Extracellular vesicles derived from human foreskin cells (hFS-Exo) accelerate cell migration and angiogenesis through MAPK pathway: an in vitro study.

BACKGROUND: Wound healing is one of the important processes in the body. Attempts to create new drugs are of interest due to the side effects of natural and chemical wound healing compounds. To overcome this obstacle, stem cells have been used as healing agents. However, both difficulties in collection and risks such as rejection and teratoma in the recipient body have limited the use of stem cells, directly. Since the potential content of the stem cells can be transferred to the recipient cells by vesicles, small extracellular vesicles have recently become prominent agents. METHODS AND RESULTS: The wound-healing effect of extracellular vesicles derived from foreskin cells was investigated in both keratinocyte and endothelial cells. Migration assay, RT-PCR, Col1a1 ELISA and Western Blot experiments were utilized to reveal healing effect of EVs and its possible molecular pathways. EV-treated groups exhibited more proliferative, invasive, and migrative characteristics. When comparing to the control group, new vessel formation was induced in EV groups. An increase in gene levels of growth factors related to wound healing and change in the mitogen-activated protein kinase (MAPK) signaling pathway proteins in EV-treated groups were determined. Possible molecular mechanisms underlying cell movements were associated with the MAPK pathway. It was found that human foreskin cell EVs (hFS-Exo) may have a potential to heal wounds in a short period of time by triggering the MAPK pathway. CONCLUSIONS: hFS-Exo could be a new promising wound healing agent in the future.

Histological and morphological development of the prepuce from birth to prepubertal age.

PURPOSE: To study the histological changes of the preputial tissue from birth to prepubertal age in order to define unnoticed morphological changes. MATERIALS AND METHODS: Prepuce samples were obtained from 79 healthy boys who underwent routine ritual circumcision. Specimens were divided into six groups according to the boys' age: newborn, 0-1 year of age, 2-3 years of age, 4-5 years of age, 6-7 years of age, and 8-9 years of age. Histologic analysis of the specimens was performed by H&E, Masson's trichrome, Verhoeff-Von Gieson, immunohistochemical staining. RESULTS: Microscopic examinations showed that average epithelial thickness increased after the neonatal period (p=0.001). When collagen fiber density was evaluated, no significant differences between groups were found (p=0.083). When the elastic fibers in the dermis were evaluated, it was determined that the number and thickness of elastic fibers increased with age. Immunohistochemical examinations showed that the number of peripheral nerves marked with S100 was lower in the neonatal period than at other ages (p=0.048). When the vessels marked with CD105 antibody were counted, there was no significant difference between the groups (p=0.078). CONCLUSIONS: This is the first study to examine the age-related structure of connective tissue elements in the foreskin. Our results showed that the prepuce's prepubertal maturation process is continuous, and the first 2 years of life are appropriate not only in relation to the physiological effects of age but also the optimum structural changes for wound healing, such as vessel diameter, epithelium thickness, peripheral nerve count.

Phimosis in Adults: Narrative Review of the New Available Devices and the Standard Treatments.

BACKGROUND: Phimosis is the inability to completely retract the foreskin and expose the glans. The treatment of phimosis varies depending on the age of the patient and the severity of the disease; a great number of conservative or surgical treatments are currently available. AIM: To provide the first review summarizing the available options for the treatment of adult phimosis. METHODS: A PubMed, Cochrane and Embase search for peer-reviewed studies, published between January 2001 and December 2022 was performed using the search terms "phimosis AND treatment". RESULTS: A total of 288 publications were initially identified through database searching. Thirty manuscripts were ultimately eligible for inclusion in this review. Conservative treatment is an option. and it includes topical steroid application and the new medical silicon tubes (Phimostop™) application for gentle prepuce dilation. Concerning the surgical approach, the gold-standard treatment is represented by circumcision in which tissue synthesis after prepuce removal can be also obtained with barbed sutures, fibrin glues or staples. Laser circumcision seems to be providing superior outcomes in terms of operative time and postoperative complication rate when compared to the traditional one. Several techniques of preputioplasty and use of in situ devices (which crush the foreskin and simultaneously create haemostasis) have been also described. These in situ devices seem feasible, safe and effective in treating phimosis while they also reduce the operative time when compared to traditional circumcision. Patient satisfaction rates, complications and impact on sexual function of the main surgical treatments are presented. CONCLUSION: Many conservative and surgical treatments are available for the treatment of adult phimosis. The choice of the right treatment depends on the grade of phimosis, results, complications, and cost-effectiveness.

