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LB5 Transmitters contributing to the voiding contraction in female rats
LB5 TRANSMITTERS CONTRIBUTING IN FEMALE RATS Strew
T.‘, Tale A.2, Andersson
TO THE
VOIDING
CONTRACTION
K.E.’
‘Lund University Hospital, Department of Clinical and Experimental Pharmacology. Sweden, 2Division of Genetics and Physiology. Laboratory of Animal Physiology, Finland
LB6 LDL RECEPTOR LIKE SCAVANGER RECEPTOR PROSTATE TISSUE
Lund, Turku,
INTRODUCTION & OBJECTIVES: It is well established that there are both cholinergic (acetylcholme) and purinergic (ATP) components in rat bladder contractmn, and that both contribute to bladder emptying. However. there are possibly other transmitters that contribute to bladder emptying. Typically, the micturitian cycle in the rat can be divided into four phases, as recorded by transvesical cystomehy. However, it has still not been established how the transmitters contribute to the different phases, including the intraluminal pressure high frequency oscdlations (IPHFOs). In this study, we used c&methylene-ATP (desensitizing purinoceptors) and atropine (blocking muscarmic receptors) to study m detail the contribution of acetylcholine and ATP to the different phases of the voiding contraction, rhabdosphincter electromyography (RB EMG), and to urine flow. MATERIAL & METHODS: Adult anesthetized (chloral hydrate, 0.9 g/kg ip) female Sprague-Dawley rats, weighing 225.270 g, were used in the study. A 20 gauge iv infusion cannula was inserted through the bladder apex into the lumen for saline (0.9 % NaCl) infusion (0.185 mlimin) and intravesicalpressure recordings. An ultrasonic flow probe connected to a flowmeter was used for measurement of the urine flow from the distal urethra. An 1~ cannula was introduced into the femoral vein. After baseline recordings, a&opine (1 mgikg). a,~-methylene-ATP (0.5 mgikg), or both, were injected iv. Both drugs were dissolved in saline. Three representative voidings were chosen for analysis before and after a drug administration. RESULTS: cr,R-Methylene-ATP decreased significantly (n=8; ~~0.05) the maximal bladder pressure durmg the first phase, whereas atropine was less effective. The maximal bladder pressure during the second phase was also reduced. The IPHFOs were seen after both treatments. Atropine reduced significantly (n=9; p
Sonnleithner
M.‘:
PROTEI1C’ CLASS
Rapp A.‘, Grill
‘Hanusch Krankenhaus, Krankenhaus, Department
R.‘,
(LRP) AND THE HDL BINDING B (SRB) ARE EXPRESSED IN
Stijgermayer
F.’
Department of Urology, Vienna, of Pathology, Vienna, Austria
Austria,
‘Hanusch
1NTRODUCTIO.U & OBJECTIVES: Epidemiological studies suggest that dietary fat might play a role as a risk factor for prostate cancer. Biochemical evidence to substantiate a link between components of dietary fat and prostate cancer cells has been lacking. The expression of the LDL receptor has been reported. As an initial step, we wanted to find out, whether prostate-tissue expresses other lipoprotein receptors, like the LDL receptor like protein (LRP) and the HDL binding scavanger receptor class B (SRB). MATERIAL & .METHODS: From 20 patients which underwent a TUR Prostate we took a sample of the TZ from the right prostate wall (standardized location at seven o’clock). This sample was sent immediately to the pathologist, who performed a frozen analysis (Schnellschnitt?).The other part of the same sample was frozen with N,. 16 samples were identified as BPH, 4 samples were identified as PCA (Gleason 7-9). From these samples we did a western blot analysis to assess LRP and SRB expression. RESULTS: We could demonstrate with the westem blot method the LRP receptor in 15 samples of BPH and in all prostate SRB was expressed in 13 BPH and in 3 PCA samples.
the expression of cancer samples.
CONCLUSIONS: We could show for the first time the expression of LRP and SRB in prostate tissue. From the data acquired we assume an involvement of the low-density lipoprotein receptor related protein (LRP) in the basal cholesterolmetabolism, present in prostate samples. In addition, we suggest that the scavanger receptor class B (SRB) might play a key-role in retrograde cholesterol transport. It may be down-regulated in malign tissues. Yet, only a small number of patients were enrolled for this study. More work has to be done on this issue.
