Klaus Weichert-Jacobsen

Artificial neural network analysis (ANNA) of prostatic transrectal ultrasound

Our purpose was to determine the diagnostic potential of a new, computerized method of interpreting transrectal ultrasound (TRUS) information by artificial neural network analysis (ANNA). This method was developed to resolve the current dilemma of visual differentiation between benign and malignant tissue on TRUS. To train and objectively evaluate ANNA, a new precise method of computerized virtual correlation of preoperative ultrasound findings and radical prostatectomy histopathology was devised. After training with this pathologically confirmed digitized TRUS information, ANNA was tested in a blinded study.

Expression of human beta-defensins 1 and 2 in kidneys with chronic bacterial infection

BackgroundConstitutive expression and localization of antimicrobial human β-defensin-1 (HBD-1) in human kidneys as a potential mechanism of antimicrobial defense has been previously reported. Inducible expression of human β-defensin-2 (HBD-2) has been described in various epithelial organs but not for the urogenital tract.MethodsWe investigated the gene- and protein expression of HBD-1 and HBD-2 by reverse transcriptase-polymerase chain reaction, and immunohistochemistry in 15 normal human kidney samples and 15 renal tissues with chronic bacterial infection. Additionally, cell culture experiments were performed to study HBD gene expression by real-time RT-PCR in response to inflammatory cytokines TNFα and IL-1β as well as lipopolysaccharide from Gram-negative bacteria.ResultsConstitutive HBD-1 gene- and protein expression was detected in normal renal tissue and kidneys with chronic infection. As a novel finding, inducible HBD-2 gene- and protein expression was demonstrated in tubulus epithelia with chronic infection but not in normal renal tissue. In pyelonephritic kidneys HBD-1 and HBD-2 expression showed a similar pattern of localizaton in distal tubules, loops of Henle and in collecting ducts of the kidney. Furthermore, real-time RT-PCR of kidney derived cell lines stimulated with inflammatory agents TNF-α, IL-1β and LPS revealed a strong increase in relative HBD-2 transcription level and also a slight increase in relative HBD-1 transcription level.ConclusionsUpregulated HBD-2 expression in renal tubulus epithelium indicates a role of a wider range of human defensins for antimicrobial host defense in the urogenital tract than previously recognized.

Cytokeratin–20 Reverse–Transcriptase Polymerase Chain Reaction as a New Tool for the Detection of Circulating Tumor Cells in Peripheral Blood and Bone Marrow of Bladder Cancer Patients

Objectives: Systemic progression is the prevalent form of bladder tumor recurrence after radical cystectomy. The ability to detect circulating tumor cells in peripheral blood or bone marrow could be of prognostic value for the disease with the consequence of early adjuvant chemotherapy. We established a sensitive and specific method using a double cytokeratin–20 (CK–20) reverse–transcriptase polymerase chain reaction (RT–PCR) to detect circulating bladder cancer cells in venous blood and bone marrow Material and Methods: The sensitivity of the detection method was determined by a serial dilution of bladder cancer cells from the cell line HT1376 in whole blood. Bone marrow from 20 bladder cancer patients was drawn prior to radical cystectomy and CK–20 cDNA was amplified by RT–PCR. Additionally, pre– and postoperative venous blood samples from 11 of these patients with bone marrow aspirates and 9 patients undergoing only transurethral resection of the bladder as well as blood samples of 25 healthy volunteers were investigated by CK–20 RT–PCR. Results: The detection limit of the method was 2 bladder cancer cells/ml whole blood containing one million peripheral blood mononuclear cells. The positive detection rate in bone marrow was 7 of 20 (35%) for bladder cancer patients of all stages. However, investigation of the preoperatively collected venous blood samples from 20 patients revealed onyl 2 positive findings, belonging to advanced tumor stages pT4pN0M0 and pT3pN2M0. In contrast, CK–20 was detected in 3 of 20 postoperatively collected venous blood samples from patients with low tumor stages (pTaNXM0 and pT1NXM0) as well as from 1 patient with pelvic lymph node metastases (pT3apN2M0). All venous blood samples of the control group (n = 25) were negative for CK–20. Conclusion: The detection of circulating bladder tumor cells in venous blood and bone marrow by the CK–20 RT–PCR is a promising approach that could improve risk assessment and the identification of bladder cancer patients who would benefit from adjuvant chemotherapy.

