Christian Radmayr

Regulation of prostatic growth and function by peptide growth factors

Polypeptide growth factors are positive and negative regulators of prostatic growth and function. Expression and biological effects of epidermal growth factor (EGF), transforming growth factors (TGFs) α and β, fibroblast growth factors (FGFs), and insulin‐like growth factors (IGFs) in the prostate have been extensively studied. EGF and TGFα, which share the same receptor, are strong mitogens for prostatic epithelial and stromal cells. Their paracrine mode of action in normal tissue and early‐stage tumors is apparently altered towards an autocrine stimulation in hormone‐independent tumors, which gain the ability to produce TGFα by themselves. TGFβ has a dual role in the regulation of prostatic growth. It inhibits growth of prostatic epithelial cells in culture and mediates programmed cell death after androgen withdrawal. However, advanced prostatic carcinomas become insensitive to the inhibitory effect of TGFβ. Several members of the FGF family have been identified in the prostate. They are mainly or exclusively expressed in the stromal cells, and stimulate the epithelial cells. In the rat Dunning tumor model, progression is accompanied by distinct changes in the expression of FGFs and their receptors. In the hyperplastic tissue, basic FGF (bFGF) is accumulated. This growth factor is also a potent angiogenic inducer, expression of which may determine the metastatic capability of a tumor. IGFs are paracrine growth stimulators in the normal and hyperplastic prostate. It is still under consideration whether prostatic cancer cells gain the ability to produce IGF‐I by themselves and thus shift to an autocrine mode of IGF‐I stimulation. Growth factors also interact with the androgen‐signaling pathway. IGF‐I in particular, other growth factors as well, can activate the androgen receptor.

CELLULAR AND HUMORAL IMMUNE RESPONSES IN PATIENTS WITH METASTATIC RENAL CELL CARCINOMA AFTER VACCINATION WITH ANTIGEN PULSED DENDRITIC CELLS

PURPOSE Dendritic cells are the most potent stimulators of immune responses including antitumor responses. We performed a pilot study of cultured antigen loaded dendritic cells in patients with metastatic renal cell carcinoma. MATERIALS AND METHODS Dendritic cells were obtained by culturing plastic adherent mononuclear cells from peripheral blood for 5 days in the presence of granulocyte-macrophage colony-stimulating factor and interleukin-4. Day 5 dendritic cells were loaded with cell lysate from cultured autologous tumor cells and with the immunogenic protein keyhole-limpet hemocyanin (KLH) which serves as a helper antigen and as a tracer molecule. During the antigen pulse dendritic cells were activated with a combination of tumor necrosis factor-alpha and prostaglandin E2. Dendritic cells were administered by 3 intravenous infusions at monthly intervals. Cellular and humoral immune responses to KLH and cell lysate were measured in vitro before and after the vaccinations. RESULTS Preparation of 12 dendritic cell vaccines from patients with advanced renal cell carcinoma was successful. Treatment with fully activated CD83+ dendritic cells was well tolerated with moderate fever as the only side effect. Potent immunological responses to KLH and, most importantly, against cell lysate could be measured in vitro after the vaccinations. CONCLUSIONS Our data demonstrate that a dendritic cell based vaccine can induce antigen specific immunity in patients with metastatic renal cell carcinoma. Dendritic cell based immunotherapy represents a feasible, well tolerated and promising new approach for the treatment of advanced renal cell carcinoma.

Human renal‐cell carcinoma tissue contains dendritic cells

Immune surveillance of cancer requires antigen‐presenting cells which activate T cells specific for tumor‐associated antigens. We show here that substantial numbers of dendritic cells, which are the most potent antigen‐presenting cells, emigrate from renal‐tumor explants in organ culture. Tumor‐derived dendritic cells presented with all characteristics of mature dendritic cells. Dendritic cells could be identified by typical cytoplasmic projections (=veils). They expressed high levels of MHC products and of the co‐stimulator CD86 (B7‐2). Dendritic cells expressed the CD45RO isoform but not CD45RA. The most important point was that up to 9% of the emigrating leukocytes expressed the CD83 antigen, a specific marker for mature dendritic cells. CD83+ cells were approximately 40‐fold enriched in the tumor tissue as compared to the peripheral blood. In contrast to cultured blood dendritic cells, tumor‐emigrant dendritic cells had a reduced potential to capture soluble antigen, as shown by the exclusion of fluoresceinated Dextran molecules. Finally, in mixed leukocyte reactions, tumor‐derived dendritic cells were able to stimulate naive T cells from cord blood, which is a unique feature of dendritic cells. This study demonstrates that genuine dendritic cells reside in or infiltrate renal‐cell carcinoma tissue. The failure of patients with renal‐cell carcinoma to mount an anti‐tumor immune response despite the presence of professional antigen‐presenting cells in the tumor tissue suggests that tumor‐associated dendritic cells are suppressed in situ, in a similar way to that described for tumor‐infiltrating lymphocytes.

Androgen receptor status of lymph node metastases from prostate cancer.