LB7 CLASSIFICATION BLADDER TISSUE
OF HEALTHY BLADDER BY RAMAN SPECTROSCOPY
TISSUE
AND
TUMOUR
De Jonrr B.‘, Van der Kwast T.‘, Bangma C.‘, Maquelin K.“, Puppels G.“, Kok D.J.’ ‘Erasmus MC, Department of Paediatric Urology, Rotterdam, The Netherlands, ‘Erasmus MC. Department of Pathology, Rotterdam, The Netherlands, iErasmus MC, Department of Urology, Rotterdam, The Netherlands. 4Erasmus MC, Department of General Surgery, Rotterdam, The Netherlands INTRODUCTIOiK & OBJECTIVES: Cystoscopy, the standard tool for diagnosing bladder tumoul-s, often misses small lesions. New techniques are developed to reveal either the presence of a turnour (urine markers), changes in tissue morphology or the location of a turnour area (e.g. 5.ALA fluorescence). However, no technique has been apphed before that both localizes suspected areas and characterizes the morphological and compositional changes in a single measurement without the use of contrasting agents. Raman spectroscopy (RS) is a nondestructive optical technique provides detailed information on the biochemical composition of the tissue analyzed. Because Raman spectra are molecule-specific, different tissues can be distinguished based on their different biochemical composition. Similarlv. biochemical alterat& within a tissue, due to pathogenesis, can be &alyred. A may& advantaee is that RS can also be annlied in 1;1vo usine fiber ootic orobes. hl viva RS inside the bladderallows for nondestructivy detection and crharacter~zati~n of diseased tissue areas. Diagnosis is therefore not limited by a restricted amount of biopsies. For this initial s’rudy on bladder carcinomas we applied in Ctm RS mappmg, which allows for detailed characterization of all molecular changes in a tissue section, as a fundamental step towards in vilio applications. MATERIAL & METHODS: RS mapping was performed on frozen sections cut from bladder biopsies that were air-dried but otherwise untreated. Raman spectra were obtained from a grid-of well-defined positions in the tissue sections. using a m&scope as a sampling interface. All soectra from a RS man that have similar soectral features are identified using hierarchical cl&er analysis. These ‘clusters of similar spectra were found to correlate wit; areas of normal, inflamed or tumour bladder tissue. A HE stain was performed on the section after each RS mapping measurement Photo images were taken and compared to the outlines of the clustering results for characterization of the spectra. In total 19 RS maps were identically analyzed. Averaged spectra of all clusters from all RS maps were comb&d in one dataset. Onlv snectra that could be correctlv identified accordine to the histolozv were used to build a pred&n model based on a linear hiscriminant analysig The model was developed to classify independent spectra as normal or cancerous. Subsequently, the model determines the presence of inflammation in normal or cancerous bladder tissue. RESULTS: In our prelimmary results, the model showed an accuracy of > 95% between normal tissue spectra and tumonr tissue spectra. Within the normal group nomx+l tissue was discriminated from inflammation with an accuracy oft 97%. In the tUmonr group there was an accuracy of 100% in discrnninating tumour tissue and turnour tissue with inflammation. CONCLUSIONS: In conclusioni with RS. we could discriminate normal bladder tissue from turnour bladder tissue very effectively. This may open the way for nondeshxctive in VIYO RS bladder tumour diagnosis.
LB8 PRESENCE HUMAN
Pomara ~.
OF POLYOMAVIRUS ADRENAL
G.‘.
