ROR2 expression predicts human induced pluripotent stem cell differentiation into neural stem/progenitor cells and GABAergic neurons.

Despite the development of various in vitro differentiation protocols for the efficient derivation of specific cell types, human induced pluripotent stem cell (hiPSC) lines have varing ability to differentiate into specific lineages. Therefore, surrogate markers for accurately predicting the differentiation propensity of hiPSC lines may facilitate cell-based therapeutic product development and manufacture. We attempted to identify marker genes that could predict the differentiation propensity of hiPSCs into neural stem/progenitor cells (NS/PCs). Using Spearman's rank correlation coefficients, we investigated genes in the undifferentiated state, the expression levels of which were significantly correlated with the neuronal differentiation propensity of several hiPSC lines. Among genes significantly correlated with NS/PC differentiation (P < 0.01), we identified ROR2 as a novel predictive marker. ROR2 expression in hiPSCs was negatively correlated with NS/PC differentiation tendency, regardless of the differentiation method, whereas its knockdown enhanced differentiation. ROR2 regulates NS/PC differentiation, suggesting that ROR2 is functionally essential for NS/PC differentiation. Selecting cell lines with relatively low ROR2 expression facilitated identification of hiPSCs that can differentiate into NS/PCs. Cells with ROR2 knockdown showed increased efficiency of differentiation into forebrain GABAergic neurons compared to controls. These findings suggest that ROR2 is a surrogate marker for selecting hiPSC lines appropriate for NS/PC and GABAergic neuronal differentiations.

Implantation and Decidualization in PCOS: Unraveling the Complexities of Pregnancy.

Polycystic Ovary Syndrome (PCOS) is a prevalent endocrine disorder in women of reproductive age, affecting 5-15% globally with a large proportion undiagnosed. This review explores the multifaceted nature of PCOS and its impact on pregnancy, including challenges in fertility due to hormonal imbalances and insulin resistance. Despite restoring ovulation pharmacologically, women with PCOS face lower pregnancy rates and higher risks of implantation failure and miscarriage. Our review focuses on the complexities of hormonal and metabolic imbalances that impair endometrial receptivity and decidualization in PCOS. Disrupted estrogen signaling, reduced integrity of endometrial epithelial tight junctions, and insulin resistance impair the window of endometrial receptivity. Furthermore, progesterone resistance adversely affects decidualization. Our review also examines the roles of various immune cells and inflammatory processes in the endometrium, contributing to the condition's reproductive challenges. Lastly, we discuss the use of rodent models in understanding PCOS, particularly those induced by hormonal interventions, offering insights into the syndrome's impact on pregnancy and potential treatments. This comprehensive review underscores the need for advanced understanding and treatment strategies to address the reproductive complications associated with PCOS, emphasizing its intricate interplay of hormonal, metabolic, and immune factors.

Unilateral renal agenesis and ipsilateral absence of the vas deferens in a cryptorchid infant with Klippel-Feil syndrome.

We report an exceedingly rare case of Klippel-Feil syndrome (KFS), compounded by ipsilateral absence of the vas deferens, renal agenesis, and diaphragmatic hernia. Unilateral absence of the vas deferens was found incidentally during orchidopexy. To the best of our knowledge, no case of unilateral absence of the kidney and vas deferens has been reported in children with KFS.

A Case of Solitary Kidney and Single-system Ectopic Ureter Draining Into the Vestibule in an Infant With Müllerian Duct Defect: Was the Bladder Missing?

We report a case of a solitary kidney and a single-system ectopic ureter draining into the vestibule in an infant with a Müllerian duct defect. Due to the absence of the upper vagina and uterus, an extremely underdeveloped bladder masqueraded as the vagina, and bladder agenesis was suspected preoperatively. Urinary continence was achieved using staged bladder surgery without augmentation or urinary diversion.

Single-Cell RNA-Seq Reveals LRRC75A-Expressing Cell Population Involved in VEGF Secretion of Multipotent Mesenchymal Stromal/Stem Cells Under Ischemia.