Can paediatric surgical registrars safely perform supervised hypospadias surgery?

INTRODUCTION: Hypospadias repair is regarded as a technically demanding, complex procedure, with variable outcomes. Therefore, it tends to be performed by consultants, with limited trainee involvement. We aimed to study the clinical outcomes of supervised registrars performing proximal and distal hypospadias repairs, compared to their consultant mentors. METHODS: We undertook a retrospective review of all primary hypospadias repairs performed between April 2013-April 2022 at our tertiary paediatric urology centre. Redo repairs and patients lost to follow-up were excluded. Pre-operative anatomy, theatre time, grade of primary surgeon (registrar (trainees and non-training middle grades) or consultant), operative technique, follow-up duration, complications, and reoperation rates were recorded. The procedures were assessed in two groups according to the primary operator: registrar or consultant. The Zwisch scale is used to describe level of consultant support. Registrars as primary operators received "passive help" or "supervision" (Zwisch levels 3/4). Consultants as primary operators provided registrars with "show-and-tell" or "active help" (Zwisch levels 1/2). RESULTS: 270 procedures performed on 228 patients met the inclusion criteria. 109 were performed by registrars and 161 by consultants. In both groups, median age was two years (p = 0.23). Median theatre time was similar (registrars 2.8 h vs. consultants 2.7 h, p = 0.88), as was median follow-up (registrars 25months, vs. consultants 21months, p = 0.99). Operations performed by registrars were 76% distal and 24% proximal; and by consultants were 62% distal and 38% proximal. The overall urethroplasty complication rate was similar, at 24% for registrars and 23% for consultants (p = 0.89). The summary table shows the distribution of different complications. Re-operation rate was 16% in both groups (p = 0.99). Complications were further assessed according to operation type (TIP vs. two-stage repair). DISCUSSION: Contrary to popular belief amongst hypospadiologists, we found complication rates were similar for registrar and consultant surgeons. We question that involvement of registrars increases complications. The literature demonstrates safety of trainee performance of limited steps of the procedure. However our institution permits registrars to perform up to the whole hypospadias repair under direct supervision, with no predefined limit to their involvement. CONCLUSION: Paediatric surgical registrars can be safely supervised to have substantial involvement in proximal and distal hypospadias repair, without compromising the duration or outcomes of surgery. We hope that allowing more registrar involvement can lead to faster acquisition of surgical skills, whilst remaining under the safety of senior supervision. Increasing opportunities for those with an aptitude for hypospadias repair can equip them with skills and confidence for entering fellowship training.

Long-term outcomes of foreskin reconstruction in distal hypospadias; a cohort study spanning twenty years.

INTRODUCTION: Surgical correction of hypospadias aims to achieve normal functionality and appearance. This entails foreskin reconstruction (FR) in countries where the uncircumcised penis constitutes the norm. Long-term data are however scarce. OBJECTIVE: To investigate the long-term outcome of FR in cohort of patients operated for distal hypospadias combined with approximately 20 years after surgery. METHODS: The hospital management system was searched for patients operated for distal hypospadias in conjunction with FR between 1997 and 2004. Prospective participants were invited to participate in an online questionnaire. Signed consent allowed for extended medical chart review, with regards to hypospadias grade, surgical procedure and complications. RESULTS: Response rate of 44.6 %. For 113 participants, median age at primary surgery was 5.2 (1.0-15.5) years. Two-thirds had a distal meatus while the remaining, meatus was mid to distal shaft. Urethroplasties performed were mainly glanular approximation procedures and meatal based flap procedures in 85 %. Foreskin fistula developed in 15 % of cases. There was no significant relationship between urethroplasty procedure or meatal position and risk of foreskin complications. Three layer closure of foreskin resulted in significantly less complications than two layer closure. Twenty years on 95 % of the men still had an intact foreskin, of whom 16.8 % had received treatment for phimosis. Foreskin was retractable in 92.5 % and 74.7 % in the flaccid and erect states respectively. Ninety intact men had had their sexual debut and in those 23.3 % reported foreskin related issues with intercourse. Evolution of foreskin retractability can be seen in the figure. DISCUSSION: Current results show that three layer FR in conjunction with hypospadias surgery is feasible and that short-term complication rates were comparable with what has previously been published in the literature. Long-term results indicate that FR is durable with regards to anatomical reconstruction however foreskin function especially in relation to sexual function was compromised in about 25 %. Foreskin retractability after surgery predicted retractability in adulthood for the flaccid but not erect penis. Limitations of this study include the retrospective nature of data collection, and that the questionnaire used was not validated. We however achieved a decent response rate and were able to capture important long-term data. CONCLUSIONS: FR has an acceptable complication rate. Long-term results two decades on are remarkably durable with regards to the anatomical preservation of the prepuce, however functionality was compromised with regards to retractability and sexual function in approximately 25 %.

