Preliminary report: Surgical outcomes following genitoplasty in children with moderate to severe genital atypia.

INTRODUCTION: Prior studies of outcomes following genitoplasty have reported high rates of surgical complications among children with atypical genitalia. Few studies have prospectively assessed outcomes after contemporary surgical approaches. OBJECTIVE: The current study reported the occurrence of early postoperative complications and of cosmetic outcomes (as rated by surgeons and parents) at 12 months following contemporary genitoplasty procedures in children born with atypical genitalia. STUDY DESIGN: This 11-site, prospective study included children aged ≤2 years, with Prader 3-5 or Quigley 3-6 external genitalia, with no prior genitoplasty and non-urogenital malformations at the time of enrollment. Genital appearance was rated on a 4-point Likert scale. Paired t-tests evaluated differences in cosmesis ratings. RESULTS: Out of 27 children, 10 were 46,XY patients with the following diagnoses: gonadal dysgenesis, PAIS or testosterone biosynthetic defect, severe hypospadias and microphallus, who were reared male. Sixteen 46,XX congenital adrenal hyperplasia patients were reared female and one child with sex chromosome mosaicism was reared male. Eleven children had masculinizing genitoplasty for penoscrotal or perineal hypospadias (one-stage, three; two-stage, eight). Among one-stage surgeries, one child had meatal stenosis (minor) and one developed both urinary retention (minor) and urethrocutaneous fistula (major) (Summary Figure). Among two-stage surgeries, three children developed a major complication: penoscrotal fistula, glans dehiscence or urethral dehiscence. Among 16 children who had feminizing genitoplasty, vaginoplasty was performed in all, clitoroplasty in nine, external genitoplasty in 13, urethroplasty in four, perineoplasty in five, and total urogenital sinus mobilization in two. Two children had minor complications: one had a UTI, and one had both a mucosal skin tag and vaginal mucosal polyp. Two additional children developed a major complication: vaginal stenosis. Cosmesis scores revealed sustained improvements from 6 months post-genitoplasty, as previously reported, with all scores reported as good or satisfied. DISCUSSION: In these preliminary data from a multi-site, observational study, parents and surgeons were equally satisfied with the cosmetic outcomes 12 months after genitoplasty. A small number of patients had major complications in both feminizing and masculinizing surgeries; two-stage hypospadias repair had the most major complications. Long-term follow-up of patients at post-puberty will provide a better assessment of outcomes in this population. CONCLUSION: In this cohort of children with moderate to severe atypical genitalia, preliminary data on both surgical and cosmetic outcomes were presented. Findings from this study, and from following these children in long-term studies, will help guide practitioners in their discussions with families about surgical management.

Monoclonal antibodies to guinea pig interferon-gamma: tools for cytokine detection and neutralization.

We have generated polyclonal antisera and monoclonal antibodies against recombinant guinea pig IFN-gamma. These antibodies were used to inhibit the function of IFN-gamma in vitro and to establish a capture ELISA system for the detection and quantitation of this cytokine. Although recombinant protein expressed in E. coli was available in abundance, it was only of limited value to develop a capture ELISA which detects the native cytokine, since only a limited number of monoclonal antibodies reacted both with the recombinant and the native protein. Positive test results in an initial ELISA setup with recombinant IFN-gamma were not predictive for the detection of IFN-gamma from activated T-lymphocytes in the same assay. After evaluating several different combinations of rabbit antisera and monoclonal antibodies, an assay system was established which uses two mouse monoclonal antibodies as capture and detecting reagents. Three of the monoclonal antibodies and the rabbit antisera were able to block the function of guinea pig IFN-gamma when assayed in a luciferase reporter assay.

Prospective assessment of cosmesis before and after genital surgery.

