One model fits all: Combining inference and simulation of gene regulatory networks.

The rise of single-cell data highlights the need for a nondeterministic view of gene expression, while offering new opportunities regarding gene regulatory network inference. We recently introduced two strategies that specifically exploit time-course data, where single-cell profiling is performed after a stimulus: HARISSA, a mechanistic network model with a highly efficient simulation procedure, and CARDAMOM, a scalable inference method seen as model calibration. Here, we combine the two approaches and show that the same model driven by transcriptional bursting can be used simultaneously as an inference tool, to reconstruct biologically relevant networks, and as a simulation tool, to generate realistic transcriptional profiles emerging from gene interactions. We verify that CARDAMOM quantitatively reconstructs causal links when the data is simulated from HARISSA, and demonstrate its performance on experimental data collected on in vitro differentiating mouse embryonic stem cells. Overall, this integrated strategy largely overcomes the limitations of disconnected inference and simulation.

Three classes of epigenomic regulators converge to hyperactivate the essential maternal gene deadhead within a heterochromatin mini-domain.

The formation of a diploid zygote is a highly complex cellular process that is entirely controlled by maternal gene products stored in the egg cytoplasm. This highly specialized transcriptional program is tightly controlled at the chromatin level in the female germline. As an extreme case in point, the massive and specific ovarian expression of the essential thioredoxin Deadhead (DHD) is critically regulated in Drosophila by the histone demethylase Lid and its partner, the histone deacetylase complex Sin3A/Rpd3, via yet unknown mechanisms. Here, we identified Snr1 and Mod(mdg4) as essential for dhd expression and investigated how these epigenomic effectors act with Lid and Sin3A to hyperactivate dhd. Using Cut&Run chromatin profiling with a dedicated data analysis procedure, we found that dhd is intriguingly embedded in an H3K27me3/H3K9me3-enriched mini-domain flanked by DNA regulatory elements, including a dhd promoter-proximal element essential for its expression. Surprisingly, Lid, Sin3a, Snr1 and Mod(mdg4) impact H3K27me3 and this regulatory element in distinct manners. However, we show that these effectors activate dhd independently of H3K27me3/H3K9me3, and that dhd remains silent in the absence of these marks. Together, our study demonstrates an atypical and critical role for chromatin regulators Lid, Sin3A, Snr1 and Mod(mdg4) to trigger tissue-specific hyperactivation within a unique heterochromatin mini-domain.

The male external urethral sphincter is autonomically innervated.

INTRODUCTION: The aim of the present study was to describe autonomic urethral sphincter (US) innervation using specific muscular and neuronal antibody markers and 3D reconstruction. MATERIAL AND METHODS: We performed en-bloc removal of the entire pelvis of three male human fetuses between 18 and 40 weeks. Serial whole mount sections (5 µm intervals) were stained and investigated. The sections were stained with Masson's trichrome and Eosin Hematoxylin, and immunostained with: anti-SMA antibody for smooth muscle; anti-S100 antibody for all nerves; and anti-PMP22 antibody, anti-TH antibody, anti-CGRP antibody, anti-NOS antibody for somatic, adrenergic, sensory and nitrergic nerve fibers, respectively. The slides were digitized for 3D reconstruction to improve topographical understanding. An animated reconstruction of the autonomic innervation of the US was generated. RESULTS: The external and internal US are innervated by autonomic nerves of the inferior hypogastric plexus (IHP). These nerves are sympathetic (positive anti-TH antibody), sensory (positive anti-CGRP antibody), and nitrergic (positive anti-NOS antibody). Some autonomic fibers run within the neurovascular bundles, posterolaterally. Others run from the IHP to the posteromedial aspect of the prostate apex, above an through the rectourethral muscle. The external US is also innervated by somatic nerves (positive anti-PMP22 antibody) arising from the pudendal nerve, joining the midline but remaining below the rectourethral. CONCLUSIONS: This study provides anatomical evidence of an autonomic component in the innervation of the external US that travels in the neurovascular bundle. During radical prostatectomy, the rectourethral muscle and the neurovascular bundles are to be preserved, particularly during apical dissection.