[Balanoposthitis in the clinical practice of a urologist and dermatovenereologist: an interdisciplinary problem].

Balanoposthitis is a common inflammatory disease of the male genitals, which occurs more often in uncircumcised men. The cause of balanoposthitis can be an infectious, inflammatory or autoimmune process, as well as traumatization. In most cases, after proper intimate hygiene and the use of neutral moisturizers, the symptoms of balanoposthitis are completely stopped. In the case of torpid course of balanoposthitis and in the absence of improvement after drug therapy, it is necessary to exclude the malignant process. In the review article, the authors present the data of modern scientific research on the qualitative and quantitative composition of the microbiome in balanoposthitis. Differences in the composition of the microbiome were revealed in patients with balanoposthitis and healthy patients from the control group with excess foreskin. It was found that in patients with balanoposthitis, a impaired in hydration of the skin of the glans penis was revealed. Staphylococcus warneri and Prevotella bivia are the most common species associated with balanoposthitis and positively correlate with the severity of the disease. Candida infection, as an etiological factor of balanoposthitis, often occurs in children and may be associated with diaper rash. The prevalence of Gardnerella vaginalis as a pathogen in the male urogenital tract has not been fully studied. Currently, there are no reliable scientific studies that make it possible to attribute G. vaginalis to the etiological factor of balanoposthitis in men. However, it should be borne in mind that balanoposthitis may have a polymicrobial and synergistic etiology with the participation of G. vaginalis and anaerobic bacteria in the lower genital tract of men. The review article is clearly illustrated with clinical examples of the disease from the personal practice of the authors.

Prevalence of urogenital disease in male adolescent outpatient visits. A descriptive, cross-sectional study. / Prevalencia de patología urogenital en la consulta ambulatoria de varones adolescentes. Estudio descriptivo transversal.

Introduction. The male genital exam is a simple and quick assessment to look for urogenital disease. Data on the prevalence of urogenital disease in male adolescents are limited. Our objective was to describe the prevalence of urogenital disease in male adolescents. Population and methods. Descriptive, cross-sectional study conducted at the Department of Adolescence of a public hospital in the City of Buenos Aires. The medical records of male patients aged 9 to 20 years seen between 2008 and 2018 were retrospectively reviewed; all those with a genital exam were included. Data on age, pubertal stage, orchidometry, and urogenital disease before adolescence and at the time of consultation were recorded. The prevalence was described as percentage and 95% confidence interval (CI). As per estimations, 1167 medical records had to be included to establish the population sample. Results. A total of 2129 medical records were assessed and 1429 were included. No genital exam had been conducted in 686 cases. The median age of the population was 12 years (interquartile range: 11-14 years). Urogenital disease before adolescence was detected in 72 boys (5.7%; 95% CI: 4.5- 7.2). Urogenital disease was found in 272 adolescents (14.8%; 95% CI: 13.1-16.7); the most common conditions were balanopreputial adhesions in 5.3% (95% CI: 4.2-6.6), varicocele in 2.7% (95% CI: 2-3.7), and phimosis in 1.8% (95% CI: 1.2-2.6). Conclusions. A genital exam allowed to detect that 14.8% of adolescent boys had a urogenital disease. The most common conditions were balanopreputial adhesions, varicocele, and phimosis.