Human multipotent mesenchymal stromal/stem cells (MSCs) have been utilized in cell therapy for various diseases and their clinical applications are expected to increase in the future. However, the variation in MSC-based product quality due to the MSC heterogeneity has resulted in significant constraints in the clinical utility of MSCs. Therefore, we hypothesized that it might be important to identify and ensure/enrich suitable cell subpopulations for therapies using MSC-based products. In this study, we aimed to identify functional cell subpopulations to predict the efficacy of angiogenic therapy using bone marrow-derived MSCs (BM-MSCs). To assess its angiogenic potency, we observed various levels of vascular endothelial growth factor (VEGF) secretion among 11 donor-derived BM-MSC lines under in vitro ischemic culture conditions. Next, by clarifying the heterogeneity of BM-MSCs using single-cell RNA-sequencing analysis, we identified a functional cell subpopulation that contributed to the overall VEGF production in BM-MSC lines under ischemic conditions. We also found that leucine-rich repeat-containing 75A (LRRC75A) was more highly expressed in this cell subpopulation than in the others. Importantly, knockdown of LRRC75A using small interfering RNA resulted in significant inhibition of VEGF secretion in ischemic BM-MSCs, indicating that LRRC75A regulates VEGF secretion under ischemic conditions. Therefore, LRRC75A may be a useful biomarker to identify cell subpopulations that contribute to the angiogenic effects of BM-MSCs. Our work provides evidence that a strategy based on single-cell transcriptome profiles is effective for identifying functional cell subpopulations in heterogeneous MSC-based products.

Prenatally diagnosed giant umbilical cord resulted from patent urachus in a monochorionic diamniotic twin with selective intrauterine growth restriction.

A long diffuse giant umbilical cord (GUC), caused by umbilical cord edema associated with a patent urachus, is an extremely rare anomaly. While patients with diffuse GUC appear to experience no significant complications and a good prognosis, little is known about their etiology and prenatal course. Here, we report the first case of prenatally diagnosed diffuse GUC resulted from patent urachus in a monochorionic diamniotic twin with selective intrauterine growth restriction. This case indicates that GUC is epigenetic and unrelated to multiple births.

Steroid hormone signaling: multifaceted support of testicular function.

Embryonic development and adult physiology are dependent on the action of steroid hormones. In particular, the reproductive system is reliant on hormonal signaling to promote gonadal function and to ensure fertility. Here we will describe hormone receptor functions and their impacts on testicular function, focusing on a specific group of essential hormones: androgens, estrogens, progesterone, cortisol, and aldosterone. In addition to focusing on hormone receptor function and localization within the testis, we will highlight the effects of altered receptor signaling, including the consequences of reduced and excess signaling activity. These hormones act through various cellular pathways and receptor types, emphasizing the need for a multifaceted research approach to understand their critical roles in testicular function. Hormones exhibit intricate interactions with each other, as evidenced, for example, by the antagonistic effects of progesterone on mineralocorticoid receptors and cortisol's impact on androgens. In light of research findings in the field demonstrating an intricate interplay between hormones, a systems biology approach is crucial for a nuanced understanding of this complex hormonal network. This review can serve as a resource for further investigation into hormonal support of male reproductive health.

Immune Cells as Critical Regulators of Steroidogenesis in the Testis and Beyond.

Steroidogenesis is an essential biological process for embryonic development, reproduction, and adult health. While specific glandular cells, such as Leydig cells in the testis, are traditionally known to be the principal players in steroid hormone production, there are other cell types that contribute to the process of steroidogenesis. In particular, immune cells are often an important component of the cellular niche that is required for the production of steroid hormones. For several decades, studies have reported that testicular macrophages and Leydig cells are intimately associated and exhibit a dependency on the other cell type for their proper development; however, the mechanisms that underlie the functional relationship between macrophages and Leydig cells are unclear. Beyond the testis, in certain instances immune cells themselves, such as certain types of lymphocytes, are capable of steroid hormone production, thus highlighting the complexity and diversity that underlie steroidogenesis. In this review we will describe how immune cells are critical regulators of steroidogenesis in the testis and in extra-glandular locations, as well as discuss how this area of research offers opportunities to uncover new insights into steroid hormone production.

