  Last updated 14.04.04 A.U. |
Head of section: Professor Knut-Jan Andersen , dr.philos.
Technical staff:
Aud Utheim,
Senior experimental officer
Behzad Gharehnia,
Senior experimental officer
Asta Sande Urtegaard,Senior
experimental officer
Brief history: The section was originally formed in 1972 when Knut-Jan
Andersen (biochemist) was appointed as Assistant Professor in Medical Department
A, to aid expertise in biochemical techniques for ongoing research in Gastroenterology,
Nephrology and Cardiology. Based on departemental leadership the ongoing research
was first focused on Gastroenterology (Vitamin B12 absorption; Enzyme activities
in normal and diseased intestinal mucosa). Later the focus was shifted to Nephrology
(Lysosomal function; Tubular Reabsorption and protein breakdown), although a
range of other major projects also were conducted including the detection and
determination of trace elements in biological fluids and tissues, and cytotoxicity
and mechanistic studies (phase 1 and 4) of x-ray contrast media. Parallel to
this, the section has continuously given advice and practical help with laboratory
assays and techniques to other investigators (including hospital doctors), although
this has not always been reflected in the list of publications.
Based on experience and collaboration, with both international and national
research groups, the main research activities in the section are today focused
on:
Research activities: Today the section has two major ongoing research projects both focusing on proteolytic activity in tumour tissues and cells.
Collaborators:
Associate
Professor Nils-Petter Aardal, MD., Ph.D., The Gade institute,
Dept. of Pathology ,UoB, Haukeland University Hospital.
Senior
Consultant Arild Horn, MD., Ph.D. Institute of Surgical Sciences, Division
of Gastroenterology ,UoB, Haukeland University Hospital.
The presence of increased activity for various proteolytic enzymes has been
demonstrated for most GI cancers, of great importance for both tumour growth
and invasion. Very limited information has been available on possible regional
differences, which enzymes are involved, or possible correlation of prognostic
criteria based on morphological observations and increased proteolytic enzyme
activity measured by biochemical assays.
We have so far studied tumours and normal tissue from more than 300 patients
with GI cancers (pancreas, stomach, colon, and rectum). Samples from tumour
and normal mucosa (10 cm from the tumour edge) are fixed for histological examination
at the Department
of Pathology. Tissue homogenates are assayed in our laboratory for cathepsin
B, cathepsin H, cathepsin L, dipeptidyl peptidase I (cathepsin C), acid -galactosidase,
acid -glucosidase, dipeptidyl peptidase II, tripeptidyl peptidase I, acid methyl
amino peptidase, N-acetyl- -glucosaminidase, and acid phosphatase, (lysosomal),
alkaline phosphatase, alanyl amino peptidase, dipeptidyl peptidase IV, -glutamyl
transferase, leucine amino peptidase, neutral methyl amino peptidase, neutral-
-glucosidase, and tripeptidyl peptidase II (plasma membrane/brush border), mono
amino oxidase (mitochondria), and tryptase (mast cells) in addition to DNA and
protein.
A preliminary examination of the data revealed that 12 enzymes showed increased
activity in stomach cancers, 9 in colon cancers, and 13 in rectum cancers. Increased
activity of several of the proteolytic enzymes measured was also found to correlate
with morphological observations classified according to Jass with respect to
unfavorable prognosis. A complete statistical examination, combining morphological
observations and biochemical data, in order to validate the use of biochemical
marker enzymes as a tool for assessing the prognosis in gastrointestinal will
be performed. Also, due to the time span of this project (10 years), permission
has been given by The Norwegian Data
Inspectorate to examine for a possible correlation between survival
and the experimental data obtained.
As a part of this study, samples of 100 l of extracts from both cancer and normal
tissue from some patients were also analyzed by gelatine substrate zymography
(11% gels) to look for possible increased Matrix Metalloproteinase activity.
The zymogram techniques revealed the presence of several bands of gelantinolytic
activity in tumour tissues from colon, rectum, and pancreas. This activity was
not detected in normal tissue from the same patients, and might therefore be
of specific tumour origin. When tissue samples were preincubated with selected
proteinase inhibitors prior to analysis, the gelantinolytic activity was inhibited
100%. These pilot observations led to Bioprospecting; - the search for biochemical
and genetic resources in plants, animals and microorganisms, to find possible
anti-tumour agents.
Collaborators (International):
Professor Erik Ilsø
Christensen, MD., Ph.D., Department of CellBiology, Institute of Anatomy,
University of Aarhus, Denmark, on correlating cell ultrastructure and function
using electron microscopy in combination with physiological experiments, cell
fractionation, cytochemistry, immuneohistochemistry, immunoelectron microscopy
and autoradiography.
