Faculty Bibliography

Urothelial tumors develop along two distinctive phenotypic pathways (superficial papillary non-invasive tumors versus flat carcinoma in situ lesions), with markedly different biological behavior and prognosis. Although multiple genetic alterations have been identified in human bladder cancer, their cause-effect relationship with the two pathways has not been firmly established. Using a urothelium-specific promoter of the uroplakin II gene, we showed earlier in transgenic mice that the urothelial expression of SV40T antigen, which inactivates p53 and pRb, induced carcinoma in situ and invasive and metastatic bladder cancer. In striking contrast, we demonstrate here that the urothelial expression of an activated Ha-ras in transgenic mice caused urothelial hyperplasia and superficial papillary non-invasive bladder tumors. These results provide strong, direct experimental evidence that the two phenotypical pathways of bladder tumorigenesis are caused by distinctive genetic defects. Our results indicate that Ha-ras activation can induce urothelial proliferation in vivo; and that urothelial hyperplasia is a precursor of low-grade, superficial papillary bladder tumors. Our transgenic models provide unique opportunities to study the detailed molecular events underlying different types of bladder neoplasms, and can serve as useful preclinical models for evaluating the in vivo efficacy of preventive and therapeutic agents that act on various signaling pathways in bladder cancer

A prostatic smooth muscle cell line (PSMC1) was established from the dorsolateral prostate of p53 null mice. The cell line is nontumorigenic when inoculated subcutaneously, under the renal capsule or intraprostatically in syngeneic mice. These cells express alpha-smooth muscle actin (alpha-SMA), indicating their smooth muscle origin, and TGF-beta significantly enhances expression of alpha-SMA. The cells express both androgen receptor (AR) mRNA and protein, and respond mitogenically to physiological concentrations of androgens. PSMC1 cells produce significant amounts of TGF-beta, which stimulates growth by an autocrine mechanism. Dihydrotestosterone (DHT) increases proliferation of PSMC1 cells by promoting TGF-beta secretion. Considering the significant inhibitory effect of TGF-beta on prostatic epithelial cells and its stimulatory effect on the PSMC1 cells, we postulate that TGF-beta produced by prostatic smooth muscle cells may have a paracrine effect on the prostatic epithelium. We also postulate that TGF-beta may be involved in the etiology of benign prostatic hyperplasia (BPH) by stimulating excessive stromal proliferation. Line PSMC1 is the first reported androgen-responsive murine smooth muscle cell line. It will be useful for in vivo and in vitro experiments to study the mechanisms of androgen action on prostatic stroma and for delineating the interactions that occur between prostatic smooth muscle and epithelium that may lead to prostatic diseases such as BPH

Urothelium synthesizes a group of integral membrane proteins called uroplakins, which form two-dimensional crystals (urothelial plaques) covering >90% of the apical urothelial surface. We show that the ablation of the mouse uroplakin III (UPIII) gene leads to overexpression, defective glycosylation, and abnormal targeting of uroplakin Ib, the presumed partner of UPIII. The UPIII-depleted urothelium features small plaques, becomes leaky, and has enlarged ureteral orifices resulting in the back flow of urine, hydronephrosis, and altered renal function indicators. Thus, UPIII is an integral subunit of the urothelial plaque and contributes to the permeability barrier function of the urothelium, and UPIII deficiency can lead to global anomalies in the urinary tract. The ablation of a single urothelial-specific gene can therefore cause primary vesicoureteral reflux (VUR), a hereditary disease affecting approximately 1% of pregnancies and representing a leading cause of renal failure in infants. The fact that VUR caused by UPIII deletion seems distinct from that caused by the deletion of angiotensin receptor II gene suggests the existence of VUR subtypes. Mutations in multiple gene, including some that are urothelial specific, may therefore cause different subtypes of primary reflux. Studies of VUR in animal models caused by well-defined genetic defects should lead to improved molecular classification, prenatal diagnosis, and therapy of this important hereditary problem

BACKGROUND: Approximately 15% of recipients of live attenuated varicella vaccine may develop mild breakthrough varicella months to years after immunization. Although some vaccinees will develop zoster, it is less common in recipients of vaccine than in those who have had natural varicella. OBJECTIVE: To determine the varicella-zoster virus (VZV) strain responsible for breakthrough varicella and zoster in recipients of varicella vaccine. METHODS: A PCR assay capable of distinguishing wild-type from vaccine strain VZV was performed on samples from skin lesions from vaccinees with breakthrough varicella and zoster. RESULTS: All of 57 vaccinees with breakthrough varicella, clinically diagnosed on the basis of a generalized maculopapular or vesicular rash, in which there was amplifiable DNA [corrected], had wild-type VZV infection based on analysis of viral DNA. The Oka vaccine strain of VZV was not identified in any of these cases. In contrast, in 32 patients with zosteriform rashes, the vaccine strain was identified in 22 samples, and the wild-type strain was identified in 10 samples. CONCLUSIONS: Wild-type virus was identified in all generalized rashes occurring after the immediate 6-week postvaccination period. When reactivation of vaccine strain occurred, it presented as typical zoster. We find no evidence that reactivation of vaccine virus occurs with the clinical picture of generalized rash