[CLINICAL AND IMAGING FINDINGS OF ACUTE EPIDIDYMITIS IN CHILDREN].

(Objective) The etiology of acute epididymitis in children remains poorly understood. Several studies have demonstrated that urine tests are negative in the majority of children with acute epididymitis, and the condition is self-limiting. The need for radiological evaluation of the urinary tract in children with acute epididymitis is still debatable. The aim of this study was to describe clinical and imaging findings in children with acute epididymitis. (Methods) We identified 47 children with acute epididymitis at our institute between 2017 and 2021.We retrospectively reviewed their clinical features and radiological and laboratory data. All children underwent ultrasonography of the kidney and urinary tract. (Results) Median patient age was 9 years (range, 6 months-16 years) and 60% of the cases occurred between the ages of 7 and 12 years. Thirteen children (28%) had a past history of genitourinary malformations. The common malformations were hypospadias in eight children and bladder dysfunction in three. Ultrasound revealed no new urinary tract abnormalities in the remaining 34 children. Urinalysis were performed in 27 children, nine of whom (33%) had pyuria. Urine culture was positive in two children. Of the nine children with genitourinary malformations, eight had pyuria. All 18 children without genitourinary malformations had a negative urinalysis except for one patient (p< 0.0001). (Conclusions) Acute epididymitis is a common cause of acute scrotum in pediatric patients. In this study, one-third of acute epididymitis cases presented pyuria, and about 30% had a past history of genitourinary malformations. The presence of pyuria was associated with a past history of genitourinary malformations. For children with no previous genitourinary malformations, routine use of ultrasound for the detection of urinary tract abnormalities is questionable due to the low yield.

Unilateral Fibrous Hyperplasia of the Labium Majus in Pre- and Early Pubertal Girls: A Report on 3 Cases.

BACKGROUND: In recent years, certain prepubertal cases of asymmetric enlargement of the labium majus came to be recognized as a physiological condition in response to hormonal stimulation. However, there exist only scattered case reports or a few case series on this entity, with different terminology. CASE: We present 3 cases of unilateral asymptomatic swelling of the labium majus in otherwise healthy pre- and early pubertal girls. Physical examination revealed painless, fluctuating, nontender bulging that was recognized for a few years on one side of the labium majus. Ultrasound and magnetic resonance imaging showed increased labial soft tissue without any masses. In all cases, surgical excision was performed for cosmetic request. Histopathological diagnosis was fibrous hyperplasia of the labium majus in all cases. Recurrence occurred in one of 3 cases a year after surgery. SUMMARY AND CONCLUSION: Due to the site- and age-specific non-neoplastic physiological condition, we recommend prepubertal unilateral fibrous hyperplasia of the labium majus (PUFHLM), as the terminology accurately reflects this entity. For the differential diagnosis of genital disorders in children, recognition of PUFHLM is important to avoid unnecessary biopsy or invasive procedures. Surgical excision for cosmetic reasons should not be considered because of the benign nature of this entity and a high recurrence rate in childhood and early adolescence.

Association of Lower Urinary Tract Symptoms Based on the International Prostate Symptom Score and Cardiovascular Disease.

BACKGROUND: Because it is unclear whether lower urinary tract symptoms (LUTS) are associated with cardiovascular disease (CVD) in the Japanese population, we explored the association in general Japanese men aged 55-75 years.Methods and Results:The cross-sectional study included male participants who had both national health checkup data and the International Prostate Symptom Score (IPSS) in the same calendar year between 2009 and 2017. LUTS severity was evaluated by IPSS. A robust Poisson regression model was used to assess the association between LUTS severity and the composite CVD outcome [coronary artery disease (CAD), stroke, or atrial fibrillation (AF)] and each component of the composite outcome. Prevalence ratio (PR) was adjusted for conventional cardiovascular risk factors. Of 16,781 male participants (mean age, 67±5 years), mild LUTS were observed in 9,243 (55.1%); moderate, 6,445 (38.4%); and severe, 1,093 (6.5%). Compared with the mild LUTS group, moderate LUTS [PR 1.18, 95% confidence interval (CI) 1.10-1.25, P<0.001] and severe LUTS (PR 1.38, 95% CI 1.24-1.53, P<0.001) were significantly associated with a higher prevalence of CVD. LUTS severity was associated with higher prevalence of CAD and stroke, but not AF. CONCLUSIONS: The severity of LUTS was associated with a higher prevalence of CVD, especially CAD and stroke, independent of conventional CVD risk factors.