Professor Eremitus J. Ken. McDonald,
Ph.D., Department of Biochemistry and Molecular Biology, Medical University
of South Carolina , Charleston, SC, USA, on proteases (endo- and exo-peptidases),
with emphasis on collagen degrading proteases.
Dr. Miloslav
Dobrota, Ph.D., Senior advisory consultant, School of Biomedical and
Life Sciences
Matrix metalloproteinases (MMP) are a family of membrane-bound proteases able
to degrade all components of the extracellular matrix. There is considerable
evidence that the MMPs have a major role in diverse physiologic and pathologic
processes, including aspects of embryonic development, tissue morphogenesis,
wound repair, inflammatory diseases, and cancer. An imbalance between MMPs and
associated Tissue Inhibitors of Metalloproteinases (TIMPs) may play a significant
role in the invasive phenotype of malignant tumours. Numerous studies have shown
that there is a close relation between the expression of various members of
the MMP family by tumours and their proliferative and invasive behavior and
metastatic potential.
The role proteases play in cancer cells growth and spreading has been the focus
for the research in our laboratory for many years. Due to pilot observations
where Zymogram techniques revealed the presence of several cancer-specific MMPs
not detected in normal tissue from the same patient (control), and the inhibition
of activity caused by certain flavonoids, our interest has focused on possible
natural sources of protease inhibitors that can effectively inhibit MMP activity
and thereby having an inhibitory effect on the growth and spreading of cancer
cells.
Several Tissue Culture Model Systems have been established to study effects
on cell proliferation (DNA synthesis), tumour cell invasion, three-dimensional
growth of tumour cells, confrontation of tumour and normal cell spheroids, and
apoptosis. In addition, zymogram techniques are used to examine possible inhibition
of MMP activity in human tumour tissues. Using these techniques very promising
results have been obtained on certain flavonoids, naturally occurring compounds
in most fruits and berries.
Our studies have been expanded to include certain extracts of different types
of marine microalgae and cyanobacteria and have been found to inhibit tumour
cell proliferation (DNA synthesis), tumour invasion, and three-dimensional growth
of tumour spheroids. Organic and water extracts of different types of marine
microalgae and cyanobacteria also demonstrated the presence of high MMP activity
(mainly water extracts) in some species while others showed MMP inhibition (mainly
organic extracts).
Based on these results, a broad national and international collaboration has
been established to test for potential inhibitory activity on MMP activity and
tumour cell growth and behavior. Furthermore, based on previous experience through
collaborative on cellular toxicity of x-ray contrast media, pre-clinical testing
(Phase 1 studies) of the most potent compounds and extracts are performed in
addition to mechanistic studies in tissue culture of normal and cancer cells.
The main goal of this project is to identify, isolate and characterize, biochemical and chemical compounds in plants, animals and microorganisms, with a focus on protease inhibitors; and to understand and acquire information on mechanisms acting, in order to find potential new drugs for the future.
Also, to collaborate with other research groups and provide an experimental tool for cytotoxicity and pre-clinical testing of active bio components from different courses.
And, to teach students the importance of Bioprospecting when doing their Master and Ph.D. as part of this project.
Kimberley Hatfield: Possible inhibitory compounds of matrix metalloproteases
extracted from marine cyanobacteria and microalgae.
Asta Urtegaard: Effect of an algae of the Chlorella-type on cancer
cells.
Anne Gjeitle: Studies extracts from the Diatome algae and their
effect on proteolytic enzymes from gastrointestinal tumours.
David Kamal: Purification, characterization and comparison of
matrix degrading enzymes from gastrointestinal tumours.
Inderjit Daphu: Studies on the effect of resveratrol on tumour
cells and matrix metalloproteinase activity.
Stig Bruhjell: Production of protease inhibitors in algae during exponential
stationary growth.
Anette Matland: Protease and protease inhibitor interaction in the Diatome
algae.
At present there is a waiting list of students that want to do their M.Sc.
thesis in connection with the project on bioprospecting. The limiting factors
being the laboratory space available, and the capacity to supervise more than
5 students at the time.
The projects within our section have received funding from several minor grants
in addition to support from the Norwegian cancer society "Støtteforeningen
for Kreftrammede".
We are at present funded by major grant from The Research Council of Norway.
Address: Research Laboratory A, Institute of Medicine,UiB,
Haukeland University Hospital, N-5021 Bergen
Visiting address: Main Hospital Building, Room A215
Phone: (+47)55 973052 Fax: (+47) 55
972950
E-mail: Knut-Jan.Andersen@med.uib.no
Last updated 14.04.04 A.U. |