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Intratumoral lymphangiogenesis and VEGF-C expression in pancreatic neuroendocrine tumors
Posters: Pancreasand SalivaryGlands/Pathology -Research and Practice 200 (2004) 317-320 fected by electroporation. The vectors (Clontech) studied are: pEGFP-actin (green), pEYFP-tubulin (yellow), pECFP-nucleus (cyan). Fluorescing cells were observed 24-72 hours post-transfection by inverted fluorescence microscopy (IX70, Olympus) and visualised by a high-sensitive digital imaging system (TILL Photonics). Results: Acinar cells were transfected with the three tested Living Colors Subcellular Localisation Vectors (LCSLV) without loss of cell functions, but mostly with low efficiency. The highest expression rates (1,2%) were detected after 24 hours following electroporation. Green fluorescent pEGFP-actin or yellow-shifted pEYFPtubulin were integrated into the filamentous and microtubular systems of the cellular cytoskeleton. Co-transfection with pECFP-nucleus and pEGFP-actin or pEYFP-tubulin resulted in dual labeling of nuclei and cytoskeleton in some cells. Spatial and temporal changes of fluorescence labeled actin and tubulin structures have been observed in real time after stimulation of the transfected cells with the secretagogue caerulein (10 nM). Conclusions: LCSLV are useful tools for studying structures and cellular events in pancreatic acinar cells without fixation or chemical staining. This innovative method will help to answer some current questions of onset of pancreatitis on the subcellular level.
240 Intratumoral lymphangiogenesis and VEGF-C expression in pancreatic neuroendocrine tumors B. SIPOS 1, W. KLAPPER 1, M.L. KRUSE 2, D. KERJASCHKP, H. KALTHOFF3, G. KLOPPEL / 1Institut ftir Allgemeine Pathologie 2 1. Medizinische Klinik 3Molekulare Onkologie, Klinik fOx Chirurgie, Universit~itskl. Schleswig-Holstein, Campus Kiel 4 Institut fox Pathologie, AKH, Wien
Aims: The role of lymphangiogenic cytokines and intratumoral lymphangiogenesis in most human malignant neoplasms is unknown. We investigated the expression of lymphangiogenic factors and intratumoral lymphangiogenesis in pancreatic neuroendocrine tumors (NETs) and correlated these data with clinico-pathological parameters. Methods: 111 pancreatic NETs were investigated by immunohistochemistry for VEGF, bFGF and VEGF-C expression. VEGF-C and VEGF-D mRNA was quantified with real-time RT-PCR. Lymph vessels (LV) were recognized by anti-podoplanin antiserum. Intratumoral lymph vessel density (LVD) was assessed with digitalized imaging in 10 disease-free pancreata and in 111 NETs. Double labeling for podoplanin and PCNA was used for detection of proliferating LVs. Results: Moderate-to-strong VEGF-C expression was encountered in 25 of 111 NETs and its expression was associated with glucagon and PP expression, with high LVD and with a malignant phenotype of functioning NETs. VEGF-D was not detectable. NETs exhibited higher LVD than disease-free pancreata, but LVD did not discriminate between benign and malignant NETs. LVD was associated with VEGF-C expression but was independent of VEGF and bFGF expression. In NETs with high LVD, proliferating LV were identified. Malignant NETs showing high LVD more frequently exhibited LV invasion, however LVD was not associated with lymph node metastasis. Conclusion: Pancreatic NETs express VEGF-C associated with glucagon and PP expression. NETs reveal definite intratumoral lymphangiogenesis mediated partly by VEGF-C. Apart from the promoting effect on LV invasion, intratumoral lymphangio-genesis has little effect on the malignant progression of NETs.