Introducción. El examen genital en varones es una evaluación clínica simple y rápida para detectar patología urogenital. Los datos sobre prevalencia de patología urogenital en varones adolescentes son limitados. Nuestro objetivo fue describir la prevalencia de patología urogenital en varones adolescentes. Población y métodos. Estudio descriptivo transversal en el Servicio de Adolescencia de un hospital público de la Ciudad de Buenos Aires. Se evaluaron retrospectivamente las historias clínicas (HC) de varones de 9 a 20 años atendidos entre 2008 y 2018; se incluyeron las que tenían examen genital. Se recabaron datos de edad, estadio puberal, orquidometría, patología urogenital antes de la adolescencia y al momento de la consulta. La prevalencia se expresó en porcentaje e intervalo de confianza del 95 % (IC95%). Se estimó necesario incluir 1167 HC como muestra poblacional. Resultados. Se evaluaron 2129 HC; se incluyeron 1429. En 686 casos no se hizo el examen genital. La población tuvo una mediana de edad de 12 años (rango intercuartílico 11-14 años). En 72 varones (5,7 %; IC95% 4,5-7,2), se halló una enfermedad genitourinaria antes de la adolescencia. Se detectó al menos una patología urogenital en 272 adolescentes (14,8 %; IC95% 13,1-16,7); las más frecuentes fueron adherencia balanoprepucial 5,3 % (IC95% 4,2-6,6), varicocele 2,7 % (IC95% 2-3,7) y fimosis 1,8 % (IC95% 1,2-2,6). Conclusiones. El examen genital permitió detectar que el 14,8 % de los varones adolescentes atendidos presentó alguna patología urogenital. Las entidades más frecuentes fueron adherencia balanoprepucial, varicocele y fimosis.

Unusual presentation of penile giant condyloma acuminatum with spontaneous prepuce perforation: A case report.

BACKGROUND: Giant condyloma acuminatum (GCA) is an uncommon condition affecting the anogenital area. CASE SUMMARY: Here, we report an 88-year-old male patient who presented with a 5-year history of a progressive appearance of multiple cauliflower-like warts over his penile shaft, pubis region, and groin as well as urine leakage along the distal penile shaft. Physical examination revealed an ulcerative skin perforation with pus discharge under the distal prepuce base, which was initially suspected to be a urethral fistula. However, during surgery, it was discovered that the perforation was caused by a giant condyloma lesion that had obliterated the prepuce opening, with infection and high pressure causing subsequent skin perforation. He underwent circumcision and wide excision with electrocauterization of the warts. He was discharged after the surgery, and the residual lesion was treated with imiquimod and low-dose oral tegafur-uracil. CONCLUSION: Penile GCA can cause prepuce perforation and can be postoperatively treated with imiquimod and low-dose oral tegafur-uracil.

Correction of curvature in single stage hypospadias repair with foreskin reconstruction.

INTRODUCTION: Many surgeons offer foreskin reconstruction (FR) as a routine part of hypospadias repair. We present a step-by-step video of the procedure of Tubularised Incised Plate (TIP) repair, FR and dorsal plication through a ventral skin incision. MATERIALS AND METHODS: A ventral incision is made between the inner preputial mucosa and the outer skin extending below the meatus. Ventral degloving is carried out. The dissection is extended laterally around the corporal bodies. The point of maximal curvature (PMC) is marked on the dorsal midline. A vertical incision is made and closed transversely with 5-0 prolene suture in a Heineke- Mikulicz fashion. Urethroplasty is performed in 2 layers using 7-0 polydioxanone (PDS). Spongioplasty and ventral dartos are used as barrier layers. Glansplasty is performed in 2 layers.FR is carried out in 3 layers. DISCUSSION: Curvature correction is key to good outcome. Dorsal degloving can be achieved through a ventral incision allowing exposure of the dorsal midline for plication sutures. RESULTS: The patient had good cosmetic and functional outcome at 1 month follow up. CONCLUSION: FR can be safely performed during TIP repair for distal hypospadias repair. Curvature of less than 30° can be corrected through a ventral incision only.

Xeno-free generation of new Yazd human embryonic stem cell lines (Yazd4-7) as a prior stage toward good manufacturing practice of clinical-grade raw materials from discarded embryos: A lab resources report.

Background: Xeno-free generation of human embryonic stem cells (hESCs) is important to prevent potential animal contaminations in culture for advanced cell-based therapeutic applications. Xeno-free production of hESCs is the first step for manufacturing clinical-grade hESC lines. Objective: To produce new hESC lines in xeno-free condition. Materials and Methods: This lab resources report was conducted at Stem Cell Biology Research Center, Yazd, Iran from 2019-2022. 4 new hESC lines from 11 (10 fresh and 1 frozen) donated surplus discarded human embryos were established. In this study, we report the xeno-free derivation of new Yazd hESC lines (Yazd4-7), without using immunosurgery, by culturing intact zona-free blastocysts obtained from discarded embryos onto the YhFF#8 cells as a feeder layer in a microdrop culture system. The pluripotency gene expression profile of the cell lines was assessed by reverse transcription polymerase chain reaction and the expression of specific surface markers was detected using immunofluorescent staining. In vitro differentiation was induced using embryoid body formation and gene expression profile of 3 germ layers and germ cells. Reverse transcriptase polymerase chain reaction was investigated to prove their pluripotent capacity. Results: In sum, we have been able to generate 4 new hESC lines (Yazd4-7) from 11 discarded embryos in xeno-free culture conditions using a micro drop culture system and YhFF#8 as a human source feeder layer. Conclusion: The outcome of this work can be the foundation for the future allogeneic cell-based therapeutic application using clinical grade good manufacturing practice-derived hESC derivatives.