To date androgen receptor (AR) expression and structure in human prostatic cancer have been studied in primary tumor specimens and in cell lines. Investigation of alterations in the androgen‐signalling transduction cascade in prostatic carcinoma metastases is important to improve our understanding of tumor progression towards androgen insensitivity. In the present study we have collected data comparing AR expression in both the primary tumors and the respective pelvic lymph node metastases. Formalin‐fixed and paraffin‐embedded tissues derived from the primary tumors and positive lymph nodes of 12 patients undergoing radical prostatectomy were immunostained for the AR and prostate‐specific antigen (PSA). AR expression was evaluated with the polyclonal antibody PG‐21, which is directed against amino acids 1–21 in the N‐terminal region of the AR. All primary tumors stained for the AR. In 8 of the 12 lymph nodes examined more than 50% of the tumor cells were AR positive and displayed a uniform staining pattern; in one lymph node metastasis remarkable heterogeneity in AR expression was observed. In two cases less than 10% of the tumor cells stained for the AR. In one case the lymph node metastasis was immunohistochemically negative for the AR, whereas the primary tumor obtained from the same patient displayed intense staining for the AR. PSA was expressed in all metastases and primary tumors. Our data demonstrate that loss of the AR in lymph node metastases from prostatic carcinoma is a rare event.

Bacillus Calmette‐Guérin mycobacteria stimulate human blood dendritic cells

Bacillus Calmette‐Guérin (BCG) mycobacteria have been used as adjuvant in the active immunotherapy of various human cancers. In addition, dendritic cells, which are the most potent antigen‐presenting cells, have been shown to be capable of initiating anti‐tumor immune responses. Here we investigated the effects of BCG on dendritic cells cultured from human blood. Addition of BCG resulted in rapid homotypic adhesion of dendritic cells. Moreover, BCG concentrations ranging from 104 to 106 bacteria/ml enhanced expression of the dendritic‐cell‐maturation antigen CD83 and of the T‐cell co‐stimulator CD86 (B7‐2) in a dose‐dependent manner. Concomitant with the increase of CD83 and CD86 expression, the cells lost the ability to capture soluble antigens, as determined by the exclusion of fluoresceinated Dextran molecules. Strikingly, the same dosages of BCG‐bacteria stimulated TNF‐α‐gene transcription and TNF‐α‐protein release from dendritic cells in a dose‐dependent fashion. BCG infection of dendritic cells in the presence of a neutralizing antibody directed against TNF‐α inhibited CD83 expression by more than 50% indicating that the BCG‐induced maturation of dendritic cells was at least partially mediated by dendritic‐cell‐derived TNF‐α. The finding that BCG activates the most potent antigen‐presenting cells reveals a plausible immunological mechanism of the occasionally observed anti‐tumor activity of BCG.

LAPAROSCOPIC HEMINEPHROURETERECTOMY IN PEDIATRIC PATIENTS

PURPOSE An increasing number of operative procedures in pediatric urology can be performed by laparoscopy. We report our experience with laparoscopic heminephroureterectomy, which is a typical operation in pediatric patients. MATERIALS AND METHODS Laparoscopic heminephroureterectomy was performed in 14 consecutive children. In 12 cases 7 upper renal poles were removed for ectopic refluxing megaureter and obstructive ureterocele in 5 and 2, respectively. In 5 children lower poles were destroyed by reflux nephropathy. In 2 children laparoscopic upper pole heminephroureterectomy for obstructive ureterocele was combined with a Pfannenstiel incision for reimplantation of the refluxing lower pole ureter. RESULTS All operations were completed as planned. Operative time was 180 to 330 minutes (mean 222) in group 1 and 345 to 510 (mean 427) in group 2. Blood loss was minimal (10 to 30 ml.) and there were no intraoperative or postoperative complications. Mean postoperative hospital stay in groups 1 and 2 was 4.4 and 7.5 days, respectively. CONCLUSIONS Laparoscopic heminephroureterectomy in children is feasible and associated with minimal blood loss, low morbidity and a low complication rate. The disadvantage is the long operative time. This technically demanding procedure should be performed only at specialized centers.

