Lipid Deposition Profiles Influence Foreign Body Responses.

Fibrosis remains a significant cause of failure in implanted biomedical devices and early absorption of proteins on implant surfaces has been shown to be a key instigating factor. However, lipids can also regulate immune activity and their presence may also contribute to biomaterial-induced foreign body responses (FBR) and fibrosis. Here it is demonstrated that the surface presentation of lipids on implant affects FBR by influencing reactions of immune cells to materials as well as their resultant inflammatory/suppressive polarization. Time-of-flight secondary ion mass spectroscopy (ToF-SIMS) is employed to characterize lipid deposition on implants that are surface-modified chemically with immunomodulatory small molecules. Multiple immunosuppressive phospholipids (phosphatidylcholine, phosphatidylinositol, phosphatidylethanolamine, and sphingomyelin) are all found to deposit preferentially on implants with anti-FBR surface modifications in mice. Significantly, a set of 11 fatty acids is enriched on unmodified implanted devices that failed in both mice and humans, highlighting relevance across species. Phospholipid deposition is also found to upregulate the transcription of anti-inflammatory genes in murine macrophages, while fatty acid deposition stimulated the expression of pro-inflammatory genes. These results provide further insights into how to improve the design of biomaterials and medical devices to mitigate biomaterial material-induced FBR and fibrosis.

Clinically translatable cytokine delivery platform for eradication of intraperitoneal tumors.

Proinflammatory cytokines have been approved by the Food and Drug Administration for the treatment of metastatic melanoma and renal carcinoma. However, effective cytokine therapy requires high-dose infusions that can result in antidrug antibodies and/or systemic side effects that limit long-term benefits. To overcome these limitations, we developed a clinically translatable cytokine delivery platform composed of polymer-encapsulated human ARPE-19 (RPE) cells that produce natural cytokines. Tumor-adjacent administration of these capsules demonstrated predictable dose modulation with spatial and temporal control and enabled peritoneal cancer immunotherapy without systemic toxicities. Interleukin-2 (IL2)-producing cytokine factory treatment eradicated peritoneal tumors in ovarian and colorectal mouse models. Furthermore, computational pharmacokinetic modeling predicts clinical translatability to humans. Notably, this platform elicited T cell responses in NHPs, consistent with reported biomarkers of treatment efficacy without toxicity. Combined, our findings demonstrate the safety and efficacy of IL2 cytokine factories in preclinical animal models and provide rationale for future clinical testing in humans.

Overview of Interventional Pulmonology for Radiologists.

Interventional pulmonology is a growing field specializing in minimally invasive procedures of the mediastinum, lungs, airways, and pleura. These procedures have both diagnostic and therapeutic indications and are performed for benign and malignant diseases. Endobronchial US has been combined with transbronchial needle aspiration to extend tissue sampling beyond the airways and into the lungs and mediastinum. Recent innovations extending the peripheral access of bronchoscopy include electromagnetic navigational bronchoscopy and thinner bronchoscopes. An important indication for therapeutic bronchoscopy is the relief of central airway obstruction, which may be severe and life threatening. Techniques for restoring patency of the central airways include mechanical debulking and multiple modalities for ablation, stent placement, and balloon bronchoplasty. Bronchoscopic lung volume reduction improves quality of life in certain patients with severe emphysema and is an important less invasive alternative to lung volume reduction surgery. Bronchial thermoplasty is likewise a nonpharmacologic treatment in patients with severe uncontrolled asthma. Many of these procedures have unique selection criteria that require precise evaluations at preprocedure imaging. Postprocedure imaging is also essential in determining outcome success and the presence of complications. Radiologists should be familiar with these procedures as well as the relevant imaging features in both planning and later surveillance. Evolving techniques that may become more widely available in the near future include robotic-assisted bronchoscopy, bronchoscopic transparenchymal nodule access, transbronchial cryobiopsy, ablation of early-stage cancers, and endobronchial intratumoral chemotherapy. An invited commentary by Wayne et al is available online. ©RSNA, 2021.

Assessment of Cancer Survivorship Training and Knowledge Among Resident Physicians in Obstetrics and Gynecology.