319
241 Immune-mediated experimental chronic pancreatitis in rats J. EMMRICH, K. RIECKEHEER, G. SPARMANN, H. D. KLEINE I, R. JASTER, S. LIEBE Abteilung ftir Gastroenterologie Abteilung fox H~imatologie, Klinik fox Innere Medizin, Universit~it Rostock
Aims: There is evidence that immunological mechanisms are involved in the pathogenesis of chronic pancreatitis (CP). To investigate the role of lymphocytes in CP we transfered lymphocytes from animals with CP in animals with mild acute pancreatitis (AP) Methods: AP was induced by i.v. application of 4 mg/kg b.w. dibutyltin dichloride (DBTC) whereas for CP 8 mg/kg b.w. were given. On day 28 of the CP course, lymphocytes were obtained from blood, lymph nodes and spleen and were injected into rats 4 days after induction of AP. 2, 3, and 4 weeks after lymphocyte transfer, pancreata were examined histologically as well as immunohistochemically for lymphocytes, macrophages, and neutrophils using the APAAP technique. Expression of IL- 1~, IL-2, IL6, IL-10, and IFN,/was analysed with RT-PCR. Results: In DBTC-induced AP there was no inflammatory reaction after two weeks. In contrast, in animals with AP and lymphocyte transfer, focal periductal fibrosis with lymphocyte infiltration has been developed, followed by panreatic duct dilatation. Pancreatic tissue was infiltrated with macrophages and lymphocytes. The expression of the investigated cytokines was elevated after lymphocyte transfer. Conclusions: The results suggest a specific immunologicalreaction of lymphocytes against pancreatic tissue during the course of CP.
242 Prognostic relevance of histomorphologic factors and adjuvant treatment in pancreatic carcinoma G. ASSMANN ~, A. NOLTE 2, R. WILKOWSKI2, J. DIEBOLD 1, U. LOHRS 1 1Pathologisches Institut der Universitat Mtinchen 2Klinik und Poliklinik ftir Strahlentherapie der Universitgt Mtinchen
Aims: Pancreatic carcinoma is a frequent malignant tumour in industrial countries. The impact of histomorphological factors and type of adjuvant therapy on patient outcome has not been clearly elucidated so far. Methods: Retrospective analysis of 57 patients with curatively resected carcinoma of the head of pancreas, which had undergone additionally either radiotherapy (RTX), simultaneous radiochemotherapy (sRCTX) or simultaneous-sequential radiochemotherapy (ssqRCTX). Mostly stage pT3 (UICC 2002). Grading according to the WHO guidelines comprising glandular differentiation, mucine production, mitotic count and nuclear morphology. In addition assessment of perineural invasion, presence of lymphangiosis or hemangiosis and the status of the tissue margins in the specimens. To date univariate analyses of the different factors in relation to survival (multivariate analysis in prep.). Results: Patients with ssqRCTX showed a highly significantlybetter long-time survival compared to patients with RTX or sRCTX, respectively (p = 0.0038; p = 0.0039). In the ssqRCTX-group a tumour-free tissue margin was associated with a significantly better long-time outcome. Low mitotic count, mild nuclear polymorphism and absence of perineural invasion distant from the turnout seemed to have positive effects on survival (not significant due to small number of patients).
240 Intratumoral lymphangiogenesis and VEGF-C expression in pancreatic neuroendocrine tumors B. SIPOS 1, W. KLAPPER 1, M.L. KRUSE 2, D. KERJASCHKP, H. KALTHOFF3, G. KLOPPEL / 1Institut ftir Allgemeine Pathologie 2 1. Medizinische Klinik 3Molekulare Onkologie, Klinik fOx Chirurgie, Universit~itskl. Schleswig-Holstein, Campus Kiel 4 Institut fox Pathologie, AKH, Wien
Aims: The role of lymphangiogenic cytokines and intratumoral lymphangiogenesis in most human malignant neoplasms is unknown. We investigated the expression of lymphangiogenic factors and intratumoral lymphangiogenesis in pancreatic neuroendocrine tumors (NETs) and correlated these data with clinico-pathological parameters. Methods: 111 pancreatic NETs were investigated by immunohistochemistry for VEGF, bFGF and VEGF-C expression. VEGF-C and VEGF-D mRNA was quantified with real-time RT-PCR. Lymph vessels (LV) were recognized by anti-podoplanin antiserum. Intratumoral lymph vessel density (LVD) was assessed with digitalized imaging in 10 disease-free pancreata and in 111 NETs. Double labeling for podoplanin and PCNA was used for detection of proliferating LVs. Results: Moderate-to-strong VEGF-C expression was encountered in 25 of 111 NETs and its expression was associated with glucagon and PP expression, with high LVD and with a malignant phenotype of functioning NETs. VEGF-D was not detectable. NETs exhibited higher LVD than disease-free pancreata, but LVD did not discriminate between benign and malignant NETs. LVD was associated with VEGF-C expression but was independent of VEGF and bFGF expression. In NETs with high LVD, proliferating LV were identified. Malignant NETs showing high LVD more frequently exhibited LV invasion, however LVD was not associated with lymph node metastasis. Conclusion: Pancreatic NETs express VEGF-C associated with glucagon and PP expression. NETs reveal definite intratumoral lymphangiogenesis mediated partly by VEGF-C. Apart from the promoting effect on LV invasion, intratumoral lymphangio-genesis has little effect on the malignant progression of NETs.