INTRODUCTION: Little data exist about the surgical interventions taking place for children with disorders of sex development (DSD). Most studies that have evaluated cosmetic outcomes after genitoplasty have included retrospective ratings by a physician at a single center. OBJECTIVE: The present study aimed to: 1) describe frequency of sex assignment, and types of surgery performed in a cohort of patients with moderate-to-severe genital ambiguity; and 2) prospectively determine cosmesis ratings by parents and surgeons before and after genital surgery. STUDY DESIGN: This prospective, observational study included children aged <2 years of age, with no prior genitoplasty at the time of enrollment, moderate-to-severe genital atypia, and being treated at one of 11 children's hospitals in the United States of America (USA). Clinical information was collected, including type of surgery performed. Parents and the local pediatric urologist rated the cosmetic appearance of the child's genitalia prior to and 6 months after genitoplasty. RESULTS: Of the 37 children meeting eligibility criteria, 20 (54%) had a 46,XX karyotype, 15 (40%) had a 46,XY karyotype, and two (5%) had sex chromosome mosaicism. The most common diagnosis overall was congenital adrenal hyperplasia (54%). Thirty-five children had surgery; 21 received feminizing genitoplasty, and 14 had masculinizing genitoplasty. Two families decided against surgery. At baseline, 22 mothers (63%), 14 fathers (48%), and 35 surgeons (100%) stated that they were dissatisfied or very dissatisfied with the appearance of the child's genitalia. Surgeons rated the appearance of the genitalia significantly worse than mothers (P < 0.001) and fathers (P ≤ 0.001) at baseline. At the 6-month postoperative visit, cosmesis ratings improved significantly for all groups (P < 0.001 for all groups). Thirty-two mothers (94%), 26 fathers (92%), and 31 surgeons (88%) reported either a good outcome, or they were satisfied (see Summary Figure); there were no significant between-group differences in ratings. DISCUSSION: This multicenter, observational study showed surgical interventions being performed at DSD centers in the USA. While parent and surgeon ratings were discordant pre-operatively, they were generally concordant postoperatively. Satisfaction with postoperative cosmesis does not necessarily equate with satisfaction with the functional outcome later in life. CONCLUSION: In this cohort of children with genital atypia, the majority had surgery. Parents and surgeons all rated the appearance of the genitalia unfavorably before surgery, with surgeons giving worse ratings than parents. Cosmesis ratings improved significantly after surgery, with no between-group differences.

Increased expression of type 2 3alpha-hydroxysteroid dehydrogenase/type 5 17beta-hydroxysteroid dehydrogenase (AKR1C3) and its relationship with androgen receptor in prostate carcinoma.

Type 2 3alpha-hydroxysteroid dehydrogenase (3alpha-HSD) is a multi-functional enzyme that possesses 3alpha-, 17beta- and 20alpha-HSD, as well as prostaglandin (PG) F synthase activities and catalyzes androgen, estrogen, progestin and PG metabolism. Type 2 3alpha-HSD was cloned from human prostate, is a member of the aldo-keto reductase (AKR) superfamily and was named AKR1C3. In androgen target tissues such as the prostate, AKR1C3 catalyzes the conversion of Delta(4)-androstene-3,17-dione to testosterone, 5alpha-dihydrotestosterone to 5alpha-androstane-3alpha,17beta-diol (3alpha-diol), and 3alpha-diol to androsterone. Thus AKR1C3 may regulate the balance of androgens and hence trans-activation of the androgen receptor in these tissues. Tissue distribution studies indicate that AKR1C3 transcripts are highly expressed in human prostate. To measure AKR1C3 protein expression and its distribution in the prostate, we raised a monoclonal antibody specifically recognizing AKR1C3. This antibody allowed us to distinguish AKR1C3 from other AKR1C family members in human tissues. Immunoblot analysis showed that this monoclonal antibody binds to one species of protein in primary cultures of prostate epithelial cells and in LNCaP prostate cancer cells. Immunohistochemistry with this antibody on human prostate detected strong nuclear immunoreactivity in normal stromal and smooth muscle cells, perineurial cells, urothelial (transitional) cells, and endothelial cells. Normal prostate epithelial cells were only faintly immunoreactive or negative. Positive immunoreactivity was demonstrated in primary prostatic adenocarcinoma in 9 of 11 cases. Variable increases in immunoreactivity for AKR1C3 was also demonstrated in non-neoplastic changes in the prostate including chronic inflammation, atrophy and urothelial (transitional) cell metaplasia. We conclude that elevated expression of AKR1C3 is highly associated with prostate carcinoma. Although the biological significance of elevated AKR1C3 in prostatic carcinoma is uncertain, AKR1C3 may be responsible for the trophic effects of androgens and/or PGs on prostatic epithelial cells.

Influence of DNA superstructural features and histone amino-terminal domains on nucleosome positioning.