The Lid/KDM5 histone demethylase complex activates a critical effector of the oocyte-to-zygote transition.

Following fertilization of a mature oocyte, the formation of a diploid zygote involves a series of coordinated cellular events that ends with the first embryonic mitosis. In animals, this complex developmental transition is almost entirely controlled by maternal gene products. How such a crucial transcriptional program is established during oogenesis remains poorly understood. Here, we have performed an shRNA-based genetic screen in Drosophila to identify genes required to form a diploid zygote. We found that the Lid/KDM5 histone demethylase and its partner, the Sin3A-HDAC1 deacetylase complex, are necessary for sperm nuclear decompaction and karyogamy. Surprisingly, transcriptomic analyses revealed that these histone modifiers are required for the massive transcriptional activation of deadhead (dhd), which encodes a maternal thioredoxin involved in sperm chromatin remodeling. Unexpectedly, while lid knock-down tends to slightly favor the accumulation of its target, H3K4me3, on the genome, this mark was lost at the dhd locus. We propose that Lid/KDM5 and Sin3A cooperate to establish a local chromatin environment facilitating the unusually high expression of dhd, a key effector of the oocyte-to-zygote transition.

Detailed muscular structure and neural control anatomy of the levator ani muscle: a study based on female human fetuses.

BACKGROUND: Injury to the levator ani muscle or pelvic nerves during pregnancy and vaginal delivery is responsible for pelvic floor dysfunction. OBJECTIVE: We sought to demonstrate the presence of smooth muscular cell areas within the levator ani muscle and describe their localization and innervation. STUDY DESIGN: Five female human fetuses were studied after approval from the French Biomedicine Agency. Specimens were serially sectioned and stained by Masson trichrome and immunostained for striated and smooth muscle, as well as for somatic, adrenergic, cholinergic, and nitriergic nerve fibers. Slides were digitized for 3-dimensional reconstruction. One fetus was reserved for electron microscopy. We explored the structure and innervation of the levator ani muscle. RESULTS: Smooth muscular cell beams were connected externally to the anococcygeal raphe and the levator ani muscle and with the longitudinal anal muscle sphincter. The caudalmost part of the pubovaginal muscle was found to bulge between the rectum and the vagina. This bulging was a smooth muscular interface between the levator ani muscle and the longitudinal anal muscle sphincter. The medial (visceral) part of the levator ani muscle contained smooth muscle cells, in relation to the autonomic nerve fibers of the inferior hypogastric plexus. The lateral (parietal) part of the levator ani muscle contained striated muscle cells only and was innervated by the somatic nerve fibers of levator ani and pudendal nerves. The presence of smooth muscle cells within the medial part of the levator ani muscle was confirmed under electron microscopy in 1 fetus. CONCLUSION: We characterized the muscular structure and neural control of the levator ani muscle. The muscle consists of a medial part containing smooth muscle cells under autonomic nerve influence and a lateral part containing striated muscle cells under somatic nerve control. These findings could result in new postpartum rehabilitation techniques.

The Bile Acid Nuclear Receptor FXRα Is a Critical Regulator of Mouse Germ Cell Fate.

Spermatogenesis is the process by which spermatozoa are generated from spermatogonia. This cell population is heterogeneous, with self-renewing spermatogonial stem cells (SSCs) and progenitor spermatogonia that will continue on a path of differentiation. Only SSCs have the ability to regenerate and sustain spermatogenesis. This makes the testis a good model to investigate stem cell biology. The Farnesoid X Receptor alpha (FXRα) was recently shown to be expressed in the testis. However, its global impact on germ cell homeostasis has not yet been studied. Here, using a phenotyping approach in Fxrα-/- mice, we describe unexpected roles of FXRα on germ cell physiology independent of its effects on somatic cells. FXRα helps establish and maintain an undifferentiated germ cell pool and in turn influences male fertility. FXRα regulates the expression of several pluripotency factors. Among these, in vitro approaches show that FXRα controls the expression of the pluripotency marker Lin28 in the germ cells.