PURPOSE: Although a dual origin of the vagina has been popularized, other theories support a mullerian or wolffian duct origin or various combinations of these structures and the urogenital sinus. Uroplakins are specialized membrane proteins of the urothelial plaque, constituting the asymmetrical unit membrane of the bladder, and represent specific molecular markers of urothelial differentiation. We hypothesize that the epithelium of the dorsal wall of the urogenital sinus is involved in the formation of the sinovaginal bulbs and will express uroplakins. In addition, localization of the androgen receptor and its temporal expression during development may in part explain the varied effects of androgens on the lower female genital tract in congenital adrenal hyperplasia. MATERIALS AND METHODS: Lower genitourinary tracts from 4 human female fetuses (9 to 18 weeks) were serially sagittally sectioned. Representative sections were stained with hematoxylin and eosin, rabbit antibodies against panuroplakin and antibodies to the androgen receptor. RESULTS: At 9 weeks of gestation the urogenital sinus showed evidence of evagination and the formation of the sinovaginal bulbs. The urothelium of the entire urogenital sinus expressed uroplakins including the region of the dorsal wall involved in evagination and formation of the sinovaginal bulbs. The mullerian ducts were in direct contact with the area of urogenital sinus evagination but were not in continuity with the sinus. Androgen receptors were expressed in the epithelium and the stroma of the urogenital sinus, sinovaginal bulbs, and mullerian and wolffian ducts. By 14 weeks androgen receptor expression was almost absent in the urothelium of the urogenital sinus, and the epithelium and surrounding stroma of the lower vagina and mullerian ducts. CONCLUSIONS: The area of evagination of the urogenital sinus expresses uroplakins, is involved in the formation of the sinovaginal bulbs and further substantiates the urogenital sinus origin of the lower vagina. Since testosterone inhibits formation of the lower vagina, the timing of exposure to systemic testosterone in congenital adrenal hyperplasia will determine the phenotypic appearance of the external genitalia and effect of testosterone on the development of the lower genital tract. If exposure to testosterone occurs after 12 weeks only clitoromegaly occurs. Androgen receptor is absent in the urogenital sinus urothelium, vaginal epithelium and mullerian ducts by 14 weeks, suggesting that these tissues become androgen insensitive and vaginal development will proceed normally after that critical time

The first rate-limiting step in the conversion of arachidonic acid to PGs is catalyzed by cyclooxygenase (Cox). Two isoforms of Cox have been identified, Cox-1 (constitutively expressed) and Cox-2 (inducible form), which are the products of two different genes. In this study we describe the immunohistochemical localization of Cox-1 and -2 in the human male fetal and adult reproductive tracts. There was no Cox-1 expression in fetal samples (prostate, seminal vesicles, or ejaculatory ducts), and only minimal expression in adult tissues. There was no expression of Cox-2 in the fetal prostate. In a prepubertal prostate there was some Cox-2 expression that localized exclusively to the smooth muscle cells of the transition zone. In adult hyperplastic prostates, Cox-2 was strongly expressed in smooth muscle cells, with no expression in the luminal epithelial cells. Cox-2 was strongly expressed in epithelial cells of both fetal and adult seminal vesicles and ejaculatory ducts. The Cox-2 staining intensity in the fetal ejaculatory ducts during various times of gestation correlated with previously reported testosterone production rates by the fetal testis. These data indicate that Cox-2 is the predominant isoform expressed in the fetal male reproductive tract, and its expression may be regulated by androgens. The distinct cell type-specific expression patterns of Cox-2 in the prostate (smooth muscle) vs. the seminal vesicles and ejaculatory ducts (epithelium) may reflect the different roles of PGs in these tissues

Two prostatic epithelial lines, one of basal origin and one of luminal origin, were established from the dorsolateral prostates of p53 null mice. The cell lines are nontumorigenic when inoculated subcutaneously under the renal capsule or intraprostatically in syngeneic mice. The luminal cell line (PE-L-1) expresses cytokeratins 8 and 18 and the basal cell line (PE-B-1) expresses cytokeratins 5 and 14. The basal cells require serum for growth, whereas the luminal cells grow only in serum-free medium. Both cell lines require the presence of growth factors for optimal growth in culture, with EGF and FGF-2 having the greatest effect on the growth rate. Both lines express androgen receptor (AR) mRNA and protein. Androgen stimulates growth of the basal cell line, indicating that the ARs are functional, whereas growth of the luminal cells is unaffected by androgens. The luminal line is significantly inhibited by exogenous TGF-beta and produces low levels of endogenous TGF-beta. In contrast, the basal cell line produces significant amounts of TGF-beta and its growth is not influenced by this cytokine. Coculture of luminal cells with prostatic smooth muscle cells results in the generation of increased levels of biologically active TGF-beta, indicating a paracrine mechanism of TGF-beta activation that may be involved in the maintenance of normal prostatic function. To our knowledge this is the first report describing both basal and luminal prostatic cell lines from a single inbred animal species and the first indication that prostatic epithelial and stromal cells interact to generate the biologically active form of TGF-beta. These lines will provide an important model for determining basal/luminal interactions in both in vitro and in vivo assays.