Erinn H. Goldman, PhD

Anthony Atala, MD, chair of the department of urology and director of the Institute for Regenerative Medicine at Wake Forest University, Winston-Salem, NC, reviewed the current state of tissue engineering at a symposium on emerging technologies in urology. Dr. Atala told the audience that research in the field has made significant strides since it began in 1990. 

"Over 75 distinct multiple-year research studies related to the engineering of bladder tissue... were performed over a 16-year time span by over 100 researchers and clinicians on our team," he said. 

 Two groups reported novel bladder generation models derived from stem cells. Their studies established the feasibility of generating an entire functioning bladder from embryonic stem cells. Both provide a powerful new model for studying bladder development and determining the potential pathophysiologic impact of various exposures. 

Jang-Won Lee, PhD, a member of Dr. Atala's group, investigated whether nuclear transfer stem cells could differentiate and participate in de novo bladder generation. Early-stage blastocysts were microinjected with single stem cells transduced to express a reporter gene. The blastocysts were then implanted into mouse uteri to enable normal embryonic development. Bladder tissue of the resulting offspring was analyzed. 

The tissue contained differentiated stem cells expressing the appropriate reporter gene.  Through functional and molecular analyses and immunohistochemical investigation, Dr. Lee found that the injected embryonic stem cells differentiated and formed the various cell lineages of a functioning bladder, including urothelium, muscle, and nerves. 

The researchers concluded that stem cells derived through nuclear transfer can differentiate and regenerate a bladder de novo. In addition to its obvious translational application, the researchers propose that this experimental model could be used to study normal bladder development. 

Siam Oottamasathien, MD, a second-year pediatric urology fellow at Vanderbilt University Medical Center, Nashville, TN, won the AAP Section on Urology Basic Science research prize for his study using the signaling environment of the mesenchyme to induce bladder embryonic stem cell differentiation. 

Dr. Oottamasathien, working with John C. Pope, IV, MD, and colleagues, isolated embryonic bladder mesenchyme (EBLM) from embryonic day 18 rat bladders to provide the appropriate signaling environment for embryonic stem cell differentiation. Heterospecific recombinant xenografts were created by combining mouse embryonic stem cells with the isolated EBLM. Xenografts of embryonic stem cells alone, not in the presence of the EBLM signaling environment, were used as a control. The xenografts were implanted in male athymic nude mice and harvested 42 days post-grafting. The grafts were assessed for urothelial and stromal differentiation. 

"Bladder-specific stem cells would likely be a better tissue-delivery cell source than other tissue cells for rehabilitative procedures. We propose that instead of working from a solid organ and trying to find stem cells, it may be more efficient to start with pluripotent embryonic stem cells," Dr. Oottamasathien told 

			"From previous studies, we know that there's a strong inductive signaling pathway in bladder mesenchyme, so we used this environment to try to force embryonic stem cells down a lineage to create a bladder progenitor cell."			

Articles

Atlanta-Tissue engineering research has grown by leaps and bounds in the past decade and a half, yielding findings that may someday have a profound impact on the field of pediatric urology. At the 2006 American Academy of Pediatrics Section on Urology annual meeting here, several groups presented cutting-edge data on novel systems for organ generation, including research demonstrating the feasibility of generating a functioning bladder from stem cells.

Research on bladder tissue regeneration progresses

The current data, while suggesting a cause-and-effect relationship, are largely based on animal models and questionnaire-based studies. 

UCSF researchers, led by Laurence S. Baskin, MD, investigated molecular mechanisms of benzophenone-2 (BP-2)-induced hypospadias. Dr. Hsieh was the runner-up for the AAP Section on Urology Basic Science Research Prize for this work. 

BP-2 is a common additive in cosmetic products and plastic food containers. The UCSF team found that in utero exposure to BP-2 upregulates estrogen receptor (ER) beta in the genital tubercle and causes hypospadias in an ER-dependent manner. 

Pregnant mice were treated with control vehicle, BP-2 alone, or BP-2 plus the ER-antagonist EM-800 from gestational days 12 through 17. The mice were sacrificed on day 18. 