Origin and nature of pelvic ureter innervation.

AIMS: Innervation of the pelvic ureter traditionally comes from the pelvic plexus. This innervation is independent: adrenergic and cholinergic. The purpose of this study was to describe more precisely the origin and nature of its innervation (adrenergic, cholinergic, nitrergic, and somatic). METHODS: Six specimens of normal human fetal pelvis (four male and two female) from 20 to 30 weeks gestation were studied. The sections of these fetuses, carried out every 5 µm without interval, were treated with Hematoxylin Eosin (HE), with Masson's trichrome (TriM), immunolabeling of smooth muscle cells with smooth anti-actin, of nerves with anti-S100 protein, anti-tyrosine hydroxylase, anti-VAChT, anti-nNOS, and with anti- peripheral myelin protein 22 (PMP 22). The slides were scanned and two-dimensional images reconstructed in 3D, and analyzed. RESULTS: The terminal pelvic ureter travels above and inside the inferior hypogastric plexus (IHP). The nerve fibers that innervate the ureterovesical junction come mainly from the superior hypogastric plexus (SHP) which gives off the hypogastric nerves and pelvic branches of the sacral plexus that form the IHP. Most nerve fibers meet below the ureter, behind the bladder to form an ascending bundle, which innervates the pelvic ureter. Immunohistochemical analysis shows that the nerves of the pelvic ureter consist of adrenergic, cholinergic, and nitrergic fibers. CONCLUSION: The innervation of the distal ureter depends mainly on the SHP. This innervation is adrenergic, cholinergic, and nitrergic. It innervates the pelvic ureter in an ascending manner. This anatomical information can change rectal resection and ureteral reimplantation techniques and drug treatments for pelvic ureter stones. Neurourol. Urodynam. 36:271-279, 2017. © 2015 Wiley Periodicals, Inc.

Testicular Shear Wave Elastography in Normal and Infertile Men: A Prospective Study on 601 Patients.

Our aim in the study described here was to prospectively establish the feasibility of using and reproducibility of testicular shear-wave elastography in the assessment of testicular stiffness in 62 normal patients and 539 infertile men with obstructive azoospermia (OA), non-Klinefelter syndrome non-obstructive azoospermia (non-KS NOA), Klinefelter syndrome NOA (KS NOA), oligoasthenoteratozoospermia (OAT) or a left varicocele. The feasibility rate was 96.9%, with an intra-class correlation coefficient of 0.85 (95% confidence interval: 0.83-0.88). Median stiffness (interquartile range) values were 2.4 kPa (2.0, 2.9), 2.1 kPa (1.8, 2.5), 2.4 kPa (2.0, 2.7), 2.0 kPa (1.7, 2.4), 2.6 kPa (2, 3.2) and 2.2 kPa (1.8, 2.6) for men with a normal testis (n = 108), OAT (n = 689), OA (n = 119), non-KS NOA (n = 183), KS NOA (n = 70) and varicocele (n = 132), respectively. Testicular shear wave elastography is a feasible and reproducible technique. A significant positive association was found between stiffness and testis volume (p = 0.001). Testicular stiffness was higher in OA than in non-KS NOA populations (p = 1.e-10) and in KS NOA than in NOA populations (p = 2.0e-8), but the substantial number of overlapping values limited the clinical impact.

Burned-Out Testis Tumors in Asymptomatic Infertile Men: Multiparametric Sonography and MRI Findings.

Multiparametric testicular ultrasound and magnetic resonance imaging (MRI) findings were analyzed in a series of 10 infertile asymptomatic men presenting with pathologically confirmed burned-out testicular tumors. Color/power Doppler ultrasound (CDUS), shear wave elastography (SWE), contrast-enhanced ultrasonography (CEUS), and MRI were performed on 10, 5, 6, and 7 patients, respectively. All lesions appeared as a hypoechoic and hypovascular nodular area at CDUS, SWE, CEUS CDUS, and CEUS (if performed). Shear wave elastography showed a stiffer nodular area compared with the surrounding/contralateral tissues (13 versus 2 kPa); MRI revealed a well-delineated nodular area in hypointense signal on T2, a high apparent diffusion coefficient value, and a lack of enhancement.