Barzon
AND HERPERVIRUS
DXA SEQUENCES
1N
TUMOURS
L.‘,
De Maria
M.‘,
Evangelisti
S.‘, Carlino
F.‘, Morelli
G.‘,
Paid G.‘. Selli C. ’ ‘S. Chiara Hospital, Padova, Department Italy
Urology Unit, Department of Histology, Microbiology
of Surgery. Piss, Italy, ‘University of and Medical Biotechnologies, Padova,
INTRODUCTION & OBJECTIVES: The diagnosis of adrenal tumours is presently more frequent but their pathogenesis is still largely unknown. Several viruses, including polyomavimses and herpesviruses, can cause cancer in humans and some of them have been demonstrated to present a tropism for the adrenal gland. However human tumour samples have never been systematically screened for the presence of these DNA viruses. MATERIAL & METHODS: A sensitive real-time PCR technique was employed to detect polyomavims (SV40, JCV, BKV) and herpesvirus (HSV-I, HSV-2, HHV8, EBV, CMV) DNA sequences in a large series of adrenal turnours, including 7 aldosteroneproducing adenomas (APA), 12 cortisol-producing adenomas/hyperplasia (CPA), 16 non-functioning adrenal adenomas (NFA), 9 adrenocortical carcinomas (ACC), 6 pheochromocytomas (including I malignant), 2 myelolipomas, and 1 case of primary bilateral adrenal non-Hodgkin’s lymphoma. Adjacent normal adrenal tissue was available in IO cases. RESULTS: SV40 DNA was found at very high copy number in the single case of malignant pheochromocytoma, and, at lower levels, in 1 ACC and in 2 APAs: JCV DNA was detected only in the adrenal non-Hodgkin’s lymphoma sample; BKV sequences were found in 7% of adrenal tmnours, but not in the normal adrenal gland. EBV DNA was detected in 62% of adrenal tumours and in 50% of normal adrenal tissues; of note, high copy number of EBV DNA was found in the non-Hodgkin’s lymphoma. CMV DNA was positive in 27% of benign adrenal turnours, but not in ACCs. Immunohistochemical analysis demonstrated the presence of CMV Immediate-early proteins in positive samples. The prevalence of HSV and HHV8 sequences in adrenal samples was low (3% and 5%, respectively) and not associated with the histological type. Co-infections were present in 28% of cases (most frequently EBV+CMV and CMV+BKV), whereas 16% of cases showed no viral genomes. Tumours from patients with severe hypercortisolism frequently showed co-infections and higher viral titers. CONCLUSIONS: This is the first report of analysis adrenal tumours and corresponding normal tissue. The infection, which is found in about 60% of normal and or in association with CMV. SV40 could be involved pheochromocytoma and ACC.
European
Urology
Supplements
of viral sequences in the human adrenal gland is a target for EBV neoplastic adrenals, either alone in the parhogenesis of malignant
3 (2004)
No. 2, pp. 233
T.‘, Tale A.2, Andersson
TO THE
VOIDING
CONTRACTION
K.E.’
‘Lund University Hospital, Department of Clinical and Experimental Pharmacology. Sweden, 2Division of Genetics and Physiology. Laboratory of Animal Physiology, Finland
LB6 LDL RECEPTOR LIKE SCAVANGER RECEPTOR PROSTATE TISSUE
Lund, Turku,
INTRODUCTION & OBJECTIVES: It is well established that there are both cholinergic (acetylcholme) and purinergic (ATP) components in rat bladder contractmn, and that both contribute to bladder emptying. However. there are possibly other transmitters that contribute to bladder emptying. Typically, the micturitian cycle in the rat can be divided into four phases, as recorded by transvesical cystomehy. However, it has still not been established how the transmitters contribute to the different phases, including the intraluminal pressure high frequency oscdlations (IPHFOs). In this study, we used c&methylene-ATP (desensitizing purinoceptors) and atropine (blocking muscarmic receptors) to study m detail the contribution of acetylcholine and ATP to the different phases of the voiding contraction, rhabdosphincter electromyography (RB EMG), and to urine flow. MATERIAL & METHODS: Adult anesthetized (chloral hydrate, 0.9 g/kg ip) female Sprague-Dawley rats, weighing 225.270 g, were used in the study. A 20 gauge iv infusion cannula was inserted through the bladder apex into the lumen for saline (0.9 % NaCl) infusion (0.185 mlimin) and intravesicalpressure recordings. An ultrasonic flow probe connected to a flowmeter was used for measurement of the urine flow from the distal urethra. An 1~ cannula was introduced into the femoral vein. After baseline recordings, a&opine (1 mgikg). a,~-methylene-ATP (0.5 mgikg), or both, were injected iv. Both drugs were dissolved in saline. Three representative voidings were chosen for analysis before and after a drug administration. RESULTS: cr,R-Methylene-ATP decreased significantly (n=8; ~~0.05) the maximal bladder pressure durmg the first phase, whereas atropine was less effective. The maximal bladder pressure during the second phase was also reduced. The IPHFOs were seen after both treatments. Atropine reduced significantly (n=9; p
Sonnleithner
M.‘:
PROTEI1C’ CLASS
Rapp A.‘, Grill
‘Hanusch Krankenhaus, Krankenhaus, Department
R.‘,
(LRP) AND THE HDL BINDING B (SRB) ARE EXPRESSED IN
Stijgermayer
F.’
