Expression of factors involved in bile acid metabolism throughout the gastrointestinal tract
Amendement: Expression of HIF-1alpha protein in upper GI tractus

Disruptions of the bile acid cycle causes an accumulation or a deficit of bile
acids, which may lead to a great variety of diseases of th egastrointestinal
tract, including cholestatic liver disease and Barrett's esophagus [1, 2]. In
this process nuclear receptors (NRs) play an essential role. They function as
bile acid sensors and influence genes including enzymes and transporters of the
bile acid cycle. Hereby, they change the composition of circulating bile acids
as well as the toxicity and function of bile acids [1, 3].
Molecular biologic techniques have made it possible to clarify the role and
function of NRs. Although the presence of these NRs in different parts of the
gastrointestinal tract has been studied in murine models, in humans little is
known. Recent studies in human have shown that NRs such as the Farnesoid X
receptor (FXR) and Pregnane X receptor (PXR) are associated with Barrett's
esophagus and inflammatory bowel disease [4, 5]. To understand the role and
function of NRs in the development of these and other disease of the
gastrointestinal tract, knowledge of NRs and the genes that they influence
that are involved in the regulation of bile acid metabolism in the entire human
gastrointestinal tract is necessary.


Amendement: Hypoxia inducible factor 1 alpha (HIF-1 alpha) is a transcriptional
factor which is induced by hypoxia. Increased concentrations of intracellular
HIF-1 alpha molecules occur after hypoxia as a result of reduced degradation by
the ubiquitin proteasome pathway. Binding of HIF-1 alpha to the hypoxia
response element of several *HIF regulated genes* results in the increased
transcription of several proteins involved in angiogenesis, glycolysis and
erythropoiesis.1 The presence of HIF-1 alpha has been demonstrated in the
surface epithelium of normal colon and adenomas and in areas associated with
peri-necrotic areas of colon adenocarcinomas.1 In addition, HIF-1 alpha
overexpression has been shown in colon of patients with ischemic colitis2,
ulcerative colitis and Crohn*s disease3. Rather, until now the expression of
HIF-1 alpha has not been investigated in normal tissue of upper
gastrointestinal tract.

To obtain insights into the diverse nuclear receptors involved in the
regulation of the bile acid cycle across the entire human gastrointestinal
tract.


Amendement:
The aim of our study is to investigate the expression and amount of expression
of HIF-1 alpha in normal tissue of upper gastrointestinal tract

For this study we obtain material at eight specified locations of the
gastrointestinal tract of patients (± 50), which is stored for the design of
several molecular as well as immunological experiments. Herefore, we use small
pieces of tissue (biopsy specimen) of patients, which are taken during standard
double balloon enteroscopy at the department of Gastroenterology of the Erasmus
MC University Medical Center Rotterdam. Formalin fixed biopsies that are taken
standard for diagnostics will be used. This paraffin fixed materiaal serves to
determine the location of the targeted bile acid factors using immunologic
techniques. Also, extra biopsies will be transferred in a special buffer,
encoded and stored in a -80°C freezer to, at a later moment, isolate RNA from
to determine the expression levels of several bile acid genes, ie nuclear
receptors.

Amendement:
Formalin fixed biopsies that are taken standard for diagnostics will be used.
This paraffin fixed materiaal serves to determine the location of HIF 1 alpha
using immunologic techniques.

In general, the duration for taking the extra biopsies will be no longer than 5
min (duration of a standard DBE is 90 min). The scopy itself is a very low risk
intervention (less than 1 in 3000 scopies leads to complications).