Are there anatomical limiting factors to foreskin reconstruction at the time of single-stage hypospadias repair?

INTRODUCTION: Foreskin reconstruction (FR) at the time of primary hypospadias repair is a truly anatomically complete reconstruction of the hypospadic penis. We prospectively collected penile and preputial measurement of children undergoing single-stage hypospadias repair and FR with the aim to identify possible relations between penile and preputial anatomy and the likelihood to develop complications. MATERIALS AND METHODS: We prospectively studied children who underwent single stage hypospadias repair associated with FR from 2016 to 2019. We recorded intra-operative foreskin and penile measurements and post-operative outcomes. Logistic Regression analysis was performed to explore independent factors affecting urethroplasty and skin complications. Chi square test was used to compare outcomes in different groups based on ventral foreskin defect (VFD) width, Glans size, age at surgery and meatal location. RESULTS: From a total of 181 consecutive patients, 86 boys who underwent a single stage hypospadias repair combined with FR were included in the study. Patients were excluded because they were either lost at follow up (n = 10), required a 2-stage repair (n = 2), were circumcised at birth (n = 3) or parents requested a circumcision (n = 78); in 2 patients, a decision to perform circumcision was made intraoperatively due to aesthetic reasons (monk-hood deformity of the prepuce). Median age at surgery was 17 months. Mean glans width was 14.4 mm. Mean unstretched and stretched foreskin circumference were 29.5 mm and 40.9 mm, respectively. Mean VFD (the distance between the proximal insertion of the foreskin hood on either side of the midline at the level of the coronal sulcus) was 7.2 mm (Fig. 1). At median follow-up of 8 months (6-23), 9 complications were recorded (10.4%): foreskin dehiscence occurred in 1% (1/86), a foreskin fistula was noted in 4.6% (4/86), tight, non-retractile, foreskin in 1% (1/86); urethrocutaneous fistula in 2.3% (2/86) and complete dehiscence of the glans and foreskin in 1 (1.2%). Multiple logistic regression analysis demonstrated that none of the measurements obtained was an independent risk factor for developing urethroplasty or skin complications. There was no significant difference in complications between wide VFD (>7 mm) vs. narrow VFD (≤7 mm), large glans (>14 mm) vs. small glans (≤14 mm), age at surgery ≤24 months vs. > 24 month and meatal location distal (glanular, coronal, subcoronal and distal penile) vs. proximal (midpenile, proximal penile and penoscrotal). CONCLUSION: To the best of our knowledge, this is the first study reporting a prospective and objective assessment of the foreskin in the context of single stage hypospadias repair. Individual anatomical differences in preputial and penile anatomy do not seem to affect the likelihood of skin and urethroplasty complications. FR can, therefore, be offered to all boys undergoing primary single stage hypospadias repair . Further studies on larger numbers and external validation of these measurements is necessary.

Immunohistological Analysis of Lichen Sclerosus of the Foreskin in Pediatric Age: Could It Be Considered a Premalignant Lesion?

BACKGROUND: A major worry of juvenile penile LS is potential malignant degeneration to spinocellular carcinoma (SCC) in adulthood. LS is characterized by increased CD8+ and CD57+ cells, dermal sclerosis, epidermal atrophy, and hyperkeratosis. p53 and Ki67 are reliable premalignant markers. Our aim was to define the LS immunohistochemical profile of foreskin in children, focusing on tissue immune response and cell proliferation. METHODS: Thirty specimens of foreskins removed from pediatric patients during circumcision were included: six from ritual operation (A), twelve from phimosis (B), and twelve from phimosis with LS (C). Formalin-fixed paraffin-embedded sections were stained for histomorphology and immunohistochemistry. A quantitative evaluation for CD8, CD57, p53, and Ki-67 and a statistical analysis were performed. RESULTS: As compared to groups A and B, the samples from group C patients showed an acanthotic epidermis, a dermal band of lymphoid infiltrate with a significant enhancement of CD8+ CD57+ lymphocytes, and a keratinocytic hyperplasia with an overexpression of Ki67+ and p53+ cells. CONCLUSIONS: Immunohistological findings confirmed an immune reaction and proliferative behavior in juvenile LS of foreskin. We believe that radical circumcision should be the first treatment of choice in pediatric patients with clinical suspicious of LS for the potential risk of transformation to SCC in adulthood.

