the first to evaluate the colon microscopic inflammation using pCLE in
      IBS patients.

      Study design: This is an observational and analytical cross-section, population - based
      survey study, with prospective case collection, non-randomized and simple blind, performed in
      a Tertiary Academic Center.

      Setting: Ecuadorian Institute of Digestive Diseases (IECED), OmniHospital Academic Tertiary
      Center. We will include patients from October 2015 to March 2016. Patients will be recruited
      from the gastroenterology unit (IECED). The study protocol and consent form has been approved
      by the Institutional Review Board (IRB) and will be conducted according to the declaration of
      Helsinki. Patients will sign an informed consent and answer a questionnaire that includes
      constitutional data (sex, age, comorbid medical condition, medication). Patients in IBS group
      will be classified according to the symptoms as diarrhea predominant (IBS-D), constipation
      predominant (IBS-C) or mixed type (IBS-M). Endoscopic findings will be recorded in a
      database.

      Endoscopic technique: All colonoscopies will be performed after the patient had undergone
      standard bowel preparation using either polyethylene glycol electrolyte lavage solution.
      Initially a routine white light colonoscopy will be performed in IBS and healthy control
      patients, using a Pentax scope (Pentax EC-387CILK, Tokyo, Japan) with great care not to
      damage the epithelium. The Boston Bowel Preparation Scale and inflammation changes will be
      recorded.

      During withdrawal, all parts of the colon will be evaluated (cecum, right-side colon,
      transverse, left-side colon, sigmoid and rectum) using pCLE randomly in one point of each
      segment, if there are no signs of iatrogenic mucosal damage. Careful washing of the mucosa
      with water in order to prevent the inclusion of image artefacts from residual stool
      fragments. Before initiating CLE evaluation, intravenous injection of 5 ml 10% fluorescein
      will be done. Fluorescein stains vessels and gives good tissue structure, but the nuclei are
      not stained and appear as dark spots. Optimal contrast is obtained within the first 10min
      after injection, but good quality images can be acquired for a further 30-60 min.

      Currently Confocal Laser Endomicroscopy (CLE) is performed using a CLE probe (pCLE). The
      confocal microscope used in pCLE captures microscopic images of untreated in vivo tissue. The
      microscope uses focused laser light of a defined wavelength and passes it through a confocal
      aperture. Images are then reconstructed in two dimensions. For pCLE (Cellvizio by Mauna Kea
      Technologies, Paris, France), both the laser scanning unit and light source are outside the
      body. The laser beam is transported via flexible confocal miniprobes and a distal lens
      sequentially scans it through a bundle of more than 10000 optical fibers. Confocal miniprobes
      are flexible, with diameters ranging from 0.9 mm to 2.5 mm.

      In the study a pCLE ColoFlex probe will be used (Cellvizio; Mauna Kea Technology, Paris,
      France). The probe goes through the accessory channel of a conventional endoscope. The image
      rate is 0.08 s/frame at 1024×1024 pixels with a imaging depth of 20 μm. The confocal images
      will be first analyzed during the endoscopy. Then, the images will be digitally stored and
      reviewed after the procedure in order to zoom in on details for a higher magnification
      (approximately 10000 fold), with special designed software package (Cellvizio Viewer),
      allowing image correction and stabilization.

      Immediately after pCLE evaluation, and before proceeding to the next colon segment, targeted
      biopsies will be taken from the mucosal area evaluated. Bleeding can impair image quality,
      therefore probe will be placed gently in contact with the tissue in order to avoid trauma and
      pCLE imaging will be completed in a region before mucosa biopsy. The endoscopist performing
      the pCLE will be blinded to the status of the patient and the images will be later analyzed
      in detail blindly by two different persons with more than two years' experience in
      endomicroscopy.

      There are not studies that validate a classification of colon inflammation using CLE in IBS
      patients. Inflammation of the colon mucosa in ulcerative colitis (UC) has been described
      previously using CLE. Kiesslich et al. reported on a 3-grade classification of inflammation
      activity by CLE, combining crypt architecture, cellular infiltrations, and vessel
      architecture. Nevertheless the assessment of cellular infiltration has been questioned
      because of the difficulty in differentiating cell types by CLE. Chang-Qing Li, et al. used
      three CLE parameters to evaluate inflammation in UC patients: a 4-grade crypt-architecture
      classification, fluorescein leakage, and microvascular alterations. On the other hand, the
      Miami classification for pCLE defines colitis using: crypt fusion and distortion, bright
      epithelium, dilated and prominent branching vessels.In the present study the parameters used
      for CLE inflammation will be: 1.Epithelial gaps followed by leaks with secretion of
      fluorescein into the lumen. In CLE images of normal colonic mucosa, the lumen of the crypt is
      free of fluorescein and appears as a dark center in the crypt; however, in inflamed mucosa,
      fluorescein leaks into the crypt lumen; therefore, the lumen is brighter than the surrounding
      epithelium. 2.4-grade crypt-architecture classification: Types A and B will be considered as
      normal and chronic inflammation, respectively, and types C and D indicate acute inflammation.
      Type A: Regular arrangement and size of crypts. Type B: Irregular arrangement of crypts,
      enlarged spaces between crypts. Type C: Dilation of crypt openings, more irregular
      arrangement of crypts, and enlarged spaces between crypts as compared to type B. Type D:
      Crypt destruction and / or crypt abscess. 3.Dilated and prominent branching vessels

      Interobserver and Intraobserver Agreement: A data set containing CLE photographs and videos
      of the all of the colon segments will be presented to three blinded endoscopists, who will
      confirm or not the findings. Inter- and intra-observer reproducibility will be measured based
      on comparison of still images and videos between the three investigators. To evaluate the
      intra-observer agreement each investigator will assess the images two times and the answers
      will be compared. To evaluate the inter-observer agreement all answers between the three
      investigators will be compared. In general, the learning curve of CLE is short and inter or
      intra-observer agreement is good.

      Statistical analysis: Baseline characteristics will be compared between case and control
      group using Chi-square o Fisher Test for categorical variable, and for continuing variables,
      we will use the Mann-Whitney Test. Diagnosis efficacy will be measured thought sensitive,
      specificity and accuracy. All the statistical analysis will be performed using SPSS software
      suite v.22.
    

Intervention name(s): Confocal Laser Endomicroscopy
Intervention type(s): Device
Intervention description(s): During colonoscopic withdrawal, the colonic mucosa will be observed with white light and be evaluated on the different segments of the colon with the confocal laser endomicroscopy technology in order to search for microscopic inflammation signs (altered crypt architecture, fluorescein leaks and dilated and prominent branching vessels)
Observational model: Case-Control
Time perspective: Cross-Sectional
Sampling method: Non-Probability Sample
Study population: 
        Outpatients with irritable bowel syndrome according to Roma III classification: recurrent
        abdominal pain or discomfort, at least 3 days per month, in the last 3 months and symptoms
        begin at least 6 months before diagnosis, associated with 2 or more of: 1) Improvement with
        defecation, 2) Start associated with changes in the bowel frequency, 3) Start associated
        with changes in stool consistency. In the absence of alarm symptoms. It will be measured in
        all patients serology for celiac disease, serum thyroid-stimulating hormone (TSH) and
        parasitological test. It will be done, in high suspicion patients, stool culture, breath
        test for lactose intolerance and breath test for bacterial overgrowth.
      
Biospecimen description: 
      Histological biopsies of the colonic mucosal
    
Keywords: Confocal Laser Endomicroscopy; inflammation; Irritable Bowel Syndrome