EAU Guidelines on Vesicoureteral Reflux in Children

CONTEXT Primary vesicoureteral reflux (VUR) is a common congenital urinary tract abnormality in children. There is considerable controversy regarding its management. Preservation of kidney function is the main goal of treatment, which necessitates identification of patients requiring early intervention. OBJECTIVE To present a management approach for VUR based on early risk assessment. EVIDENCE ACQUISITION A literature search was performed and the data reviewed. From selected papers, data were extracted and analyzed with a focus on risk stratification. The authors recognize that there are limited high-level data on which to base unequivocal recommendations, necessitating a revisiting of this topic in the years to come. EVIDENCE SYNTHESIS There is no consensus on the optimal management of VUR or on its diagnostic procedures, treatment options, or most effective timing of treatment. By defining risk factors (family history, gender, laterality, age at presentation, presenting symptoms, VUR grade, duplication, and other voiding dysfunctions), early stratification should allow identification of patients at high potential risk of renal scarring and urinary tract infections (UTIs). Imaging is the basis for diagnosis and further management. Standard imaging tests comprise renal and bladder ultrasonography, voiding cystourethrography, and nuclear renal scanning. There is a well-documented link with lower urinary tract dysfunction (LUTD); patients with LUTD and febrile UTI are likely to present with VUR. Diagnosis can be confirmed through a video urodynamic study combined with a urodynamic investigation. Early screening of the siblings and offspring of reflux patients seems indicated. Conservative therapy includes watchful waiting, intermittent or continuous antibiotic prophylaxis, and bladder rehabilitation in patients with LUTD. The goal of the conservative approach is prevention of febrile UTI, since VUR will not damage the kidney when it is free of infection. Interventional therapies include injection of bulking agents and ureteral reimplantation. Reimplantation can be performed using a number of different surgical approaches, with a recent focus on minimally invasive techniques. CONCLUSIONS While it is important to avoid overtreatment, finding a balance between cases with clinically insignificant VUR and cases that require immediate intervention should be the guiding principle in the management of children presenting with VUR.

Optimizing the Operative Treatment of Boys with Varicocele: Sequential Comparison of 4 Techniques

PURPOSE We compared 4 techniques of varicocele ligation in boys and young adolescents to determine the optimal operative treatment that avoids varicocele recurrence and postoperative hydrocele formation. MATERIALS AND METHODS In 10 years a total of 128 varicocelectomies were performed sequentially in 121 boys and young adolescents with a mean age of 12 years using the laparoscopic, inguinal testicular artery sparing, standard Palomo (high mass retroperitoneal ligation) and modified Palomo approaches. The modified Palomo approach involved suprainguinal and retroperitoneal ligation of the veins and artery, and microsurgical sparing of the blue stained lymphatic pathway of the testis. Patients were followed a mean of 52 months. RESULTS In the 19 boys in the laparoscopy group varicocele persisted in 10% and hydrocele developed in 5%. In the 21 patients who underwent inguinal surgery with artery preservation recurrent varicoceles were identified in 14% and no hydroceles were observed. In the 32 patients who underwent the standard Palomo procedure there was no palpable varicocele persistence or recurrence, while hydroceles developed in 12%. Of the 56 patients in the modified Palomo group varicocele recurred in 1 (2%) and there were no hydroceles. No testicular atrophy developed in any patient. CONCLUSIONS Comparison of all 4 groups revealed significant differences in varicocele recurrence (p = 0.038) and hydrocele formation (p = 0.023). Pairwise group comparison showed that the modified Palomo technique resulted in a significant decrease in the incidence of postoperative hydrocele formation compared with the standard Palomo method (p = 0.015). This procedure can be recommended as the optimal surgical technique for varicocele treatment in males of this young age.

Basic fibroblast growth factor levels in cancer cells and in sera of patients suffering from proliferative disorders of the prostate.

Both benign and malignant growth of the prostate depend on the induction of a microvasculature. Basic fibroblast growth factor (bFGF), a potent angiogenic factor, is thought to play an important role in this process.

Human prostatic smooth muscle cells in culture: Estradiol enhances expression of smooth muscle cell-specific markers

Smooth muscle cells (SMCs) constitute a major cellular component of prostatic stroma. SMC tension plays an important role in urethral obstruction secondary to benign prostatic hyperplasia (BPH). We have developed an in vitro procedure for the propagation of human prostatic SMCs. Tissue specimens from patients undergoing radical prostatectomy or cystectomy were enzymatically disaggregated and cultured in MCDB‐131 medium supplemented with horse serum, insulin, conditioned medium from the tumor cell line CRL‐5813, and steroid hormones. The medium was assembled on the basis of the effects these supplements have on the growth of SMC cultures and on the expression of the two markers desmin and smooth muscle myosin. Addition of 0.1 μM of estradiol to the growth medium dramatically increased expression of these SMC‐specific markers. Dihydrotestosterone (DHT) and hydrocortisone had a similar, albeit less pronounced effect. At three to five passages, about two thirds of the cells were immunohistologically positive for smooth muscle myosin or desmin. Almost all cells were positive for the myofibroblast marker smooth muscle α‐actin throughout 10 passages and more. In SMC cultures, cells staining for smooth muscle myosin and desmin were found to seek direct contact to myofibroblasts. They grew in aggregates on a layer of myofibroblasts which adhered to the surface of the culture vessel. As revealed by transmission electron microscopy the cultured cells exhibited morphological features of myofibroblasts. Characteristics of smooth muscle cells, such as prominent bundles of microfilaments associated with dense bodies, basal laminae investing the cells, and numerous caveolae at the cell surfaces were regularly observed in cultures of low passages. After several passages, these features were markedly decreased and organelles of the biosynthetic system became more prominent. In summary, we present an in vitro model of prostatic SMCs and demonstrate that steroid hormones have characteristic effects on these cells. SMC cultures are expected to facilitate investigation of the functions and properties of human prostatic SMCs. Prostate 30:117–129, 1997.