Levator ani muscle innervation: Anatomical study in human fetus.

AIMS: To characterize the nature and function of the levator ani muscle innervation pathways and to perform a comprehensive three-dimensional reconstruction of female pelvic innervation. METHODS: A computer-assisted anatomical dissection protocol was applied to seven female human fetuses, after approval from the national biomedicine agency. Specimens were serially sectioned and immunostained for overall (antibody against protein S100), somatic (antibody against peripheral myelin protein 22), adrenergic (antibody against tyrosine hydroxylase), cholinergic (antibody against vesicular acetylcholine transferase), and nitrergic (antibody against the neural isoform of nitric oxide synthase) nerve fibers. Slides were digitized for three-dimensional reconstructions using WinSurf®. RESULTS: Three main nerve pathways to the levator ani muscle were observed: the levator ani nerve, the pudendal nerve, and the inferior hypogastric plexus. The pudendal nerve was both somatic and autonomic, located below the levator ani muscle (infralevator pathway), supplying innervation to the inferior aspect of the levator ani muscle. The levator ani nerve was solely somatic, located above the levator ani muscle (supralevator pathway), supplying innervation to the superior aspect of the levator ani muscle. The inferior hypogastric plexus nerve fibers were solely autonomic, located in between the levator ani muscle and pelvic organs (endolevator pathway), supplying innervation to the medial portion of the levator ani muscle. CONCLUSIONS: Our study provides a new representation of levator ani muscle innervation with three nerve pathways, and the levator ani muscle itself as an anatomical landmark.

Functional and structural microanatomy of the fetal sciatic nerve.

INTRODUCTION: The ultrastructure of a nerve has implications for surgical nerve repair. The aim of our study was to characterize the fascicular versus fibrillar anatomy and the autonomic versus somatic nature of the fetal sciatic nerve (SN). METHODS: Immunohistochemistry for vesicular acetylcholine transporter, tyrosine hydroxylase, and peripheral myelin protein 22 was performed to identify cholinergic, adrenergic, and somatic axons, respectively, in the human fetal SN. Two-dimensional (2D) analysis and 3D reconstructions were performed. RESULTS: The fetal SN is composed of one-third stromal tissue and two-thirds neural tissue. Autonomic fibers are predominant over somatic fibers within the neural tissue. The distribution of somatic fibers is initially random, but then become topographically organized after intra- and interfascicular rearrangements have occurred within the nerve. CONCLUSIONS: The fetal model presents limitations but enables illustration of the nature of the nerve fibers and the 3D fascicular anatomy of the SN. Muscle Nerve 56: 787-796, 2017.

Sunitinib for the treatment of benign and malignant neoplasms from von Hippel-Lindau disease: A single-arm, prospective phase II clinical study from the PREDIR group.

Von Hippel-Lindau (VHL) disease is an autosomal dominant hereditary cancer syndrome that predisposes affected individuals to the development of multiple benign and malignant tumors. One of the main manifestations of VHL is renal cell carcinoma (RCC). RCC is increasingly being treated with targeted therapies, which offer an alternative treatment option for patients with VHL disease. This study investigated the effectiveness of sunitinib in VHL patients with advanced tumors or tumors unsuitable for surgery.This multicenter, phase II, open-label study from the PREDIR VHL network, treated patients with genetically-confirmed advanced VHL disease with oral sunitinib (50 mg/day for 28 days then a 2-week rest period) until progression. Lesions were performed using magnetic resonance imaging (MRI) and computed tomographic (CT) scan. The primary endpoint was objective response rate; secondary endpoints included tolerability and overall survival.All five patients showed stable disease as best response at 6 months. Two patients showed impressive transitory clinical improvement during early cycles. No patient died during sunitinib treatment. Reasons for discontinuing sunitinib therapy were disease progression (n=1), unacceptable toxicity (n=3) and lack of clinical improvement after 7 cycles (10.5 months) with unacceptable toxicity (n=1).In conclusion, sunitinib was of limited benefit in patients with advanced VHL disease, but had better efficacy against metastatic RCC than other VHL-related lesions. Treatment-related toxicity is an important limiting factor in this frail patient population. New agents with different mechanisms of action are required to treat this disease.