319
241 Immune-mediated experimental chronic pancreatitis in rats J. EMMRICH, K. RIECKEHEER, G. SPARMANN, H. D. KLEINE I, R. JASTER, S. LIEBE Abteilung ftir Gastroenterologie Abteilung fox H~imatologie, Klinik fox Innere Medizin, Universit~it Rostock
Aims: There is evidence that immunological mechanisms are involved in the pathogenesis of chronic pancreatitis (CP). To investigate the role of lymphocytes in CP we transfered lymphocytes from animals with CP in animals with mild acute pancreatitis (AP) Methods: AP was induced by i.v. application of 4 mg/kg b.w. dibutyltin dichloride (DBTC) whereas for CP 8 mg/kg b.w. were given. On day 28 of the CP course, lymphocytes were obtained from blood, lymph nodes and spleen and were injected into rats 4 days after induction of AP. 2, 3, and 4 weeks after lymphocyte transfer, pancreata were examined histologically as well as immunohistochemically for lymphocytes, macrophages, and neutrophils using the APAAP technique. Expression of IL- 1~, IL-2, IL6, IL-10, and IFN,/was analysed with RT-PCR. Results: In DBTC-induced AP there was no inflammatory reaction after two weeks. In contrast, in animals with AP and lymphocyte transfer, focal periductal fibrosis with lymphocyte infiltration has been developed, followed by panreatic duct dilatation. Pancreatic tissue was infiltrated with macrophages and lymphocytes. The expression of the investigated cytokines was elevated after lymphocyte transfer. Conclusions: The results suggest a specific immunologicalreaction of lymphocytes against pancreatic tissue during the course of CP.
242 Prognostic relevance of histomorphologic factors and adjuvant treatment in pancreatic carcinoma G. ASSMANN ~, A. NOLTE 2, R. WILKOWSKI2, J. DIEBOLD 1, U. LOHRS 1 1Pathologisches Institut der Universitat Mtinchen 2Klinik und Poliklinik ftir Strahlentherapie der Universitgt Mtinchen
Aims: Pancreatic carcinoma is a frequent malignant tumour in industrial countries. The impact of histomorphological factors and type of adjuvant therapy on patient outcome has not been clearly elucidated so far. Methods: Retrospective analysis of 57 patients with curatively resected carcinoma of the head of pancreas, which had undergone additionally either radiotherapy (RTX), simultaneous radiochemotherapy (sRCTX) or simultaneous-sequential radiochemotherapy (ssqRCTX). Mostly stage pT3 (UICC 2002). Grading according to the WHO guidelines comprising glandular differentiation, mucine production, mitotic count and nuclear morphology. In addition assessment of perineural invasion, presence of lymphangiosis or hemangiosis and the status of the tissue margins in the specimens. To date univariate analyses of the different factors in relation to survival (multivariate analysis in prep.). Results: Patients with ssqRCTX showed a highly significantlybetter long-time survival compared to patients with RTX or sRCTX, respectively (p = 0.0038; p = 0.0039). In the ssqRCTX-group a tumour-free tissue margin was associated with a significantly better long-time outcome. Low mitotic count, mild nuclear polymorphism and absence of perineural invasion distant from the turnout seemed to have positive effects on survival (not significant due to small number of patients).