Nucleosome positioning has been studied on a strongly curved 268 bp DNA fragment from a Crithidia fasciculata kinetoplast, complexed with a histone octamer either normal or lacking amino-terminal domains. A very similar nucleosome multiple positioning, with the same rotational phasing, has been found, by Exo III mapping, in both cases. The experimental positioning is in fairly good agreement with that predicted using a theoretical method based on DNA distortion energy, derived from the nucleotide sequence. Taking into account that nucleosomes, without histone amino-terminal domains, lack thirty percent of electrostatic interactions, these results suggest a dominant role on nucleosome positioning of DNA distortion energy with respect to modifications in histone domains.

Small intestinal submucosa: a rapidly resorbed bioscaffold for augmentation cystoplasty in a dog model.

The extracellular matrix (ECM) of porcine small intestinal submucosa (SIS) has been shown to serve as a resorbable scaffold for tissue repair and remodeling in several body locations including the urinary bladder. The rate of resorption and extent of SIS degradation are unknown. Nine dogs were divided into three equal groups. Approximately 40% of the anterior dome of the urinary bladder was resected in each dog and replaced with porcine SIS. One group of dogs was sacrificed at each of 4, 8, and 12 weeks after surgery and the fate of the implanted SIS determined by immunohistochemical methods using a monoclonal antibody specific for porcine-derived SIS. By 4 weeks after surgery, only scattered remnants of SIS were present in the remodeled urinary bladder and these positively staining foci were surrounded by an extensive new host derived ECM and neovascularization. There was a continuous layer of transitional epithelium on the luminal surface by 4 weeks. No evidence for the originally implanted SIS could be found at either 8 or 12 weeks and bundles of organized smooth muscle cells were present at the operative site. In summary, SIS is rapidly and extensively degraded when used as a bioscaffold for augmentation cystoplasty in the dog model.

True hermaphroditism and normal male genitalia: an unusual presentation.

True hermaphroditism, the rarest form of intersex, is usually diagnosed during the newborn period in the course of evaluating ambiguous genitalia. As an exception, we present an unusual case of a 15-year-old boy with phenotypically normal male genitalia and bilaterally descended ovotestes, who was seen for evaluation of intermittent scrotal swelling and pain. Although rare, true hermaphroditism should be included in the differential diagnosis of apparently normal males presenting with similar complaints.

Percutaneous nephrolithotomy directly through an angiomyolipoma.

We present a patient with tuberous sclerosis and bilateral angiomyolipomas with a right partial staghorn calculi in which the calculi was managed with a percutaneous nephrolithotomy. Despite the inherent risk of hemorrhage with a percutaneous approach compounded by the fact that this was done directly through a tumor, we were able to render the patient stone free with no intraoperative bleeding, complications, or the need for postoperative blood transfusion. To our knowledge, this is the first reported case of percutaneous nephrolithotomy directly through a renal angiomyolipoma.

Penile orthoplasty using dermal grafts in the outpatient setting.

OBJECTIVES: To review and evaluate our extensive outpatient experience in the use of dermal grafts for the correction of significant residual chordee in patients requiring penile orthoplasty. METHODS: The medical records of all patients requiring surgical correction for severe penile curvature over the past 5 years were reviewed. Among these, 51 patients were identified in whom dermal grafts were used to correct severe residual chordee. Of these patients, 41 had severe hypospadias, 9 had epispadias, and 1 had urethral atresia. Graft harvest and placement technique, any resulting complications, and parental satisfaction with cosmetic and functional results were all evaluated. RESULTS: The dermal grafts were harvested from the hairless skin of the inguinal area, using one of two techniques. All were used to fill a tunical defect created by incising the tunica at the point of maximal penile curvature. No complications or postoperative hospitalizations resulted from any of the orthoplasty procedures. At a mean follow-up of 27 months, all patients had excellent cosmetic and functional results. CONCLUSIONS: In patients with significant residual chordee, a dermal graft is our preferred method of penile orthoplasty. This technique provides a straight phallus with a very minimal complication rate, it can easily and successfully be performed as an outpatient procedure, and it has assured an excellent cosmetic and functional result.

Wilms' tumor with renal cell carcinoma overgrowth in a 3-year-old child.