The number of gynecologic cancer survivors in the USA is expected to grow to nearly 2 million by 2029. Gynecologic oncologists alone will not be able to care for all of these women. Thus, preparation of general obstetrician/gynecologists (OBGYNs) to deliver this care is crucial. Our objective was to assess cancer survivorship training (CST) among OB/GYN residents and to evaluate knowledge in basic gynecologic cancer survivorship. OB/GYN residents were recruited nationally to complete a de novo questionnaire, querying demographics, hours of CST received, teaching methods used, and efficacy of those methods. Survivorship knowledge was assessed by ten questions based on the 2017 Society of Gynecologic Oncology recommendations on post-treatment surveillance, which includes topics appropriate for generalists. Analyses were done using t tests and ANOVA, with significance set at p = 0.05. In total, 128 residents responded to the survey. Observation was the most common method of CST (53%), with patient contact reported as the most effective method (42.6%). The mean score of correct responses (MSCRs) among all respondents was 61.5%. MSCR significantly improved with increasing post-graduate year (PGY) (p = 0.003). Survivorship training method was not associated with improved MSCR. Improvements in MSCR were observed with an increasing number of CST hours (p = 0.011). A total of 13.9% of residents reported feeling "very comfortable" with survivorship care, yet 88.5% of respondents indicated they did not want additional CST. More hours of CST are associated with improved resident in knowledge in cancer survivorship care, though deficits still remain. Further investigation into optimizing CST is warranted.

Tryptophan degradation is associated with risk-taking propensity in methamphetamine users with treated HIV infection.

Few studies have examined neuroimmune pathways that could contribute to impulsivity in people living with HIV who use substances. Eighty-four methamphetamine-using, sexual minority men with an undetectable HIV viral load were administered the Balloon Analogue Risk Task (BART), a behavioral measure of risk-taking propensity. We examined the associations between kynurenine/tryptophan ratio and phenylalanine/tyrosine ratio with BART scores using multiple linear regression. A higher kynurenine/tryptophan ratio was independently associated with greater BART scores (beta = 0.25; 95% CI = 0.05-1.23; p = 0.034). The phenylalanine/tyrosine ratio was not significantly associated with BART scores. Findings support the need for further research to elucidate the neuroimmune mechanisms linking tryptophan degradation with impulsivity to catalyze the development novel pharmacologic treatments for people living with HIV who use methamphetamine.

Erratum: Ocular toxicities associated with targeted anticancer agents: an analysis of clinical data with management suggestions.

[This corrects the article DOI: 10.18632/oncotarget.17634.].

Ocular toxicities associated with targeted anticancer agents: an analysis of clinical data with management suggestions.

Ocular toxicities are among the most common adverse events resulting from targeted anticancer agents and are becoming increasingly relevant in the management of patients on these agents. The purpose of this study is to provide a framework for management of these challenging toxicities based on objective data from FDA labels and from analysis of the literature. All oncologic drugs approved by the FDA up to March 14, 2015, were screened for inclusion. A total of 16 drugs (12 small-molecule drugs and 4 monoclonal antibodies) were analyzed for ocular toxicity profiles based on evidence of ocular toxicity. Trials cited by FDA labels were retrieved, and a combination search in Medline, Google Scholar, the Cochrane database, and the NIH Clinical Trials Database was conducted. The majority of ocular toxicities reported were low severity, and the most common were conjunctivitis and "visual disturbances." However, severe events including incidents of blindness, retinal vascular occlusion, and corneal ulceration occurred. The frequency and severity at which ocular toxicities occur merits a more multidisciplinary approach to managing patients with agents that are known to cause ocular issues. We suggest a standardized methodology for referral and surveillance of patients who are potentially at risk of severe ocular toxicity.

Efficacy of the Modified Broström Procedure for Adolescent Patients With Chronic Lateral Ankle Instability.

