NR5A2 connects zygotic genome activation to the first lineage segregation in totipotent embryos.

Zygotic genome activation (ZGA) marks the beginning of the embryonic program for a totipotent embryo, which gives rise to the inner cell mass (ICM) where pluripotent epiblast arises, and extraembryonic trophectoderm. However, how ZGA is connected to the first lineage segregation in mammalian embryos remains elusive. Here, we investigated the role of nuclear receptor (NR) transcription factors (TFs), whose motifs are highly enriched and accessible from the 2-cell (2C) to 8-cell (8C) stages in mouse embryos. We found that NR5A2, an NR TF strongly induced upon ZGA, was required for this connection. Upon Nr5a2 knockdown or knockout, embryos developed beyond 2C normally with the zygotic genome largely activated. However, 4-8C-specific gene activation was substantially impaired and Nr5a2-deficient embryos subsequently arrested at the morula stage. Genome-wide chromatin binding analysis showed that NR5A2-bound cis-regulatory elements in both 2C and 8C embryos are strongly enriched for B1 elements where its binding motif is embedded. NR5A2 was not required for the global opening of its binding sites in 2C embryos but was essential to the opening of its 8C-specific binding sites. These 8C-specific, but not 2C-specific, binding sites are enriched near genes involved in blastocyst and stem cell regulation, and are often bound by master pluripotency TFs in blastocysts and embryonic stem cells (ESCs). Importantly, NR5A2 regulated key pluripotency genes Nanog and Pou5f1/Oct4, and primitive endoderm regulatory genes including Gata6 among many early ICM genes, as well as key trophectoderm regulatory genes including Tead4 and Gata3 at the 8C stage. By contrast, master pluripotency TFs NANOG, SOX2, and OCT4 targeted both early and late ICM genes in mouse ESCs. Taken together, these data identify NR5A2 as a key regulator in totipotent embryos that bridges ZGA to the first lineage segregation during mouse early development.

Maturation of Nephrons by Implanting hPSC-derived Kidney Progenitors Under Kidney Capsules of Unilaterally Nephrectomized Mice.

BACKGROUND: Human pluripotent stem cell (hPSC)-derived kidney organoids may contribute to disease modeling and the generation of kidney replacement tissues. However, the realization of such applications requires the induction of hPSCs into functional mature organoids. One of the key questions for this process is whether a specific vascular system exists for nephrogenesis. Our previous study showed that short-term (2 weeks) implantation of hPSC-derived organoids below the kidney capsules of unilaterally nephrectomized and immunodeficient mice resulted in the enlargement of organoids and production of vascular cells, although signs of maturation were lacking. METHODS: Organoids were induced for 15 days in vitro and then grafted below kidney capsules of the same unilaterally nephrectomized immunodeficient mouse model to examine whether medium-term (4 weeks) implantation could improve organoid maturation and vascularization, as evaluated by immunofluorescence and transmission electron microscopy. RESULTS: We demonstrated that after 2-4 weeks of implantation, renal organoids formed host-derived vascularization and matured without any exogenous vascular endothelial growth factor. Glomerular filtration barrier maturation was evidenced by glomerular basement membrane deposition, perforated glomerular endothelial cell development, and apical, basal podocyte polarization. A polarized monolayer epithelium and extensive brush border were also observed for tubular epithelial cells. CONCLUSIONS: Our results indicate that the in vivo microenvironment is important for the maturation of human kidney organoids. Stromal expansion and a reduction of nephron structures were observed following longer-term (12 weeks) implantation, suggesting effects on off-target cells during the induction process. Accordingly, induction efficiency and transplantation models should be improved in the future.

Resection of the intracavernous sinus tumors using a purely endoscopic endonasal approach.

Resection of the cavernous sinus (CS) lesions has been a surgical challenge because the anatomy of the CS presents a high grade of complexity. This report describes the feasibility of the purely endoscopic endonasal approach to the CS. Twenty-five patients with intracavernous sinus tumors were treated with a purely endoscopic endonasal approach. The indications, efficacy, surgical techniques, and complications of this approach were discussed. Gross total resection occurred in 19 cases (76%), subtotal resection occurred in 2 cases (8%), and partial resection occurred in 4 cases (16%) including pituitary adenoma in 10 cases (total 70%; subtotal 10%; partial 20%), meningioma in 6 cases (total 66.6%; subtotal 16.7%, partial 16.7%), schwannoma in 5 cases (100%, total 5), malignant tumor in 4 cases (total 75%; subtotal 25%). All patients experienced resolution or improvement of symptoms. No patient experienced intraoperative complication and new neurological deficit. Only 1 case of postoperative cerebrospinal fluid leakage repaired via endoscopic endonasal approach on the 14th day after the surgery. The purely endoscopic endonasal approach to the CS in appropriately evaluated patients can be used to address a wide variety of benign and malignant tumor pathology with favorable outcomes and a low incidence of complications.

Transoral endoscopic odontoidectomy to decompress the cervicomedullary junction.