A selective cytotoxic adenovirus vector for concentration of pluripotent stem cells in human pluripotent stem cell-derived neural progenitor cells.

Highly sensitive detection of residual undifferentiated pluripotent stem cells is essential for the quality and safety of cell-processed therapeutic products derived from human induced pluripotent stem cells (hiPSCs). We previously reported the generation of an adenovirus (Ad) vector and adeno-associated virus vectors that possess a suicide gene, inducible Caspase 9 (iCasp9), which makes it possible to sensitively detect undifferentiated hiPSCs in cultures of hiPSC-derived cardiomyocytes. In this study, we investigated whether these vectors also allow for detection of undifferentiated hiPSCs in preparations of hiPSC-derived neural progenitor cells (hiPSC-NPCs), which have been expected to treat neurological disorders. To detect undifferentiated hiPSCs, the expression of pluripotent stem cell markers was determined by immunostaining and flow cytometry. Using immortalized NPCs as a model, the Ad vector was identified to be the most efficient among the vectors tested in detecting undifferentiated hiPSCs. Moreover, we found that the Ad vector killed most hiPSC-NPCs in an iCasp9-dependent manner, enabling flow cytometry to detect undifferentiated hiPSCs intermingled at a lower concentration (0.002%) than reported previously (0.1%). These data indicate that the Ad vector selectively eliminates hiPSC-NPCs, thus allowing for sensitive detection of hiPSCs. This cytotoxic viral vector could contribute to ensuring the quality and safety of hiPSCs-NPCs for therapeutic use.

Congenital perineal urethrocutaneous fistula without rupture in a neonate.

Congenital anterior urethrocutaneous fistula (CAUF) is a rare anomaly, commonly observed in the mid-shaft or more distal position of the penis. It can be seen in the perineum in anorectal anomalies or in cases of duplicated urethra. However, isolated CAUF opening into the perineum has not been reported. Herein, we present a case of congenital anterior urethral diverticulum and possible perineal urethrocutaneous fistula of an otherwise healthy neonate.

[A CASE OF SPLENOGONADAL FUSION PRESENTING NON-PALPABLE TESTIS AND PROXIMAL HYPOSPADIAS].

A 9-month-old boy, having a history of cardiac surgery in neonatal period, was referred to our department for evaluation of genital anomalies. The antenatal course was uneventful, except for unknown gender. His family history was unremarkable. He was delivered at full term, and his birth weight was 3,510 g. Physical examination revealed proximal hypospadias and a non-palpable testis on the left side. Chromosome studies showed a normal male karyotype with positive SRY. At the age of 14 months, he underwent hypospadias repair. Three months later, left testicular exploration was performed along with orchidopexy of an ascending testis on the contralateral side. As nothing was found through an inguinal incision on the left side, laparoscopy was indicated. Laparoscopic observation revealed a small dark reddish mass cranially connected to the left hypoplastic testis that was located high in the left iliac fossa. The epididymis and vas deference looked abnormal, and detachment to the testis was apparent. Testicular vessels were undifferentiated from the mass. Therefore, the left testis was excised with the mass. Histopathological examination confirmed the testis and spleen tissue, and the diagnosis of splenogonadal fusion was made postoperatively.

Variation of Gonadal Dysgenesis and Tumor Risk in Patients With 45,X/46,XY Mosaicism.