BACKGROUND AND PURPOSE: Lateral ankle sprains are very common, representing up to 30% of sports-related injuries. The anterior talofibular ligament (ATFL) and less commonly the calcaneofibular ligament (CFL) are injured. Surgical treatment is reserved for injuries that fail nonoperative treatment with recurrent instability. Anatomic repair using the modified Broström technique has been shown to have good clinical outcomes in the adult population. The purpose of this study was to report on the outcomes of the modified Broström technique in the pediatric and adolescent population (under 18 y old) for chronic lateral ankle instability. METHODS: Thirty-one patients over an 8-year period were included in the current study after excluding for congenital malformation or underlying connective tissue disease. All patients were treated with a modified Broström technique in which the ATFL was repaired anatomically. Twenty-four patients (77%) underwent concomitant arthroscopy for intra-articular pathology. Demographic, surgical, and clinical data were collected and outcome scores were obtained, including the Marx activity scale, University of California, Los Angeles (UCLA) activity score, and modified American Orthopedic Foot and Ankle Society (AOFAS) score. RESULTS: Mean time from initial injury to surgery averaged 27 months with an overall mean clinical postoperative follow-up of 36 months. Of the 24 patients who underwent concomitant arthroscopy, all had thickening of Bassett ligament and 3 (12.5%) had cartilage lesions. Postoperatively, the mean Marx activity score was 9.9±4.7, mean UCLA score was 9.3±1.3, and mean modified AOFAS score was 83.8±11.7. 71% (22 of 31) of patients achieved good-to-excellent results (as defined by a modified AOFAS score of 80 or greater). Two patients had superficial wound infections; no other complications were experienced in this cohort. CONCLUSIONS: Lateral ankle sprains are common injuries that can frequently be treated nonoperatively; chronic instability may result despite appropriate therapy. Surgical treatment with anatomic repair of the ATFL and CFL using the modified Broström technique in pediatric and adolescent patients results in improved stability, low complication rate, and good clinical outcome scores. LEVEL OF EVIDENCE: Level IV-prognostic retrospective case series.

Designing to the Test: Performance Standards and Technological Change in the U.S. Automobile after 1966.

Performance standards have played an important part in the regulation of technology for many years, yet they have received little attention from historians. This article presents a three-part framework of analysis for examining the role of regulatory performance standards in technological change. Parts one and two of the framework focus on how regulatory agencies construct and enforce standards for technological risks, or "negative externalities." The final part addresses the important question of whether and how firms choose to internalize these standards in their design procedures and other organizational routines. The article provides examples from the history of automotive regulation in the United States to illustrate each part of the framework, and it gives some thoughts on how regulatory performance standards fit within the voluminous literature on standardization, including other work on de facto and voluntary consensus standards.

Eleven microsatellites in an emerging invader, Phytolacca americana (Phytolaccaceae), from its native and introduced ranges.

PREMISE OF THE STUDY: To facilitate population genetic analyses, microsatellite markers were developed for pokeweed (Phytolacca americana), a large, weedy, perennial herb native to eastern North America that is emerging as a significant invasive species in China. METHODS AND RESULTS: We mined 1,100,538 Illumina MiSeq reads from genomic DNA for microsatellites and identified 58 primer pairs. We screened these primers for polymorphism in two native and two invasive populations. We identified 11 loci that amplified consistently. The number of alleles per locus ranged from two to six, and observed heterozygosity ranged from 0.00 to 1.00. All loci were largely monomorphic within populations but different among populations. The primers were of very limited use in the congener P. acinosa. CONCLUSIONS: These loci will provide a valuable resource to study the population genetics and invasion history of P. americana.

Surgical anatomy of the sternoclavicular joint: a qualitative and quantitative anatomical study.

BACKGROUND: The quantitative anatomical relationships of the main ligamentous, tendinous, and osseous structures of the sternoclavicular joint have not been widely investigated. The purpose of this study was to provide a quantitative description of the sternoclavicular joint in relation to relevant surgical landmarks. METHODS: We dissected eleven nonpaired, fresh-frozen cadaveric sternoclavicular joints from four men and seven women (mean age at death, fifty-three years; range, thirty-three to sixty-four years) and measured the ligaments, musculature, and osseous landmarks with use of a three-dimensional coordinate-measuring device. RESULTS: The clavicular pectoralis ridge, located at the 9:30 clock-face position on a right clavicle, served as a reliable osseous landmark for reference to the soft-tissue attachments around the sternoclavicular joint. The costoclavicular ligament was the largest ligament of the sternoclavicular joint, with 80% greater footprint area than that of the posterior sternoclavicular ligament. Articular cartilage covered 67% of the medial end of the clavicle and was located anteroinferiorly. The sternohyoid muscle inserted directly over the posterior sternoclavicular joint and the medial end of the clavicle, whereas the sternothyroid muscle inserted 9.5 mm inferior to the posterior-superior articular margin of the manubrium and coursed 19.8 mm laterally along the first rib. An avascular plane that can serve as a "safe zone" for posterior dissection was observed in each specimen, posterior to the sternoclavicular joint and anterior to the sternohyoid and sternothyroid muscles. CONCLUSIONS: The clavicular pectoralis ridge can be used as an intraoperative guide for clavicle orientation and tunnel placement in sternoclavicular ligament reconstruction. Sternoclavicular joint resection arthroplasty should avoid injuring the costoclavicular ligament, which is the largest sternoclavicular joint ligament. Resection of only the anteroinferior aspect of the medial end of the clavicle may provide adequate decompression while preserving the stability of the clavicle. The location of the sternohyoid and sternothyroid musculotendinous insertions appear to provide a "safe zone" for posterior clavicle and manubrial dissection.