Cas phosphorylation regulates focal adhesion assembly.

Integrin-mediated cell attachment rapidly induces tyrosine kinase signaling. Despite years of research, the role of this signaling in integrin activation and focal adhesion assembly is unclear. We provide evidence that the Src-family kinase (SFK) substrate Cas (Crk-associated substrate, p130Cas, BCAR1) is phosphorylated and associated with its Crk/CrkL effectors in clusters that are precursors of focal adhesions. The initial phospho-Cas clusters contain integrin ß1 in its inactive, bent closed, conformation. Later, phospho-Cas and total Cas levels decrease as integrin ß1 is activated and core focal adhesion proteins including vinculin, talin, kindlin, and paxillin are recruited. Cas is required for cell spreading and focal adhesion assembly in epithelial and fibroblast cells on collagen and fibronectin. Cas cluster formation requires Cas, Crk/CrkL, SFKs, and Rac1 but not vinculin. Rac1 provides positive feedback onto Cas through reactive oxygen, opposed by negative feedback from the ubiquitin proteasome system. The results suggest a two-step model for focal adhesion assembly in which clusters of phospho-Cas, effectors and inactive integrin ß1 grow through positive feedback prior to integrin activation and recruitment of core focal adhesion proteins.

Respiratory viruses induce the expression of type I and III IFNs in MSCs through RLR/IRF3 signaling pathways.

Mesenchymal stem cells (MSCs) comprise a primitive cell population and reside in various tissues and organs. These cells exhibit immunomodulatory activity and are effective in treating respiratory viral infections. The activation of type I and III interferons, which protect cells against viral infections, can be induced after pattern recognition receptors (PRRs) recognize viral nucleic acid species. Although certain viruses can upregulate IFN-ß expression in MSCs, the underlying mechanisms and responsiveness to different IFNs are unclear. We found that foreskin-derived fibroblast-like stromal cells (FDSCs), a kind of functional MSC, were permissive to IAV PR8, HCoV-229E, and EV-D68. Infection by IAV PR8 and HCoV-229E increased the expression of IFN-ß and IFN-λ species in FDSCs in an IRF-3-dependent manner. RIG-I was critical for detecting IAV PR8 in FDSCs, and IAV PR8 infection induced a significant increase in the expression of interferon signaling genes (ISGs). Interestingly, only IFN-ß, but not IFN-λ species, could induce the expression of ISGs, a finding supported by our observation that only IFN-ß induced STAT1 and STAT2 phosphorylation in FDSCs. We also proved that treatment with IFN-ß suppressed the propagation of IAV PR8 and promoted the survival of virus-infected FDSCs. Respiratory viruses could infect FDSCs and induce the expression of IFN-ß and IFN-λ1, but only IFN-ß could protect FDSCs against viral infection.

hsa_circ_0000417 downregulation suppresses androgen receptor expression and apoptotic signals in human foreskin fibroblasts via sponging miR-6756-5p.

BACKGROUND: Dysregulated apoptosis of penile mesenchymal cells during male urethragenesis has been previously demonstrated to underly hypospadiac urethral closure failure, and androgen receptor (AR) has been shown to play a central role in regulating penile mesenchyme cell proliferation and survival. However, the regulatory mechanisms upstream and downstream of AR remain poorly understood. Our clinical data and bioinformatics analysis previously indicated that hsa_circ_0000417, a circRNA significantly downregulated in hypospadias preputial specimens, may act as a ceRNA for AR via sequestering hsa_miR-6756-5p, and that the biological functions of hsa_circ_0000417 may significantly involve the PI3K/AKT pathway. In this study, we employed human foreskin fibroblasts (HFF-1) to experimentally validate this putative hsa_circ_0000417/miR-6756-5p/AR axis and its impact on penile mesenchymal cell proliferation and apoptosis. METHOD AND RESULTS: We showed that hsa_circ_0000417 knockdown significantly promoted proliferation and suppressed apoptosis of HFF-1 cells. Mechanistically, hsa_circ_0000417 functioned as a molecular sponge for miR-6756-5p in HFF-1 cells and relieved the latter's translational repression on AR mRNA, leading to decreased AKT activation and increased expression of pro-apoptotic proteins BAX and cleaved-caspase 9. Conversely, elevated levels of miR-6756-5p resulted in diminished AR expression concomitant with enhanced AKT activation and HFF-1 cell proliferation. CONCLUSIONS: Collectively, our data describe for the first time a circRNA-mediated post-transcriptional regulatory mechanism of AR and its functional consequences in penile mesenchymal cells in the context of hypospadias. These findings may contribute to advancing our current understanding of the roles of AR and mesenchymal cell fate decisions during penile morphogenesis.