Non-palpable incidentally found testicular tumors: Differentiation between benign, malignant, and burned-out tumors using dynamic contrast-enhanced MRI.

PURPOSE: To evaluate qualitative, semi-quantitative, and quantitative parameters obtained by dynamic contrast-enhanced (DCE)-MRI for the characterization of histologically proven, non-palpable, incidentally found intratesticular tumors. MATERIALS AND METHODS: From 2006 to 2014, we included men with non-palpable, incidentally found testicular tumors on ultrasound, normal tumoral marker levels,referred for surgery. DCE-MRI data were analyzed retrospectively and independently by two radiologists blinded to the histological diagnosis. The visual enhancement patterns, time-signal intensity curves, shape of the curves (type 0-3), maximal relative enhancement (Peak), initial enhancement slope (IS), time to peak (TTP), as well as transfer constants Ktrans and Kep were compared between the tumors. The interobserver correlation was evaluated. Receiver Operating Characteristic (ROC) curves and areas under the curve (AUC) were extracted. RESULTS: Thirty-one patients (mean age of 37.3 years) were included. Tumor mean size was 1.2±0.77 cm (min=0.3cm, max=2.8cm). Regarding the histology results, three groups were defined: Twelve stromal "benign tumors" (BT) exhibited more type 2 and type 3 curves than 12 "malignant tumors" (MT) and 7 "burned-out tumors" (BOT) (p<0.0001). BT had a higher peak (96 vs. 54 and 17%), shorter TTP (215 vs. 412 and 692 sec), higher IS (73 vs. 12 and 2 arbitrary units), higher Ktrans (255 vs. 88 and 14min-1*1000) and higher Kep (554 vs. 159 and 48min-1*1000) than MT and BOT, respectively (p<0.0001, p=0.0003, p<0.0001, p<0.0001 and p<0.0001, respectively). The agreement coefficient values and the AUC extracted after gathering MT with BOT varied from 0.83 to 0.96 and from 0.868 to 0.978, respectively. CONCLUSION: DCE-MRI may assist in differentiating between benign intratesticular stromal tumors,malignant and burned-out tumors.

Nestin expression on tumour vessels and tumour-infiltrating macrophages define a poor prognosis subgroup of pt1 clear cell renal cell carcinoma.

The behaviour of clear cell renal cell carcinoma (CCRCC) is highly unpredictable. Despite adequate initial surgery, 20 to 30 % of patients will develop local recurrence or metastasis during follow-up. Usual clinical and pathology parameters tend to classify most patients in an intermediate prognosis group, and molecular markers to determine prognosis more accurately are needed. A key feature of CCRCC is its abundant vascularization. Factors that upregulate angiogenesis, such as hypoxia and the presence of immune cells including macrophages, also modulate tumour proliferation and metastasis. We studied angiogenesis, as defined by nestin-positive capillaries, and tumour infiltration by macrophages especially in the good prognosis pT1 subgroup of CCRCC. We assessed whether these parameters are associated with metastatic extension and survival in CCRCC. The expression of HIF1α, CAIX, nestin, CD68 and CD163 was assessed by immunohistochemistry on a tissue microarray (TMA) containing tissue samples from 257 consecutive patients with sporadic CCRCC. Factors associated with progression-free (PFS) and overall survival (OS) were analysed. The presence of nestin-positive tumour vessels was independently associated with shorter PFS in the whole cohort and in the pT1 subgroup. The presence of tumour-infiltrating macrophages was independently associated with shorter OS in the whole cohort and in the pT1 subgroup. The presence of nestin-positive endothelial cells is associated with early relapse, especially in the pT1 subgroup and may help to select patients for antiangiogenic treatment. The presence of tumour-infiltrating M2-type macrophages is a strong predictor of short survival and may be used to adapt treatment strategy.