Arthroscopic sternoclavicular joint resection arthroplasty: a technical note and illustrated case report.

Open resection arthroplasty of the sternoclavicular (SC) joint has historically provided good long-term results in patients with symptomatic osteoarthritis of the SC joint. However, the procedure is rarely performed because of the risk of injury to vital mediastinal structures and concern regarding postoperative joint instability. Arthroscopic decompression of the SC joint has therefore emerged as a potential treatment option because of many recognized advantages including minimal tissue dissection, maintenance of joint stability, avoidance of posterior SC joint dissection, expeditious recovery, and improved cosmesis. There are, however, safety concerns given the proximity of neurovascular structures. In this article we demonstrate a technique for arthroscopic SC joint resection arthroplasty in a 26-year-old active man with bilateral, painful, idiopathic degenerative SC joint osteoarthritis. This case also highlights the pearls and pitfalls of arthroscopic resection arthroplasty for the SC joint. There were no perioperative complications. Four months postoperatively, the patient had returned to full activities, including weightlifting, without pain or evidence of SC joint instability. One year postoperatively, the patient showed substantial improvements in the American Shoulder and Elbow Surgeons score; Single Assessment Numeric Evaluation score; Quick Disabilities of the Arm, Shoulder and Hand score; and Short Form 12 Physical Component Summary score over preoperative baseline values.

Clinical and structural outcomes after arthroscopic single-row versus double-row rotator cuff repair: a systematic review and meta-analysis of level I randomized clinical trials.

BACKGROUND: The purpose of this study was to perform a systematic review and meta-analysis of all available level I randomized controlled trials comparing single-row with double-row repair to statistically compare clinical outcomes and imaging-diagnosed re-tear rates. METHODS: A literature search was undertaken to identify all level I randomized controlled trials comparing structural or clinical outcomes after single-row versus double-row rotator cuff repair. Clinical outcomes measures included in the meta-analysis were the American Shoulder and Elbow Surgeons, University of California-Los Angeles, and Constant scores; structural outcomes included imaging-confirmed re-tears. Meta-analyses compared raw mean differences in outcomes measures and relative risk ratios for imaging-diagnosed re-tears after single-row or double-row repairs by a random-effects model. RESULTS: The literature search identified a total of 7 studies that were included in the meta-analysis. There were no significant differences in preoperative to postoperative change in American Shoulder and Elbow Surgeons, University of California-Los Angeles, or Constant scores between the single-row and double-row groups (P = .440, .116, and .156, respectively). The overall re-tear rate was 25.9% (68/263) in the single-row group and 14.2% (37/261) in the double-row group. There was a statistically significant increased risk of sustaining an imaging-proven re-tear of any type in the single-row group (relative risk, 1.76 [95% confidence interval, 1.25-2.48]; P = .001), with partial-thickness re-tears accounting for the majority of this difference (relative risk, 1.99 [95% confidence interval, 1.40-3.82]; P = .039). CONCLUSION: Single-row repairs resulted in significantly higher re-tear rates compared with double-row repairs, especially with regard to partial-thickness re-tears. However, there were no detectable differences in improvement in outcomes scores between single-row and double-row repairs.

Posterior sternoclavicular joint injuries in skeletally immature patients.