OBJECTIVE: To describe the gonadal features of patients with 45,X/46,XY mosaicism, and to evaluate the prevalence of gonadal tumor in different phenotypes. MATERIALS AND METHODS: The medical records of consecutive patients with 45,X/46,XY karyotype or its variants who had undergone gonadal biopsy or gonadectomy at a single institute between 1996 and 2017 were retrospectively reviewed. RESULTS: Of 34 patients with 45,X/46,XY mosaicism, a unilateral dysgenetic testis and a contralateral streak gonad was detected in 20 patients (59%), bilateral streak gonads in 9 (26%), and bilateral dysgenetic testes in 5 (15%). A gonad composed of both streak and dysgenetic testicular portions was observed in 7 gonads of 6 patients. All streak gonads were removed, and bilateral gonadectomy was performed in 15 patients raised as girls. Pathologic examination revealed gonadal tumors in 6 of the 34 (18%) patients, including a gonadoblastoma in 7 gonads among 5 patients and an association of dysgerminoma with gonadoblastoma in 1 gonad. All 6 patients who developed gonadal tumor had female genitalia. Postoperative course was uneventful except 1 boy. A seminoma was developed in his soritaly scrotal testis at the age of 16 years. CONCLUSION: The prevalence of gonadal tumor in patients with 45,X/46,XY mosaicism may vary according to the phenotype, and high in patients with female phenotype. Considering the increased risk of gonadal tumors in such patients, early investigation and individual management, including prophylactic gonadectomy, are recommended. In male patients, a close follow-up of the preserved testes is mandatory until adulthood.

SALL3 expression balance underlies lineage biases in human induced pluripotent stem cell differentiation.

Clinical applications of human induced pluripotent stem cells (hiPSCs) are expected, but hiPSC lines vary in their differentiation propensity. For efficient selection of hiPSC lines suitable for differentiation into desired cell lineages, here we identify SALL3 as a marker to predict differentiation propensity. SALL3 expression in hiPSCs correlates positively with ectoderm differentiation capacity and negatively with mesoderm/endoderm differentiation capacity. Without affecting self-renewal of hiPSCs, SALL3 knockdown inhibits ectoderm differentiation and conversely enhances mesodermal/endodermal differentiation. Similarly, loss- and gain-of-function studies reveal that SALL3 inversely regulates the differentiation of hiPSCs into cardiomyocytes and neural cells. Mechanistically, SALL3 modulates DNMT3B function and DNA methyltransferase activity, and influences gene body methylation of Wnt signaling-related genes in hiPSCs. These findings suggest that SALL3 switches the differentiation propensity of hiPSCs toward distinct cell lineages by changing the epigenetic profile and serves as a marker for evaluating the hiPSC differentiation propensity.

Gonadal function and testicular histology in males with Prader-Willi syndrome.

CONTEXT: Cryptorchidism is common in Prader-Willi syndrome (PWS) males, but the testicular histology in childhood remains uncertain. The association between testicular histology and long-term gonadal function in PWS males is also unknown. OBJECTIVES: To evaluate the relationship between testicular histology in childhood and long-term gonadal function in PWS males. PATIENTS AND METHODS: Forty men with PWS were assessed longitudinally at our institute over the past 24 years. Clinical examinations and blood tests for LH, FSH and testosterone levels were compared with normal reference values. Tissue specimens were collected during orchiopexy and analyzed based on Nistal categories. RESULTS: Of nine testes available for pathological assessments, two showed favourable histology (Nistal I) and seven showed unfavourable histology (Nistal II or III). Of five postpubertal males with histology available, four reached puberty spontaneously, but only one reached Tanner stage 5. In a male with favourable histology, LH and FSH were high, but testosterone was normal, though below the average of the reference range. In three males with unfavourable histology, LH was normal, but FSH was highly elevated, and testosterone was at the lower limit of normal. One patient took hCG treatment to induce puberty; this patient showed favourable histology, but LH, FSH and testosterone were not elevated in adolescence. CONCLUSIONS: Testicular histology of PWS men in childhood varies from normal to Sertoli Cell-Only Syndrome. Regardless of the testicular histology in childhood, hypogonadism in PWS adults arises as a consequence of primary testicular dysfunction with highly elevated FSH and insufficient testosterone levels.

Development of selective cytotoxic viral vectors for concentration of undifferentiated cells in cardiomyocytes derived from human induced pluripotent stem cells.