STUDY DESIGN: Clinical study. OBJECTIVE: To investigate the feasibility of the transoral endoscopic odontoidectomy without occipitocervical fusion. SUMMARY OF BACKGROUND DATA: Endoscopic transnasal resection of the odontoid process is less invasive than the conventional transoral odontoidectomy. However, the endonasal approach has a much longer working distance compared with the transoral approach to the craniovertebral junction and usually the endonasal approach needs a previous occipitocervical posterior fusion. METHODS: From July 2007 to June 2010, 5 patients (3 males and 2 females, age range, 25-41 yr) with irreducible cervicomedullary junction compression were subjected to endoscopic transoral odontoidectomy without occipitocervical posterior fixation and bone fusion. RESULTS: A purely endoscopic transoral odontoidectomy for decompression of the cervicomedullary junction without the occipitocervical fusion was achieved successfully in 5 patients. None of the patients underwent tracheotomy and postoperative gastrostomy tube placement. The patients were started on liquids on the third postoperative day and advanced to a regular diet on the fourth postoperative day. There was no postoperative velopharyngeal insufficiency, cerebrospinal fluid leakage, regional infection, or meningitis. The patients were discharged in 10 to 12 days after the surgery. There were no evidence of instability at the craniovertebral junction at 12 to 47 months of follow-up and remarkable improvement in neurological function was observed in each patient. CONCLUSION: The endoscopic transoral approach may be a more direct route to C1 and the odontoid than the endoscopic endonasal approach. This approach allows complete resection odontoid to decompress the cervicomedullary junction without increasing the risk of complications such as wound infection, meningitis, and velopharyngeal insufficiency. Usually, the occipitocervical posterior fusion and tracheotomy is less necessary in this approach.

Anatomy of the petrous apex as related to the endoscopic endonasal approach.

The endoscopic endonasal approach (EEA) has been reported to be an efficient approach for treating lesions of the petrous apex. However, there have been only limited anatomic studies for the EEA. Furthermore, most of the relevant distances for EEA cannot be measured easily on a cadaveric skull. Two fresh adult cadaver heads and five formalin-fixed adult cadaver heads were dissected using the EEA to identify groups of landmarks for safe guidance during this approach. The distances between these landmarks were then measured by CT angiography by using three-dimensional software. The EEA to the petrous apex can be divided into five phases. In each phase, a group of landmarks, rather than a single landmark, can be identified easily for guiding the next phase of the approach. There was no significant difference between males and females in any of the distances reported in the present study. The EEA can be performed to manage a petrous apex lesion more safely by referring to multiple landmarks and the distances between them.

Endoscopic endonasal management of trigeminal schwannomas extending into the infratemporal fossa.

Extracranial trigeminal schwannomas extending into the infratemporal fossa are rare. The traditional surgical approaches to the infratemporal fossa are associated with complications, such as facial nerve dysfunction, hearing loss, dental malocclusion and cosmetic problems. We report eight patients (four males, four females, age range=31-62 years) who were treated between 2004 and 2009 for extracranial trigeminal schwannomas extending into the infratemporal fossa. Schwannomas were surgically removed using a purely endoscopic endonasal approach. The maximum diameters of the tumours ranged from 30 mm to 70 mm and all tumours were completely removed. There were no intraoperative or postoperative complications in this series. There were no recurrences during the follow-up period which ranged from 10 to 74 months (mean=30 months). The purely endoscopic endonasal approach may provide a minimally invasive and safe approach to remove extracranial trigeminal schwannomas extending into the infratemporal fossa. Radical resection was associated with an excellent long-term outcome in this series.

Transclival cerebrospinal fluid rhinorrhea as the initial presenting symptom of a tiny intradural chordoma.

We report a patient with a tiny intradural clival chordoma, which was identified following presentation with cerebrospinal fluid (CSF) rhinorrhea as the initial symptom. The transclival dural defect and the intradural tumor were successfully localized by both radiological investigation and intraoperative endoscopic inspection. The tumor was totally resected and the CSF fistula was repaired by an endoscopic endonasal approach. The diagnosis, possible mechanisms and management of this rare condition are discussed. The role of endoscopy in identifying and treating the clival CSF rhinorrhea is emphasized. To our knowledge, this is the first report of a clival fistula secondary to a tiny intradural chordoma.

Identification of an inhibitory cis-element in the promoter region of human homeobox gene NKX3.1.

BACKGROUND: NKX3.1 is a prostate-specific homeobox gene related strongly to prostate development and prostate cancer. To study its transcriptional regulation, a 1,040 bp promoter of human NKX3.1 gene was cloned into the upstream of the luciferase reporter gene in pGL3-basic plasmid and a 180 bp region extending from -391 to -212 in the upstream of NKX3.1 gene was identified presenting an inhibitory regulation for NKX3.1 expression in our previous experiments. In this work, it is aimed to identify precisely the functional cis-element within the 180 bp region involved in the inhibitory regulation of the NKX3.1 promoter and to identify its specific binding protein. METHODS: 5'-deletion mutation, substitution mutation, and electrophoresis mobility shift assay (EMSA) were carried out to identify precisely the functional cis-element contributing to the inhibitory regulation of NKX3.1 and its binding ability to nuclear extracts from prostate cancer cell line LNCaP. RESULTS: A 20-bp inhibitory element located between -362 and -343 in the upstream of NKX3.1 gene was identified by deletion and substitution mutation analysis and proven to be a functional inhibitory cis-element by EMSA. CONCLUSIONS: We have identified a functional inhibitory cis-element between -362 and -343 in the upstream of NKX3.1 gene; it is likely to play an important role in downregulating NKX3.1 gene transcription.