Department of Urology, Vienna, of Pathology, Vienna, Austria
Austria,
‘Hanusch
1NTRODUCTIO.U & OBJECTIVES: Epidemiological studies suggest that dietary fat might play a role as a risk factor for prostate cancer. Biochemical evidence to substantiate a link between components of dietary fat and prostate cancer cells has been lacking. The expression of the LDL receptor has been reported. As an initial step, we wanted to find out, whether prostate-tissue expresses other lipoprotein receptors, like the LDL receptor like protein (LRP) and the HDL binding scavanger receptor class B (SRB). MATERIAL & .METHODS: From 20 patients which underwent a TUR Prostate we took a sample of the TZ from the right prostate wall (standardized location at seven o’clock). This sample was sent immediately to the pathologist, who performed a frozen analysis (Schnellschnitt?).The other part of the same sample was frozen with N,. 16 samples were identified as BPH, 4 samples were identified as PCA (Gleason 7-9). From these samples we did a western blot analysis to assess LRP and SRB expression. RESULTS: We could demonstrate with the westem blot method the LRP receptor in 15 samples of BPH and in all prostate SRB was expressed in 13 BPH and in 3 PCA samples.
the expression of cancer samples.
CONCLUSIONS: We could show for the first time the expression of LRP and SRB in prostate tissue. From the data acquired we assume an involvement of the low-density lipoprotein receptor related protein (LRP) in the basal cholesterolmetabolism, present in prostate samples. In addition, we suggest that the scavanger receptor class B (SRB) might play a key-role in retrograde cholesterol transport. It may be down-regulated in malign tissues. Yet, only a small number of patients were enrolled for this study. More work has to be done on this issue.
LB7 CLASSIFICATION BLADDER TISSUE
OF HEALTHY BLADDER BY RAMAN SPECTROSCOPY
TISSUE
AND
TUMOUR
De Jonrr B.‘, Van der Kwast T.‘, Bangma C.‘, Maquelin K.“, Puppels G.“, Kok D.J.’ ‘Erasmus MC, Department of Paediatric Urology, Rotterdam, The Netherlands, ‘Erasmus MC. Department of Pathology, Rotterdam, The Netherlands, iErasmus MC, Department of Urology, Rotterdam, The Netherlands. 4Erasmus MC, Department of General Surgery, Rotterdam, The Netherlands INTRODUCTIOiK & OBJECTIVES: Cystoscopy, the standard tool for diagnosing bladder tumoul-s, often misses small lesions. New techniques are developed to reveal either the presence of a turnour (urine markers), changes in tissue morphology or the location of a turnour area (e.g. 5.ALA fluorescence). However, no technique has been apphed before that both localizes suspected areas and characterizes the morphological and compositional changes in a single measurement without the use of contrasting agents. Raman spectroscopy (RS) is a nondestructive optical technique provides detailed information on the biochemical composition of the tissue analyzed. Because Raman spectra are molecule-specific, different tissues can be distinguished based on their different biochemical composition. Similarlv. biochemical alterat& within a tissue, due to pathogenesis, can be &alyred. A may& advantaee is that RS can also be annlied in 1;1vo usine fiber ootic orobes. hl viva RS inside the bladderallows for nondestructivy detection and crharacter~zati~n of diseased tissue areas. Diagnosis is therefore not limited by a restricted amount of biopsies. For this initial s’rudy on bladder carcinomas we applied in Ctm RS mappmg, which allows for detailed characterization of all molecular changes in a tissue section, as a fundamental step towards in vilio applications. MATERIAL & METHODS: RS mapping was performed on frozen sections cut from bladder biopsies that were air-dried but otherwise untreated. Raman spectra were obtained from a grid-of well-defined positions in the tissue sections. using a m&scope as a sampling interface. All soectra from a RS man that have similar soectral features are identified using hierarchical cl&er analysis. These ‘clusters of similar spectra were found to correlate wit; areas of normal, inflamed or tumour bladder tissue. A HE stain was performed on the section after each RS mapping measurement Photo images were taken and compared to the outlines of the clustering results for characterization of the spectra. In total 19 RS maps were identically analyzed. Averaged spectra of all clusters from all RS maps were comb&d in one dataset. Onlv snectra that could be correctlv identified accordine to the histolozv were used to build a pred&n model based on a linear hiscriminant analysig The model was developed to classify independent spectra as normal or cancerous. Subsequently, the model determines the presence of inflammation in normal or cancerous bladder tissue. RESULTS: In our prelimmary results, the model showed an accuracy of > 95% between normal tissue spectra and tumonr tissue spectra. Within the normal group nomx+l tissue was discriminated from inflammation with an accuracy oft 97%. In the tUmonr group there was an accuracy of 100% in discrnninating tumour tissue and turnour tissue with inflammation. CONCLUSIONS: In conclusioni with RS. we could discriminate normal bladder tissue from turnour bladder tissue very effectively. This may open the way for nondeshxctive in VIYO RS bladder tumour diagnosis.