Dose finding study for on-demand HIV pre-exposure prophylaxis for insertive sex in sub-Saharan Africa: results from the CHAPS open label randomised controlled trial.

BACKGROUND: The efficacy of on-demand HIV pre-exposure prophylaxis (PrEP) for men in sub-Saharan Africa has not been evaluated, and the on-demand PrEP dosing requirement for insertive sex remains unknown. METHODS: HIV-negative males 13-24 years, requesting voluntary medical male circumcision (VMMC), were enrolled into an open-label randomised controlled trial (NCT03986970), and randomised 1:1:1:1:1:1:1:1:1 to control arm or one of eight arms receiving emtricitabine-tenofovir disoproxil fumarate (F/TDF) or emtricitabine-tenofovir alafenamide (F/TAF) over one or two days, and circumcised 5 or 21 h thereafter. The primary outcome was foreskin p24 concentrations following ex vivo HIV-1BaL challenge. Secondary outcomes included peripheral blood mononuclear cell (PBMC) p24 concentration, and drug concentrations in foreskin tissue, PBMCs, plasma and foreskin CD4+/CD4-cells. In the control arm, post-exposure prophylaxis (PEP) activity of non-formulated tenofovir-emtricitabine (TFV-FTC) or TAF-FTC was assessed with ex vivo dosing 1, 24, 48 or 72 h post-HIV-1 challenge. FINDINGS: 144 participants were analysed. PrEP with F/TDF or F/TAF prevented ex vivo infection of foreskins and PBMCs both 5 and 21 h after PrEP dosing. There was no difference between F/TDF and F/TAF (p24day15 geometric mean ratio 1.06, 95% confidence interval: 0.65-1.74). Additional ex vivo dosing did not further increase inhibition. In the control arm, PEP ex vivo dosing was effective up to 48 post-exposure diminishing thereafter, with TAF-FTC showing prolonged protection compared to TFV-FTC. Participants receiving F/TAF had higher TFV-DP concentrations in foreskin tissue and PBMCs compared with F/TDF, irrespective of dose and sampling interval; but F/TAF did not confer preferential TFV-DP distribution into foreskin HIV target cells. FTC-TP concentrations with both drug regimens were equivalent and ∼1 log higher than TFV-DP in foreskin. INTERPRETATION: A double dose of either F/TDF or F/TAF given once either 5 or 21 h before ex vivo HIV-challenge provided protection across foreskin tissue. Further clinical evaluation of pre-coital PrEP for insertive sex is warranted. FUNDING: EDCTP2, Gilead Sciences, Vetenskapsrådet.

[Development of circumcision rates in Germany since the approval of ritual circumcision : A population-based analysis from 2013 to 2018]. / Entwicklung der Zirkumzisionszahlen in Deutschland seit Billigung der rituellen Beschneidung : Eine bevölkerungsbezogene Analyse von 2013 bis 2018.

INTRODUCTION: The religious and cultural circumcision of male infants in Germany is controversially discussed. After the passing of the religious circumcision bill in 2012, an increase of infant circumcisions without medical indication was feared. The aim of this study was to analyze the development of the circumcision case numbers. MATERIALS AND METHODS: We used the research database of the German Institute for Applied Health Research with a representative anonymous sample of 4.9 million insured persons to estimate the annual circumcision numbers in Germany from 2013-2018. We stratified the data according to age (< 18 vs. ≥ 18 years). The number of male adolescents in the study period was taken from the database of the German Federal Statistical Office. RESULTS: In the study period, 673,819 circumcisions were performed. From 2014, there was a significant decrease in the number of cases across all age groups (p = 0.049). Thereby, circumcisions in minors significantly increased (p = 0.002) and procedures in adults significantly decreased (p = 0.01) during the entire study period. The number of male minors increased by 4% from 6,709,137 (2013) to 6,992,943 (2018). The corresponding population-based number increased from 7.5 circumcisions per 1000 minors in 2013 to 8 in 2018 (p = 0.037). CONCLUSIONS: After the passage of the circumcision bill in 2012, there was a significant increase of circumcisions in the age group of < 18 years in Germany. A major limitation of our study is that presumably many ritual circumcisions might not be provided within the health care system.