BACKGROUND: Posterior injuries to the sternoclavicular (SC) joint are uncommon. In the skeletally immature (SI) population, these injuries have been described as either dislocations of the SC joint or fractures of the medial clavicular physis. The current literature and standardized test questions state that a posterior SC injury, in a SI patient, is more likely a physeal fracture than a SC joint dislocation. However, this injury characterization is based on case reports or small case series. The purpose of this study is to characterize posterior SC injuries in SI patients in terms of the prevalence of dislocation versus medial clavicle physeal fracture. METHODS: A retrospective review was performed of 48 SI patients treated for posterior SC joint injuries over a 20-year period with a mean age of 15.4 years (range, 13 to 18 y). Forty patients underwent open reduction and internal fixation as their definitive treatment and 8 patients were treated exclusively with closed reduction. Patients treated operatively were utilized in determining the prevalence of SC joint dislocation versus physeal fracture. RESULTS: All patients treated operatively underwent primary repair without reconstruction. Twenty (50%), of the 40 patients treated operatively had a true SC joint dislocation and 20 patients (50%) had a medial clavicle physeal fracture. Twenty-two (46%) of the 48 total patients had an attempted closed reduction of which only 8 (36%) were successful. Among the 14 unsuccessful closed reductions, 12 (86%) were true dislocations (P<0.001). All successful closed reductions occurred in patients within 24 hours from injury. Eleven of the 48 (23%) patients' injuries were missed on initial presentation. CONCLUSIONS: Posterior SC joint dislocation and medial clavicular physeal fracture both occur with roughly equivalent prevalence in patients with an open medial physis. An attempted closed reduction may be more successful if performed within 24 hours after injury. Patients who fail attempts at closed reduction are more likely to have a posterior SC joint dislocation than a physeal fracture. Posterior SC joint injury may be missed in nearly 25% of patients on initial presentation. LEVEL OF EVIDENCE: Level IV-retrospective case series.

Role of preoperative computed tomography scans in operative planning for malleolar ankle fractures.

BACKGROUND: There remains no consensus regarding the role of computed tomography (CT) scans in preoperative planning for malleolar ankle fractures. The aim of this study was to determine the role of preoperative CT scans on operative planning in these fractures. METHODS: A retrospective analysis was performed on 100 consecutive patients treated at our institution for malleolar ankle fractures (AO type 44) with both preoperative radiographs and CT scans. Six study participants reviewed available radiographs and formulated an operative (or nonoperative) plan including positioning, operative approach, and fixation. Participants then analyzed CT scans of the same fractures, deciding whether (and how) they would alter operative strategy. Characteristics of fractures and radiographs were correlated with changes in operative strategy. RESULTS: Operative strategy was notably changed in 24% of cases after CT review, with strong intraclass correlation (0.733). Common changes included alterations in medial malleolar (21%) or posterior malleolar (15%) fixation and fixation of an occult anterolateral plafond fracture (9%). Notable predictors of changes in operative strategy included trimalleolar over unimalleolar fractures (29% vs 10% rate of change), preoperative dislocation over no dislocation (31% vs 20%), the presence of only radiographs with overlying plaster versus fractures with at least 1 set of radiographs without plaster (25% vs 14%), and suprasyndesmotic versus trans- and infra-syndesmotic fractures (40% vs 20% and 4%, respectively). CONCLUSIONS: CT scans may be useful adjuncts in preoperative planning for malleolar ankle fractures, most notably in fracture dislocations, cases in which all available radiographs are obscured by plaster, trimalleolar fractures, and suprasyndesmotic fractures. LEVEL OF EVIDENCE: Level III, retrospective comparative study.

Salvage esophagectomy after failed definitive chemoradiation for esophageal adenocarcinoma.

BACKGROUND: Outcomes of salvage esophagectomy after definitive chemoradiation (CRT) for squamous cell carcinoma are well defined. Previous reports of salvage esophagectomy in patients with recurrent adenocarcinoma after definitive CRT are limited by small numbers and high morbidity and mortality rates. METHODS: We reviewed our experience of 65 patients with esophageal adenocarcinoma treated from 1997 to 2010 who underwent salvage esophagectomy after failed definitive CRT. We then compared this group to 65 matched patients of 521 total patients with esophageal adenocarcinoma who received preoperative CRT followed by planned esophagectomy. Propensity matching and multivariable analysis were performed. RESULTS: Median time to surgery from completion of therapy for the salvage group was 216 days. Major postoperative events (major pulmonary event, conduit loss, leak, readmission to intensive care unit) occurred in 35% (23 of 65) of salvage patients and 31% (20 of 65) of the planned resection matched group. Anastomotic leak occurred in 18.5% (12 of 65) and 11.3 (59 of 521) of salvage and planned groups, respectively. Thirty-day mortality was 3.1% (2 of 65) after salvage resection and 4.6% (3 of 65) after planned resection. There was no difference in 3-year overall or median survival between the two groups of patients (32 months, 48% salvage, versus 40 months, 57% planned resection). Multivariable analysis did not identify salvage strategy or time from completion of therapy to resection as a predictor of major event or death. CONCLUSIONS: Postoperative morbidity, mortality, and overall survival of patients after salvage esophagectomy are comparable to matched patients after planned resection. These results suggest that patients with esophageal adenocarcinoma who fail definitive CRT and recur locoregionally should be considered for salvage esophagectomy at experienced esophageal centers.