Cell-processed therapeutic products (CTPs) derived from human pluripotent stem cells (hPSCs) have innovative applications in regenerative medicine. However, undifferentiated hPSCs possess tumorigenic potential; thus, sensitive methods for the detection of residual undifferentiated hPSCs are essential for the clinical use of hPSC-derived CTPs. The detection limit of the methods currently available is 1/105 (0.001%, undifferentiated hPSCs/differentiated cells) or more, which could be insufficient for the detection of residual hPSCs when CTPs contain more than 1 × 105 cells. In this study, we developed a novel approach to overcome this challenge, using adenovirus and adeno-associated virus (AdV and AAV)-based selective cytotoxic vectors. We constructed AdV and AAV vectors that possess a suicide gene, iCaspase 9 (iCasp9), regulated by the CMV promoter, which is dormant in hPSCs, for the selective expression of iCasp9 in differentiated cells. As expected, AdV/CMV-iCasp9 and AAV/CMV-iCasp9 exhibited cytotoxicity in cardiomyocytes but not in human induced pluripotent stem cells (hiPSCs). The vectors also induced apoptosis in hiPSC-derived cardiomyocytes, and the surviving cells exhibited higher levels of hPSC marker expression. These results indicate that the AdV- and AAV-based cytotoxic vectors concentrate cells expressing the undifferentiated cell markers in hiPSC-derived products and are promising biological tools for verifying the quality of CTPs.

[LOCALIZED CYSTIC DISEASE OF THE KIDNEY IN A 6-YEAR-OLD GIRL].

Localized cystic disease of the kidney (LCDK) is a rare, non-hereditary, non-progressive benign cystic renal condition. It is found primarily in adults, and is rarely reported in children. To date, only 5 prepubertal cases of LCDK have been reported in the literature. In this report, we present a case of LCDK that was difficult to differentiate from malignant renal tumor in a 6-year-old girl. Nephron-sparing surgery could not be performed.A 6-year-old girl with no past medical history presented with gross hematuria and right-sided abdominal pain. An abdominal ultrasound at a local hospital showed multiple variable-sized cysts throughout the right kidney. She was referred to our hospital for further evaluation. Computed tomography demonstrated that the region between cysts was slightly enhanced in some part of the lower pole and was diagnosed III in the Bosniak classification. Nephrectomy was performed for possible cystic renal neoplasm. After surgery, a diagnosis of LCDK was made by histopathological examination.

Tumorigenicity-associated characteristics of human iPS cell lines.

Human induced pluripotent stem cells (hiPSCs) represent promising raw materials of human cell-based therapeutic products (hCTPs). As undifferentiated hiPSCs exhibit intrinsic tumorigenicity properties that enable them to form teratomas, hCTPs containing residual undifferentiated hiPSCs may cause tumor formation following transplantation. We first established quantitative and sensitive tumorigenicity testing of hiPSCs dissociated into single cells using NOD/Shi-scid IL2Rγnull (NOG) mice by inhibiting apoptosis of hiPSCs with a Rho kinase inhibitor. To examine different features in tumorigenicity of various hiPSCs, 10 commonly available hiPSC lines were subjected to in vivo tumorigenicity testing. Transplanted hiPSC lines showed remarkable variation in tumor incidence, formation latency, and volumes. Most of the tumors formed were classified as immature teratomas. However, no signs of malignancies, such as carcinoma and sarcoma, were recognized in the tumors. Characteristics associated tumorigenicity of hiPSCs were investigated with microarray analysis, karyotype analysis, and whole exome sequencing. Gene expression profiling and pathway analysis supported different features of hiPSC lines in tumorigenicity. hiPSC lines showed chromosomal abnormalities in some lines and 61-77 variants of cancer-related genes carrying effective nonsynonymous mutations, which were confirmed in the COSMIC databases. In this study, the chromosomal abnormalities and cancer-related gene mutations observed in hiPSC lines did not lead to the malignancy of tumors derived from hiPSCs. Our results suggest that the potential tumorigenicity risk of hCTPs containing residual undifferentiated hiPSCs is dependent on not only amounts of undifferentiated hiPSCs but also features of the cell lines used as raw materials, a finding that should be considered from the perspective of quality of hCTPs used.