Experimental Support for Human Papillomavirus Genome Amplification Early after Infectious Delivery.

Even though replication and transcription of human papillomavirus type 16 (HPV16) has been intensively studied, little is known about immediate-early events of the viral life cycle due to the lack of an efficient infection model allowing genetic dissection of viral factors. We employed the recently developed infection model (Bienkowska-Haba M, Luszczek W, Myers JE, Keiffer TR, et al. 2018. PLoS Pathog 14:e1006846) to investigate genome amplification and transcription immediately after infectious delivery of viral genome to nuclei of primary keratinocytes. Using 5-ethynyl-2'-deoxyuridine (EdU) pulse-labeling and highly sensitive fluorescence in situ hybridization, we observed that the HPV16 genome is replicated and amplified in an E1- and E2-dependent manner. Knockout of E1 resulted in failure of the viral genome to replicate and amplify. In contrast, knockout of the E8^E2 repressor led to increased viral genome copy number, confirming previous reports. Genome copy control by E8^E2 was confirmed for differentiation-induced genome amplification. Lack of functional E1 had no effect on transcription from the early promoter, suggesting that viral genome replication is not required for p97 promoter activity. However, infection with an HPV16 mutant virus defective for E2 transcriptional function revealed a requirement of E2 for efficient transcription from the early promoter. In the absence of the E8^E2 protein, early transcript levels are unaltered and even decreased when normalized to genome copy number. Surprisingly, a lack of functional E8^E2 repressor did not affect E8^E2 transcript levels when normalized to genome copy number. These data suggest that the main function of E8^E2 in the viral life cycle is to control genome copy number. IMPORTANCE It is being assumed that human papillomavirus (HPV) utilizes three different modes of replication during its life cycle: initial amplification during the establishment phase, genome maintenance, and differentiation-induced amplification. However, HPV16 initial amplification was never formally proven due to a lack of an infection model. Using our recently established infection model (Bienkowska-Haba M, Luszczek W, Myers JE, Keiffer TR, et al. 2018. PLoS Pathog 14:e1006846), we demonstrate herein that viral genome is indeed amplified in an E1- and E2-dependent manner. Furthermore, we find that the main function of the viral repressor E8^E2 is to control viral genome copy number. We did not find evidence that it regulates its own promoter in a negative feedback loop. Our data also suggest that the E2 transactivator function is required for stimulation of early promoter activity, which has been debated in the literature. Overall, this report confirms the usefulness of the infection model for studying early events of the HPV life cycle using mutational approaches.

Efficacy and safety evaluation of the scrotal skin transfer method in the treatment of pediatric concealed penis: a six-month follow-up data.

Background: Pediatric concealed penis affects penis development and the psychological health of the children. Current surgical methods tend to retain too much of the inner foreskin plate, resulting in unsatisfactory appearance and postoperative complications. We aimed to investigate the efficacy and safety of a new surgical intervention method using a scrotal skin transfer instead of the external foreskin plate to treat pediatric concealed penis. Methods: Sixty children (aged between 3 and 9 years and 3 months) diagnosed with concealed penis admitted to our hospital between June 2020 and June 2022 were included. All patients were randomly divided into Improve group (n=30, treated with the new surgical intervention method of scrotal skin transfer) and General group (n=30, treated with traditional Shiraki procedure). The pre-operative and post-operative penis length were collected. A follow-up of 6 months was conducted in all patients. Postoperative penile appearance, penile skin color difference, and satisfaction with the penile appearance were used to evaluate the efficacy of surgery, and the postoperative complications (penile retraction, skin edema and incision infection) were collected to observe the safety of treatment. Results: The post-operative penis length of Improve group was significantly increased compared with General group. The penile skin color was consistent, the penile scrotal angle and the penile pubic angle were formed naturally, and the penis was completely exposed in Improve group. During 6 months' follow-up, the patients in the improvement group did not have infection, penile retraction and skin color difference, and 2 patients had edema. The complication rate in the Improve group was lower than that in the General group. Conclusions: The scrotal skin transfer method shows remarkable efficacy and safety without apparent complications and results in a penis that is fully exposed and aesthetically pleasing.