Hypospadias developed in eight (14%) of 57 male fetuses of female mice treated with BP-2 alone (

=.0064). None of the fetuses of mice treated with control vehicle, EM-800, or BP-2 plus EM-800 developed hypospadias. Real-time polymerase chain reaction experiments revealed that the genital tubercles of BP-2-treated male fetuses expressed higher levels of ER-beta than did male controls (

=.04). In fact, levels of ER-beta in females and BP-2-treated males were comparable. 

"BP-2 is so ubiquitous. It's not established whether the dose we tested is relevant to human exposure," Dr. Hsieh said. "Further studies investigating the effect of BP-2 exposure in humans may be warranted." 

 Another in vitro study, performed by John P. Gearhart, MD, and colleagues at the Johns Hopkins Hospital in Baltimore, tested the effects of the phytoestrogen genistein in rats. Previous studies by this group have demonstrated that genistein exposure can lead to deleterious changes in male rat genitalia and reproductive behaviors. The current study aimed to discern the molecular basis of these abnormalities. 

Rats were fed a diet with low levels of genistein (5 mg per kg of food) or a normal diet (control). They continued this diet through timed mating, pregnancy, and weaning. RNA was extracted from the lower genitourinary tissue at gestational days 14, 16, 18, 21, and 2 weeks after birth. 

Atlanta-The impact of environmental toxins on male sexual development has become a topic of keen interest among pediatric urologists, who have released a growing body of data suggesting environmental links to hypospadias, cryptorchidism, and other urinary tract anomalies and diseases. At the 2006 American Academy of Pediatrics Section on Urology annual meeting, several groups presented data on the effect of in utero exposure to endocrine disrupters on the developing fetus.

Growing data point to toxins' role in fetal anomalies

"This was the first descriptive study to use validated questionnaires to assess how the growing population of adult spina bifida patients are doing in terms of sexual function and overall quality of life," said first author Jenny Lassmann, MD, a pediatric urology fellow who worked on the study with Howard M. Snyder, MD, and colleagues. 

Because of advancements in the treatment of spina bifida, a far greater number of patients are progressing into adulthood. As these patients age, their needs change, a fact that is causing researchers to investigate quality of life indicators in such populations. One of these indicators, sexual function, is an important component, and has been noted to be disturbed in patients with other physically limiting diseases. 			

All patients underwent an interview to determine their method of bladder emptying and degree of urinary incontinence. Additionally, each patient completed two questionnaires. The Watts sexual function questionnaire (WSFQ) was used to assess four areas of sexual experience: desire, arousal, orgasm, and satisfaction. The second questionnaire, the Medical Outcomes Study 36-item short-form (SF-36), was used to measure general health-related quality of life. 

 Of the 76 patients evaluated, 24% were sexually active, defined as having had sexual intercourse at least once during the previous 2 months. Using historical controls, sexual activity in these patients was delayed when compared with other groups, including U.S. high school students (55%) and men diagnosed with exstrophy (77%). 

"Our cohort of spina bifida patients is in continued follow-up at a tertiary pediatric center. The number of sexually active patients in our cohort was less than that published in other spina bifida series, perhaps due to the fact that our patients have more significant disease," Dr. Lassmann said. 

Interestingly, no significant differences in quality of life measures were observed between sexually active and non-sexually active spina bifida patients. 

"We were surprised to see the trend, though not statistically significant, that the mental health component of the quality of life score was lower (and therefore closer to that of the general population) in sexually active spina bifida patients than in those who are not sexually active," Dr. Lassmann noted. "Though this seems counterintuitive, it may suggest that sexually active spina bifida patients have a higher psychological and functional status that more closely mirrors the general population." 

This was supported by the finding that sexually active spina bifida patients were more likely to have no hydrocephalus and to live independently. There was no association between sexual activity and age, gender, urinary incontinence, or degree of physical disability in this population. 

In sexually active patients, their sexual function appears to be disturbed, as they scored slightly higher in the WSFQ assessment than did other published disease groups (including women with diabetes or breast cancer and patients with renal cancer or hypertension) and lower than a mixed control group. Dr. Lassmann noted that the mean age of all cohorts was higher than that of the presented spina bifida group. 