Colour Doppler and ultrasound characteristics of testicular Leydig cell tumours.

OBJECTIVE: To assess the colour Doppler and ultrasound features of testicular Leydig cell tumours (LCTs) in a population of 38 surgically proven lesions. METHODS: From August 2008 to March 2015, we retrospectively included 38 surgically proven LCTs in 36 patients. Clinical data, scrotal colour Doppler, B-mode ultrasound and videos images were reviewed for each patient. The volume, echotexture of the testis, size, shape, echogenicity and the vascularization pattern of the lesion were evaluated. The tumour margins were categorized as either smooth or lobulated. The vascularization was classified as intense, moderate or without any hypervascularization. We defined the vascularization pattern groups as central, peripheral and mixed (the latter meaning both central and peripheral). RESULTS: 26 patients were referred for infertility [5 patients were subsequently diagnosed with Klinefelter syndrome (KS) and 5 patients with cryptorchidism]. 28 patients underwent testis-sparing surgery, while 8 patients underwent a radical orchiectomy. The LCTs were mostly infracentimetric (68.4%), with a median size of 7.0 mm (ranging from 4.0 to 11 mm). 50% of the lesions had lobulated margins, and these were significantly larger than the smooth lesions (p < 0.05). The content of the lesions was markedly homogeneous and hypoechoic. All lesions had sharp demarcations from the adjacent pulp. 36/38 lesions exhibited moderate-to-intense hypervascularization, with a mixed intrinsic and peripheral rim pattern. Larger lesions were more hypervascularized (p < 0.05). LCTs in patients with KS had atypical features. CONCLUSION: Typical sporadic LCTs appeared as isolated hypoechoic, infracentimetric masses, with a clear demarcation from the adjacent pulp. They presented intrinsic and peripheral rim hypervascularization. ADVANCES IN KNOWLEDGE: By undertaking the largest imaging series of LCT to date (to our knowledge), we reassessed the typical sonographical aspects of LCTs, so as to provide guidance in regard to opting for testis-sparing surgery and for follow-up. LCTs present both intrinsic and rim vascularization detectable by colour Doppler ultrasound. Intrinsic vascularization and lobulated margins are common findings in testicular LCTs.

Testis ultrasound in Klinefelter syndrome infertile men: making the diagnosis and avoiding inappropriate management.

OBJECTIVE: To compare the testicular Color Doppler ultrasound (US), hormone levels, and histological results from 67 infertile men with Klinefelter syndrome (KS), vs. 66 non-KS non-obstructive azoospermic men. METHODS: Scrotal US images were collected from 67 infertile KS and 66 non-obstructive, non-KS azoospermic men. The testis volume, echotexture, vascularity, and microliths were evaluated and graded. We defined the following echo pattern alteration groups: normal, striated, coarse, and measurable nodules. The vascularization was classified as low, normal, moderate, or strong. Testosterone, follicle-stimulating hormone, luteinizing hormone, and inhibin B levels were determined. Large testicular nodules were removed. A testicular biopsy and sperm extraction was performed in 18 of the KS, and all of the 66 non-KS men. RESULTS: The mean testis volume was low in the KS, compared to the non-KS patients: i.e., 2 vs. 8 mL (P < 0.0001). The distributions in the echotexture groups differed markedly, with coarse or nodular patterns in the KS men, and normal/striated patterns in the control patients (P < 0.0001). The vascularization and microlithiasis grades were higher in the KS patients than the control men (P < 0.0001 and P < 0.001, respectively). All of the nodules removed from the KS patients were benign Leydig cell tumors, and all of the biopsies showed marked Leydig cell hyperplasia, with spermatogenesis in only two patients. The non-KS biopsies were predominantly Sertoli cell-only syndrome. CONCLUSIONS: Small testes, with a coarse or nodular echotexture, hypervascularization, and microlithiasis are associated with KS. The KS nodules were benign Leydig cell tumors/hyperplasias.