Weiterentwicklung des transrektalen Ultraschalls Artifizielle neuronale Netzwerkanalyse (ANNA) in der Erkennung und Stadieneinteilung des Prostatakarzinoms

ZusammenfassungDas prostataspezifische Antigen (PSA) ist heutzutage der meistgenutzte Marker in der Diagnostik des Prostatakarzinoms. Hieraus resultiert eine vermehrte Anzahl von asymptomatischen Männern, die allein durch eine PSA-Werterhöhung Kandidaten für eine weiterführende Prostatadiagnostik werden. Ein deutlich erhöhter PSA-Serumwert (>20 ng/ml) lässt mit hoher Wahrscheinlichkeit auf das Vorhandensein eines Prostatakarzinoms schließen. Im sog. Graubereich zwischen 4 und 10 ng/ml ist der Gewebemarker PSA meist durch gutartige Veränderungen beeinflusst, so dass eine Unterscheidung zwischen maligner und benigner Ursache aufgrund des PSA-Wertes allein nicht möglich ist [1–4]. Darüber hinaus findet man Karzinome bei Patienten, die ein PSA unter dem Normwert von 4 ng/ml aufweisen.Die Methoden, die bislang für die Früherkennung oder Erkennung des Prostatakarzinoms zur Verfügung standen (Tastbefund und Ultraschall) sind unzureichend. So sind ca. 70% der palpablen Tumoren nicht mehr organbegrenzt [5, 6]. Das klassische Problem der visuellen Ultraschallbeurteilung ist die mangelnde Spezifität, insbesondere bei geringer Erfahrung mit der Methode [7–11].Um die diagnostischen Möglichkeiten des transrektalen Ultraschalls (TRUS) in der Prostatakarzinom-Früherkennung und -Stadieneinteilung zu erhöhen, wird in der hier vorgestellten Studie eine Artifizielle Neuronale Netzwerkanalyse (ANNA) eingesetzt, die zusätzliche subvisuelle, graustufendifferente Informationen des TRUS erfassen und auswerten kann [12–14]. Dieser Ansatz erscheint vielversprechend, da Artifizielle Neuronale Netzwerke die im Ultraschallbild vorhandenen komplexen Datenformationen erkennen können, sie gleichsam “lernen” und diese dann bei noch nicht gesehenen Datenformationen wiedererkennen und korrekt klassifizieren können [15].AbstractAs a result of the enhanced clinical application of prostate specific antigen (PSA), an increasing number of men are becoming candidates for prostate cancer work-up. A high PSA value over 20 ng/ml is a good indicator of the presence of prostate cancer, but within the range of 4–10 ng/ml, it is rather unreliable. Even more alarming is the fact that prostate cancer has been found in 12–37% of patients with a “normal” PSA value of under 4 ng/ml (Hybritech). While PSA is capable of indicating a statistical risk of prostate cancer in a defined patient population, it is not able to localize cancer within the prostate gland or guide a biopsy needle to a suspicious area. This necessitates an additional effective diagnostic technique that is able to localize or rule out a malignant growth within the prostate. The methods available for the detection of these prostate cancers are digital rectal examination (DRE) and Transrectal ultasound (TRUS). DRE is not suitable for early detection, as about 70% of the palpable malignancies have already spread beyond the prostate. The classic problem of visual interpretation of TRUS images is that hypoechoic areas suspicious for cancer may be either normal or cancerous histologically. Moreover, about 25% of all cancers have been found to be isoechoic and therefore not distinguishable from normal-appearing areas. None of the current biobsy or imaging techniques are able to cope with this dilemma. Artificial neural networks (ANN) are complex nonlinear computational models, designed much like the neuronal organization of a brain. These networks are able to model complicated biologic relationships without making assumptions based on conventional statistical distributions. Applications in Medicine and Urology have been promising. One example of such an application will be discussed in detail: A new method of Artificial Neural Network Analysis (ANNA) was employed in an attempt to obtain existing subvisual information, other than the gray scale, from conventional TRUS and to improve the accuracy of prostate cancer identification.

p53 expression, proliferation marker Ki-S5, DNA content and serum PSA: Possible biopotential markers in human prostatic cancer