LB8 PRESENCE HUMAN
Pomara ~.
OF POLYOMAVIRUS ADRENAL
G.‘.
Barzon
AND HERPERVIRUS
DXA SEQUENCES
1N
TUMOURS
L.‘,
De Maria
M.‘,
Evangelisti
S.‘, Carlino
F.‘, Morelli
G.‘,
Paid G.‘. Selli C. ’ ‘S. Chiara Hospital, Padova, Department Italy
Urology Unit, Department of Histology, Microbiology
of Surgery. Piss, Italy, ‘University of and Medical Biotechnologies, Padova,
INTRODUCTION & OBJECTIVES: The diagnosis of adrenal tumours is presently more frequent but their pathogenesis is still largely unknown. Several viruses, including polyomavimses and herpesviruses, can cause cancer in humans and some of them have been demonstrated to present a tropism for the adrenal gland. However human tumour samples have never been systematically screened for the presence of these DNA viruses. MATERIAL & METHODS: A sensitive real-time PCR technique was employed to detect polyomavims (SV40, JCV, BKV) and herpesvirus (HSV-I, HSV-2, HHV8, EBV, CMV) DNA sequences in a large series of adrenal turnours, including 7 aldosteroneproducing adenomas (APA), 12 cortisol-producing adenomas/hyperplasia (CPA), 16 non-functioning adrenal adenomas (NFA), 9 adrenocortical carcinomas (ACC), 6 pheochromocytomas (including I malignant), 2 myelolipomas, and 1 case of primary bilateral adrenal non-Hodgkin’s lymphoma. Adjacent normal adrenal tissue was available in IO cases. RESULTS: SV40 DNA was found at very high copy number in the single case of malignant pheochromocytoma, and, at lower levels, in 1 ACC and in 2 APAs: JCV DNA was detected only in the adrenal non-Hodgkin’s lymphoma sample; BKV sequences were found in 7% of adrenal tmnours, but not in the normal adrenal gland. EBV DNA was detected in 62% of adrenal tumours and in 50% of normal adrenal tissues; of note, high copy number of EBV DNA was found in the non-Hodgkin’s lymphoma. CMV DNA was positive in 27% of benign adrenal turnours, but not in ACCs. Immunohistochemical analysis demonstrated the presence of CMV Immediate-early proteins in positive samples. The prevalence of HSV and HHV8 sequences in adrenal samples was low (3% and 5%, respectively) and not associated with the histological type. Co-infections were present in 28% of cases (most frequently EBV+CMV and CMV+BKV), whereas 16% of cases showed no viral genomes. Tumours from patients with severe hypercortisolism frequently showed co-infections and higher viral titers. CONCLUSIONS: This is the first report of analysis adrenal tumours and corresponding normal tissue. The infection, which is found in about 60% of normal and or in association with CMV. SV40 could be involved pheochromocytoma and ACC.
European
Urology
Supplements
of viral sequences in the human adrenal gland is a target for EBV neoplastic adrenals, either alone in the parhogenesis of malignant
3 (2004)
No. 2, pp. 233