Alkaline Phosphatase Positive Reticular Cell Network Recovery after Radiation-induced Marrow Aplasia in Mice

s of the papers presented during the meeting of January 26, 2002, Université Libre de Bruxelles, Hôpital Erasme 95 ABSTRACTS OF THE PAPERS PRESENTED DURING THE MEETING OF JANUARY 26, 2002, UNIVERSITÉ LIBRE DE BRUXELLES, HÔPITAL ERASME Acta Clinica Belgica, 2002; 57-2 BOLD CONTRAST WITHIN TUMORS : HOW DOES IT CORRELATE TO PO 2 ? SIMULTANEOUS AND CO-LOCALIZED MEASUREMENT USING GRADIENTECHO FMRI AND OXYLITE-OXYFLO Christine Baudelet and Bernard Gallez Laboratory of Biomedical Magnetic Resonance and Laboratory of Medicinal Chemistry and Radiopharmacy, Université Catholique de Louvain, B-1200 Brussels, Belgium. BOLD imaging is often proposed as a tool to evaluate the responsiveness of tumors to oxygen changes. However, the signal is combination of flow and oxygen dependant parameters. Up to-now, the validity of the approach relies only on the comparison of BOLD signal with oxygen measurements carried out on different animals or animals measured at different times and location. We have developed an experimental setup which allows a simultaneous and co-localized measurement of pO2/flow and gradient echo fMRI. Isoflurane anesthetized mice were imaged at 4.7 Tesla. The slice for imaging was carefully selected such that it was positioned on the extremity of the Oxylite-/Oxyflo probe inserted into the intramuscularly implanted tumor. The spatial resolution for imaging was chosen to get voxels dimensions compatible with the sensitive area of the probe, while keeping a good BOLD contrast. GE images (200/20/45°) were continuously acquired during modification of gas breathing (air-50%O2-air-100%O2-air-Carbogen-air). A set of multi-echo GE images (200/ 6.1/6/45°) were then also acquired following the same protocol. From Multi-echo images, both T2* map and S 0 map were created. We computed (pixel-by-pixel analysis) the correlation of the three time series with the signals coming from OxyLite and OxyFlo. A typical co-localized and simultaneous recording of the GE signal and tumor pO 2 : EXPRESSION OF VEGF AND ITS RECEPTORS IN HUMAN ILIAC CREST AND FEMORAL BONE MARROW Z. Belaid, A. Thiry, A. Rodriguez, J. Boniver, Ch.M.Lapière, BV. Nusgens and MP. Defresne, Departement of Histology and Anatomy pathology, Departement of Orthopedics Surgery, Laboratory of Connectives Tissues Biology, University of liege, Sart-Tilman, Belgium The understanding of the mechanisms regulating hematopoiesis in normal bone marrow is required for progressing in the elucidation of the alteration occurring in many hematological malignancies such as myelodysplastic syndromes and leukemias. The aim of this study is to further explore the hypothesis that normal human hematopoiesis may be regulated by paracrine regulatory loops and by cross-talk between hematopoietic cells and their environment. The vascular network is the first candidate, which can be implicated, in these regulatory loops. We investigated by RT-PCR the mRNA level of the vascular endothelial cell growth factor (VEGF), its splice variants and its receptors, the VEGR-1/Flt1, the VEGFR-2/KDR and the specific VEGF165 isoform co-receptor, the neuropilin-1 (NRP-1) as well as its ligand semaphorin-III (SEM-III) known to function as VEGF antagonist. Bone marrows were collected from the iliac crest and the femur from 11 donors. VEGF121 and 165 are the main isoforms expressed at variable levels in iliac crest and femur while the extracellular matrix binding-VEGF189 is present in all samples, although at low level. Neither the 145 nor the 206 variants are detected. The VEGFR-2 is similarly expressed in all samples. Interestingly we observed a higher steady state level of VEGFR-1, NRP-1 and SEM-III in the femoral bone marrow, rich in adipocytes than in the hematopoietic iliac crest. To assess the cellular origin of this overexpression in femoral bone marrow, adipocytes were isolated from collagenase-digested femoral biopsies on the basis of their floating properties. The purified adipocyte population effectively expressed more NRP-1 mRNA than the sedimented cells population (stromal cells and hematopoietic cells). These results will be confirmed by immunostaining and western-blot analysis at the protein level. In conclusion, this preliminary study showed that the expression of NRP-1 in adipocytes is high when hematopoiesis is low, suggesting that these cells may play an active role in regulation of hematopoiesis and angiogenesis. In conclusion, BOLD contrast imaging is a powerful tool to appreciate the efficacy of a treatment of O 2 breathing. In carbogen breathing protocols, the variation in signal intensity is higher than using the oxygen protocol; this effect is likely to be due to the combination of an inflow effect together (increase in So) with a higher increase in pO 2 (increase in T2*). BOLD MRI is a non invasive method that allows a mapping of the response, and the variations observed are directly related to pO 2 changes. However, this technique has no value to estimate the basal pO 2 before treatment and BOLD MRI should be considered as a technique complementary to other methods allowing absolute quantification of the pO 2 . QUANTIFICATION OF MICROMETASTATIC DISEASE IN BREAST CANCER PATIENTS : OPTIMALISATION OF THE METHOD USING REAL-TIME QUANTITATIVE PCR FOR THE DETECTION OF CK19 MRNA IN BLOOD I. Benoy, H. Elst, P. Van Dam, P. Huget, S. Scharpé, E. Van Marck, L. Dirix, Oncology Center, AZ Sint-Augustinus, Antwerp, Belgium Department of Pathology, University Hospital Antwerp, Belgium Department of Clinical Biochemistry, University of Antwerp, Belgium Introduction : Earlier diagnosis has led to stage migration and better prognosis, but a subgroup of patients with early breast cancer continue to succumb to metastatic disease. Nodal status remains the most valuable marker to predict for the presence of occult spread. Morphological and molecular techniques suffer from low sensitivity and a lack of quantification. We have developed a sensitive and quantitative assay to detect breast cancer cells in venous blood using a quantitative RT-PCR identifying transcripts of the CK19 gene Methods : Total RNA was isolated from full blood samples of patients and healtzy volunteers using the Qiagen Rneasy ® total RNA kit. For generation of first strand cDNA, 2μg of total RNA was reverse transcripted with the high-Capacity cDNA Archive Kit (Applied Biosystems). A cDNA-specific CK19 Taqman TM primer and probe set was developed using Primer Express ® software. PCR was performed on the ABI Prism 7700 system and the standard thermal cycling protocol. The cycle at witch the emission intensity rises above baseline is referred to as cycle