In preparation for future analyses, intestine tissue biopsies were either (i) stored in RNAlater for subsequent analysis of mRNA expression (AM7021; Applied Biosystems, Carlsbad, CA, USA), (ii) fixed in neutral-buffered formalin for histological assessment, or (iii) immediately frozen in liquid nitrogen for proteomic analyses. Histological assessment was performed by a pathologist (EU) who was blinded to patient clinical and laboratory data, utilizing the histological grading scale introduced by Geboes et al.37 (link). Six histological features were evaluated as previously described12 (link). Due to the limited available biopsy material, distinct investigations were performed based on mRNA analysis. However key findings were validated at the protein level.
Liver tissue specimens were collected as tissue blocks (∼1 cm3), which were immediately frozen in liquid nitrogen and stored at −75 °C until use. For analysis, samples were powdered and homogenized in an appropriate lysis buffer to extract either total RNA or protein.
Liver tissue specimens were collected as tissue blocks (∼1 cm3), which were immediately frozen in liquid nitrogen and stored at −75 °C until use. For analysis, samples were powdered and homogenized in an appropriate lysis buffer to extract either total RNA or protein.
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