Internet information quality for ten common foot and ankle diagnoses.

BACKGROUND: Patients use the Internet regularly to access health-related information. This study's goal was to assess the quality and content of Internet-based information for common foot and ankle diagnoses. METHODS: We identified the ten most common foot and ankle diagnoses in our academic foot and ankle practice. Ten websites for each diagnosis were identified using two large Internet search engines. A custom grading form was used to determine website quality, based upon the Health On the Net Foundation (HON) principles, and information content. Four independent reviewers graded each website. RESULTS: One hundred thirty-six unique websites were reviewed. Average HON score was 62.4 (range, 52.3-68.8) and content score was 49.7 (range, 33.8-62.1) out of a maximum of 100. Interobserver variability was low. CONCLUSIONS: The overall quality of Internet information for common foot and ankle diagnoses is variable, raising concerns about what information is currently available to patients.

Neoadjuvant chemoradiotherapy followed by surgery for esophageal adenocarcinoma: significance of microscopically positive circumferential radial margins.

OBJECTIVES: The incidence and consequence of an isolated involved circumferential radial margin (CRM) after resection for esophageal adenocarcinoma in the setting of neoadjuvant chemoradiotherapy (CRT) has not been reported. We aimed to determine the frequency and significance of a close (<1 mm) or involved CRM in patients undergoing esophagectomy after CRT. METHODS: We retrospectively analyzed the data from patients undergoing resection from 1997 to 2008 for esophageal adenocarcinoma after neoadjuvant CRT. A positive CRM was defined as microscopic tumor at or less than 1 mm of the radial margin. An R1 resection was tumor at the radial margin. Only patients with ypT3 or greater tumors were included. R2 resections were excluded. Statistical comparisons were performed using Cox regression and Kaplan-Meier analyses. RESULTS: A total of 160 patients met the inclusion criteria, 42 (26%) had a positive CRM. The median survival did not significantly differ between the CRM-negative and -positive groups (28 vs 50 months, P = .84). A propensity score matching analysis also failed to find a significant difference in outcomes. When analyzed by tumor present at the margin (R1), R0 patients had a longer median survival compared with R1 patients (28 vs 8 months, P = .01). This difference, however, was not seen on propensity score matching. CONCLUSIONS: Resections of locally advanced esophageal adenocarcinoma with residual transmural viable tumor after CRT frequently showed involvement of the radial margin with tumor either close to or at the margin. Tumor close (<1 mm) to the radial margin did not result in a significant decrease in overall or disease-free survival or increase in local recurrence.

A transgenic wnt8a:PAC reporter reveals biphasic regulation of vertebrate mesoderm development.

Vertebrate wnt8a links anteroposterior and dorsoventral axis patterning, but the regulation of wnt8a expression and its relationship to mesoderm induction and maintenance pathways is unclear. To address this, we have generated zebrafish transgenic for a modified genomic PAC clone that expresses EGFP from the wnt8a locus. The EGFP reporter transgene is expressed in a pattern nearly identical to wnt8a, including maternal deposition, expression in the ventrolateral mesoderm and in the yolk syncytial layer. Loss of function studies show that wnt8a expression is under biphasic control by Nodal and No Tail/Brachyury, whereby early phase expression is Nodal-dependent but late phase expression is Ntl/Bra dependent. EGFP fluorescence persists in cells that transcribe the reporter, thus comprising a tracer for ventrolaterally derived mesodermal lineages. We use this property to show that wnt8a expression marks Nodal-independent tail mesoderm formation and that Ntl/Bra predominantly regulates wnt8a in paraxial mesoderm progenitors.