Atlanta-Sexual activity is lower and more disturbed in adult patients with spina bifida than it is in the general population, according to a recent study by researchers from the Children's Hospital of Philadelphia. Interestingly, however, this diminished sexual activity does not appear to significantly affect spina bifida patients' quality of life, researchers said at the 2006 American Association of Pediatrics Section on Urology annual meeting here.

Adult spina bifida patients less sexually active

-A study funded by the National Institutes of Health and a new registry promise to provide needed information on the incidence, prevalence, and consequences of two important urologic conditions in children: vesicoureteral reflux and prenatal hydronephrosis. Both the study and the registry were highlighted at the American Academy of Pediatrics Section on Urology annual meeting here. 

Saul Greenfield, MD, director of the division of pediatric urology at Women and Children's Hospital in Buffalo, introduced the NIH-funded Randomized Intervention for Children for Vesicoureteral Reflux (RIVUR) study. This randomized study will evaluate the impact of antimicrobial prophylaxis in improving the symptoms and consequences of VUR. RIVUR investigators will create a database of more than 600 children with VUR and track their outcomes. 

"The incidence of scarring in association with urinary tract infection in children who have reflux is low. There have been challenges to the conventional wisdom that continual prophylaxis is needed in all children with VUR," Dr. Greenfield told the AAP audience. 

The primary endpoint of the study is number, timing, and recurrence of urinary tract infections. The secondary endpoints are renal scarring, as measured by DMSA scan, and antimicrobial resistance in urinary pathogens. 

Other factors that will likely be addressed by RIVUR are compliance with chronic medical therapy, voiding dysfunction, and chronic constipation in children over 3 years of age, antimicrobial resistance in stool cultures, and quality of life. Blood and urine specimens will be kept in a central repository for potential future genetic testing. 

Enrollment in RIVUR will begin in 2007. Investigators hope that data from the study and companion genetic investigations will answer a widely debated question in the pediatric urology community: Should all children with VUR be treated with continuous antimicrobial prophylaxis? 

 Anthony Herndon, MD, assistant professor of the division of urology at the University of Alabama in Birmingham, presented an update of the Society for Fetal Urology Prenatal Hydronephrosis (SFU/Prenatal) registry, which will collect prospective data on patients with prenatal hydronephrosis for 3 years. 

Previous studies have shown that prenatal hydronephrosis is associated with higher risk of kidney abnormalities after birth. 

"A prospective study is needed to further define these risks," Dr. Hernandon said. 

Individual study sites will have complete autonomy over diagnostic and treatment approaches. They will assign a unique identification code to all patients and transmit data to a central server at the University of Alabama. Study site representatives will have password-protected access to the database and can contact a registry manager with any technology-related concerns. Data analysis of individual sites is available for the first 5 years of the study at no cost to the participating institution. 

Open enrollment for the pilot phase of the SFU/Prenatal registry began in May 2006. Of the 63 patients currently enrolled, the majority (66%) have been diagnosed with ureteropelvic junction obstruction. Other commonly observed conditions include VUR (11%) and ureterocele (10%). 

"The registry will not address a specific hypothesis. It is a vehicle to warehouse data. Specific projects will be hypothesis driven and utilize data points that have been populated in the registry," Dr. Herndon explained. 

The architects of the SFU/Prenatal program envision this registry as a tool that will enable researchers to tackle some of the most typical questions in fetal urology, including identification of optimal postnatal treatment approaches for all stages of fetal hydronephrosis and possible refinement of the SFU grading system. 

 Comprehensive registries are the backbone of large-scale epidemiologic research. They facilitate the determination of incidence, prevalence, and treatment success for a number of disorders. 

Atlanta-A study funded by the National Institutes of Health and a new registry promise to provide needed information on the incidence, prevalence, and consequences of two important urologic conditions in children: vesicoureteral reflux and prenatal hydronephrosis. Both the study and the registry were highlighted at the American Academy of Pediatrics Section on Urology annual meeting here.

Author | Erinn H. Goldman, PhD

Articles

Research on bladder tissue regeneration progresses

Growing data point to toxins' role in fetal anomalies

Adult spina bifida patients less sexually active

Initiatives will support VUR, hydronephrosis research