OBJECTIVES The biology of prostate cancer is poorly understood. Despite established prognostic criteria, a confident prediction of the clinical outcome is not always possible. Therefore, additional and more precise information is highly desirable. In the present study, we compared potential biologic markers with the laboratory, clinical, and histopathologic parameters of prostate-specific antigen (PSA) level, tumor stage, and tumor grade. METHODS Paraffin-embedded material from 62 radical prostatectomies for prostate carcinoma was examined immunohistochemically using monoclonal antibody Ki-S5 to determine the tumor growth fraction and antibody DO-1 to assess p53 protein overexpression. Deoxyribonucleic acid-ploidy was analyzed by flow and image cytometry. Preoperative PSA levels were assessed by standard method. The tumors were categorized according to the Gleason grading system and staged postsurgery after the TNM classification. RESULTS The p53 expression, proliferation rate (Ki-S5), and rate of aneuploidy correlated closely with stage (P < 0.05) and Gleason score (P < 0.01). However, divergences were occasionally observed. The ploidy status correlated closely with proliferative activity and p53 expression. Conversely, no correlation was seen between these parameters and serum PSA content, the latter being significantly associated with the tumor stage alone. CONCLUSIONS The results characterize proliferation marker Ki-S5, p53 expression, and ploidy status as tumor biopotential markers, whereas PSA provides diagnostic information. Use of these investigative methods promises to provide additional information relevant in prognosis and therapy selection. Nonetheless, their precise prognostic value will have to be established in further studies.

Prostate-specific antigen in female urine: a prospective study involving 217 women

OBJECTIVES Histomorphologic studies have provided evidence of prostate-specific antigen (PSA)-producing tissue in the female urethra. Some urine samples from women in a small series were positive for PSA, but no systematic investigation of this subject has been done to date. METHODS In a prospective study, we analyzed whether PSA occurs in the urine of women and what factors induce detectable PSA levels. The urine samples of 217 women were analyzed (Hybritech-Tandem E-PSA) under standardized conditions. The impact of urine pH and volume was investigated, and the results were correlated with clinical data (age, residual urine, urinary tract infection and prior sexual intercourse within 48 hours). RESULTS A positive PSA level greater than the detection limit of 0.1 ng/mL was found in 11% of the analyzed samples; their mean value was 0.29 ng/mL. pH correction did not result in a significant difference. The voiding volume had no influence on the PSA level. Among the cases of detectable PSA, women younger than 50 years of age (n = 14) had a mean PSA of 0.34 ng/mL and those older than 50 years (n = 9) a mean of 0.23 ng/mL. One of 9 women with and 22 of 208 women without residual urine volume had a detectable PSA level, as did 0 of 20 with and 23 of 197 women without urinary tract infection, and 3 of 7 with and 20 of 210 women without prior sexual intercourse within the previous 48 hours. None of the differences were significant. CONCLUSIONS A urine PSA level was detected in 11% of all women studied, with PSA values apparently age dependent. Any urine portion is suitable for analysis. No influence was determined for residual urine volume or urinary tract infection. Sexual intercourse may cause detectable PSA values, but the data of this study did not provide sufficient evidence for this hypothesis.

Prognostic indicators for response to therapy and survival in patients with metastatic renal cell cancer treated with interferon alpha-2 beta and vinblastine

OBJECTIVES Only one third of all patients with metastatic renal cell carcinoma respond to immunochemotherapy. Improved patient selection could render such treatment unnecessary in many cases. The goal of the study was to test various factors for their prognostic value as predictors of success of immunochemotherapy and patient survival in metastatic renal cell carcinoma. METHODS Fifty patients with metastatic renal cell carcinoma were subjected to immunochemotherapy with interferon alpha-2 beta and vinblastine. Different variables such as age, sex, location of metastasis, primary or late metastasis, performance status, histologic status, overexpression of the p53 protein and cell proliferation as assessed by immunohistochemistry, and deoxyribonucleic acid-ploidy were considered as potential prognostic factors for response to immunochemotherapy and survival. RESULTS Thirty percent (15) of the cases responded to therapy: 2 complete and 13 partial remissions. In univariate analysis, the proliferative activity (Ki-S5 labeling index) emerged as the statistically most significant prognostic factor (P = 0.0013) for prediction of successful immunochemotherapy in metastatic renal cell carcinoma. The second most significant factor was the location of metastases (P = 0.015), whereas all other parameters did not achieve statistical significance. As to overall survival, responsiveness to therapy was the most significant predictor (P = 0.0003), followed by Ki-S5 scores (P = 0.025). All other factors, including the sites of metastasic spread (P = 0.21), were not statistically relevant. CONCLUSIONS Proliferation status in terms of Ki-S5 immunoreactive scores appears to be a valuable predictor of the responsiveness to immunochemotherapy. Overall survival appears to depend essentially on disease progression and tumor cell proliferation. Other alleged prognostic factors, such as performance status, sarcomatoid histology, and metastasis location, were not significant in this study.