Retinoic Acid Receptors Control Spermatogonia Cell-Fate and Induce Expression of the SALL4A Transcription Factor.

All-trans retinoic acid (ATRA) is instrumental to male germ cell differentiation, but its mechanism of action remains elusive. To address this question, we have analyzed the phenotypes of mice lacking, in spermatogonia, all rexinoid receptors (RXRA, RXRB and RXRG) or all ATRA receptors (RARA, RARB and RARG). We demonstrate that the combined ablation of RXRA and RXRB in spermatogonia recapitulates the set of defects observed both upon ablation of RAR in spermatogonia. We also show that ATRA activates RAR and RXR bound to a conserved regulatory region to increase expression of the SALL4A transcription factor in spermatogonia. Our results reveal that this major pluripotency gene is a target of ATRA signaling and that RAR/RXR heterodimers are the functional units driving its expression in spermatogonia. They add to the mechanisms through which ATRA promote expression of the KIT tyrosine kinase receptor to trigger a critical step in spermatogonia differentiation. Importantly, they indicate also that meiosis eventually occurs in the absence of a RAR/RXR pathway within germ cells and suggest that instructing this process is either ATRA-independent or requires an ATRA signal originating from Sertoli cells.

RAR/RXR binding dynamics distinguish pluripotency from differentiation associated cis-regulatory elements.

In mouse embryonic cells, ligand-activated retinoic acid receptors (RARs) play a key role in inhibiting pluripotency-maintaining genes and activating some major actors of cell differentiation. To investigate the mechanism underlying this dual regulation, we performed joint RAR/RXR ChIP-seq and mRNA-seq time series during the first 48 h of the RA-induced Primitive Endoderm (PrE) differentiation process in F9 embryonal carcinoma (EC) cells. We show here that this dual regulation is associated with RAR/RXR genomic redistribution during the differentiation process. In-depth analysis of RAR/RXR binding sites occupancy dynamics and composition show that in undifferentiated cells, RAR/RXR interact with genomic regions characterized by binding of pluripotency-associated factors and high prevalence of the non-canonical DR0-containing RA response element. By contrast, in differentiated cells, RAR/RXR bound regions are enriched in functional Sox17 binding sites and are characterized with a higher frequency of the canonical DR5 motif. Our data offer an unprecedentedly detailed view on the action of RA in triggering pluripotent cell differentiation and demonstrate that RAR/RXR action is mediated via two different sets of regulatory regions tightly associated with cell differentiation status.

The visceromotor and somatic afferent nerves of the penis.

INTRODUCTION: Innervation of the penis supports erectile and sensory functions. AIM: This article aims to study the efferent autonomic (visceromotor) and afferent somatic (sensory) nervous systems of the penis and to investigate how these systems relate to vascular pathways. METHODS: Penises obtained from five adult cadavers were studied via computer-assisted anatomic dissection (CAAD). MAIN OUTCOME MEASURES: The number of autonomic and somatic nerve fibers was compared using the Kruskal-Wallis test. RESULTS: Proximally, penile innervation was mainly somatic in the extra-albugineal sector and mainly autonomic in the intracavernosal sector. Distally, both sectors were almost exclusively supplied by somatic nerve fibers, except the intrapenile vascular anastomoses that accompanied both somatic and autonomic (nitrergic) fibers. From this point, the neural immunolabeling within perivascular nerve fibers was mixed (somatic labeling and autonomic labeling). Accessory afferent, extra-albugineal pathways supplied the outer layers of the penis. CONCLUSIONS: There is a major change in the functional type of innervation between the proximal and distal parts of the intracavernosal sector of the penis. In addition to the pelvis and the hilum of the penis, the intrapenile neurovascular routes are the third level where the efferent autonomic (visceromotor) and the afferent somatic (sensory) penile nerve fibers are close. Intrapenile neurovascular pathways define a proximal penile segment, which guarantees erectile rigidity, and a sensory distal segment.