Wilms' tumors that contain features of both renal cell carcinoma and classic Wilms' tumor histology are rare. Even though nine such cases have been previously reported in the literature, we report the first case of a Wilms' tumor with an overwhelmingly renal cell carcinoma histologic pattern.

Rabbit urethral regeneration using small intestinal submucosa onlay grafts.

OBJECTIVES: To determine if small intestinal submucosa (SIS) can evoke urethral regeneration. METHODS: Twenty male white New Zealand rabbits were assigned to one of three experimental groups. Group 1 (n = 4) underwent simple urethrotomy and closure. Group 2 (n = 8), a second control group, underwent an onlay urethroplasty with a graft of full-thickness preputial skin from the host rabbit. Group 3 (n = 8) underwent an onlay urethroplasty with an SIS graft. RESULTS: All eight SIS onlay grafts promoted regeneration of the normal rabbit epithelium supported by a well-vascularized collagen and smooth muscle backing. Preputial free onlay grafts maintained a keratinizing squamous cell epithelium with a poor supportive backing, which resulted in the formation of urethral diverticulum. CONCLUSIONS: SIS onlay patch grafts for urethroplasty promote rabbit urethral regeneration.

Experimental assessment of small intestinal submucosa as a bladder wall substitute.

OBJECTIVES: This study determined the feasibility of promoting urinary bladder regeneration with porcine-derived small intestinal submucosa (SIS). METHODS: Twenty-two Sprague-Dawley rats underwent partial cystectomy with immediate bladder augmentation with SIS. Bladders were harvested for histologic evaluation at 2, 4, 8, 12, 24, and 48 weeks. RESULTS: Histologically at 2 weeks, there was infiltration of the graft material with viable host cells consisting of fibroblasts, macrophages, and blood vessels covered by complete mucosal urothelium comprised of transitional cells. During the next 10 weeks, collagen formation and maturation were noted, and by the end of 12 weeks, the SIS graft was comprised of a mature collagen matrix admixed with thinly scattered disorganized smooth muscle bundles and covered by normal urothelium. At 48 weeks, all three layers of the normal bladder (urothelium, smooth muscle, and serosa) were present and were grossly and microscopically indistinguishable from the normal rat urinary bladder. CONCLUSIONS: This study further supports the concept of bladder regeneration and suggests that SIS may be a viable material for bladder augmentations.

Influence of DNA superstructural features and histones aminoterminal domains on mononucleosome and dinucleosome positioning.

Mononucleosome and dinucleosome positioning was studied in the complexes between two DNA fragments of different lengths, both containing a strongly curved sequence from Crithidia fasciculata kinetoplast, and histone octamers either normal or lacking aminoterminal domains. The results obtained by Exo III and DNase I selective digestion were: (a) The first and most stable nucleosome, formed with both types of histone octamers, is positioned on the curved sequence, showing a multiple dyad axis translational positioning with the same rotational phasing. This result is in very good agreement with the theoretical prediction, obtained by adopting a method developed by us, based on the evaluation of DNA distortion energy from the nucleotide sequence. (b) The second nucleosome has two main different positions. The first one, near the extremity of the DNA fragment opposite to the curved sequence, presents a higher frequency in the case of normal nucleosome, whereas an intermediate position appears populated with a higher frequency in the case of the "tailless' nucleosome.

Evaluation and treatment of children with neurogenic bladders.

The evaluation and treatment of children with neurogenic bladders can be difficult because of the complexity of the neurologic deficit and the subjectivity of the history and physical exam. The primary emphasis of the physicians caring for these children should be to preserve renal function and facilitate continence when possible. As knowledge of both normal and abnormal lower urinary-tract dynamics increases, so does the ability to care for children with abnormal bladder dynamics caused by various neurologic conditions. With recent advancements in medical and surgical treatment of the neurogenic bladder, most children can maintain adequate renal function and attain urinary continence. This article reviews the pertinent innervation, anatomy, and physiology of the lower urinary tract, and discusses current evaluation and treatment of children with neurogenic bladders.

A Study on the Phylogeny of the Dyer's Woad Rust Fungus and Other Species of Puccinia from Crucifers.