Experiences with the entero-ureteral anastomosis via the extramural serous-lined tunnel : Procedure of Abol-Enein

OBJECTIVES Ureteral reimplantation in any urinary diversion setting should be easy, reproducible, nonrefluxive, and without complications. In this study, we present our experiences with the entero-ureteral anastomosis via the extramural serous-lined tunnel, a technique introduced by Abol-Enein. METHODS In the period between 1995 and 1998 we performed the Abol-Enein technique in 50 patients who underwent radical cystectomy and ileal neobladder construction. Preoperatively 10 patients had unilateral ureteral dilatation. RESULTS The technique was performed easily, and any serious complications did not occur in the early postoperative period. Leakage of the urethral-intestinal anastomosis was seen in 2 patients and transient hydronephrosis in 2 patients. Among the most serious late complications were two strictures of the urethral-intestinal anastomosis and four unilateral strictures of the ureteral-intestinal anastomosis. Only 1 patient presented with unilateral grade IV urinary reflux. Four patients developed metabolic acidosis. In all cases, renal function was stabilized or improved, and preoperative dilatation of the ureters was alleviated. CONCLUSIONS The low stricture rates of the uretero-intestinal anastomosis, as well as the low reflux rates of the procedure of Abol-Enein, render this procedure a safe method of uretero-intestinal anastomosis when an orthotopic ileal neobladder is formed. The method can be performed with equal or even greater ease in very dilated ureters that in many situations other techniques either cannot deal with or require ureteral tailoring. Although this technique seems to meet most of the criteria for an ideal uretero-intestinal anastomosis, further studies that involve longer follow-up periods are necessary to allow a definitive value of this method.

Correlation between DNA ploidy, proliferation marker Ki-67 and early tumor progression in renal cell carcinoma. A prospective study.

OBJECTIVE The course of metastatic renal cell carcinoma shows a broad range of interindividual variation that cannot be sufficiently predicted by tumor stage and grade. The aim of this study was to establish the prognostic value of DNA ploidy and the proliferation marker Ki-67 in renal cell carcinoma. METHODS Both parameters were measured simultaneously in 100 tumors and then correlated with the classic prognostic criteria pathologic stage and tumor differentiation grade as well as clinical course (early tumor progression). RESULTS DNA ploidy correlated well with staging, grading and early progression. The proliferation index (Ki-67) correlated well with tumor stage and histopathological grade but not with the clinical course (early progression). CONCLUSION Due to the diverse biological potential of renal cell carcinoma observation of further clinical course, including late tumor progression, will be necessary to determine whether one of these two indicators can provide additional information beyond what differentiation grade and tumor stage already tell us.

Urinary Leakage of Tubular Enzymes after Shock Wave Lithotripsy

Objectives: Urinary loss of tubular marker enzymes following shock wave lithotripsy (SWL) suggests corresponding morphological changes in the kidney. To date, the morphological correlate of enzymuria and its dependence on the energy applied remains unclear. Methods: In an animal study, the acute morphological changes occurring in the tubulus cells as the basis of enzymuria were investigated. It was evaluated whether SWL-induced enzymuria correlates with the extent of renal damage. Results: Acute morphological changes in the tubulus cells were demonstrated beneath isolated tubulus necrosis. The mechanically induced lesions of the cell organelles included fragmentation of the lysosomes and severe alterations of the cell membrane. The tubulus damage can be quantified. With the help of histochemical N-acetyl-β-D-glucosaminidase (NAG) staining and electron microscopic observations, a significant correlation was found between the shock wave parameters number of impulses and intensity and the tubular damage. The intensity of NAG enzymuria reflected the severity of the tubular damage. Conclusions: In this animal model, NAG proved to be a suitable marker enzyme for estimation of the degree of SWL-induced tubular damage.