ABSTRACT The identity of a Puccinia species occurring on the introduced weed dyer's woad (Isatis tinctoria) was studied using sequences from the internal transcribed spacer of the nuclear ribosomal DNA. The relationship of this fungus to other Puccinia species occurring on the family Brassicaceae in Europe and North America was examined, and we tested the hypothesis that P. thlaspeos and P. monoica are correlated species. The data suggest that the Puccinia species from dyer's woad is closely related to the North American species P. consimilis and may be derived from an indigenous strain of P. consimilis that switched hosts. Thus, the Puccinia species from dyer's woad is probably native to North America and is unlikely to cause disease epidemics on indigenous plants if used as a biological control agent against dyer's woad. P. thlaspeos appears to be polyphyletic and, therefore, P. thlaspeos and P. monoica do not appear to be correlated species. Additional DNA sequence data will be needed to clarify further the phylogeny of Puccinia species on the family Brassicaceae.

Seedborne fungal contamination: consequences in space-grown wheat.

Plants grown in microgravity are subject to many environmental stresses that may promote microbial growth and result in disease symptoms. Wheat (cv. Super Dwarf) recovered from an 8-day mission aboard a NASA (National Aeronautics and Space Administration) space shuttle showed disease symptoms, including girdling of leaf sheaths and chlorosis and necrosis of leaf and root tissues. A Neotyphodium species was isolated from the seed and leaf sheaths of symptomatic wheat used in the spaceflight mission. Certain isozymes of a peroxidase unique to extracts from the microgravity-grown plants were observed in extracts from earth-grown Neotyphodium-infected plants but were not present in noninfected wheat. The endophytic fungus was eliminated from the wheat seed by prolonged heat treatment at 50 degrees C followed by washes with water at 50 degrees C. Plants from wheat seed infected with the Neotyphodium endophyte were symptomless when grown under greenhouse conditions, whereas symptoms appeared after only 4 days of growth in closed containers. Disease spread from an infected plant to noninfected plants in closed containers. Dispersion via spores was found on asymptomatic plants at distances of 7 to 18 cm from infected plants. The size and shape of the conidia, mycelia, and phialide-bearing structures and the ability to grow rapidly on carbohydrates, especially xylose, resembled the characteristics of N. chilense, which is pathogenic on orchard grass, Doctylis glomerati. The Neotyphodium wheat isolate caused disease symptoms on other cereals (wheat cv. Malcolm, orchard grass, barley, and maize) grown in closed containers.

Bioengineering organs using small intestinal submucosa scaffolds: in vivo tissue-engineering technology.

Tissue engineering is a promising field of research that has the potential to revolutionize urinary bladder reconstruction. Currently, two techniques for the induction of bladder regeneration are being researched. The first, the in vivo technique, involves the use of a biodegradable scaffold that the host bladder can use to remodel and regenerate. This technique takes advantage of the cell's natural ability to heal or regenerate itself back to a normal state. The second technology, the in vitro technique, involves establishment of primary cell cultures from the host's native bladder. These cells are seeded on a biodegradable scaffold to create a composite graft that is then transplanted back into the host for continuation of the regeneration process. Clearly, both techniques have advantages and disadvantages, and both will have some role in future urinary reconstruction. To date, the most successful results utilizing in vivo techniques have been with small intestinal submucosa (SIS). In this article, we discuss in vivo tissue engineering technology and the preclinical studies that have been performed utilizing SIS for urinary tract regeneration.

The Value of PIC Cystography in Detecting De Novo and Residual Vesicoureteral Reflux after Dextranomer/Hyaluronic Acid Copolymer Injection.

The endoscopic injection of Dx/HA in the management of vesicoureteral reflux (VUR) has become an accepted alternative to open surgery. In the current study we evaluated the value of cystography to detect de novo contralateral VUR in unilateral cases of VUR at the time of Dx/HA injection and correlated the findings of immediate post-Dx/HA injection cystography during the same anesthesia to 2-month postoperative VCUG to evaluate its ability to predict successful surgical outcomes. The current study aimed to evaluate whether an intraoperatively performed cystogram could replace postoperative studies. But a negative intraoperative cystogram correlates with the postoperative study in only 80%. Considering the 75-80% success rate of Dx/HA implantation, the addition of intraoperative cystograms cannot replace postoperative studies. In patients treated with unilateral VUR, PIC cystography can detect occult VUR and prevent postoperative contralateral new onset of VUR.