Histone demethylase JMJD3 contributes to epigenetic control of INK4a/ARF by oncogenic RAS.

The INK4a/ARF tumor suppressor locus, a key executor of cellular senescence, is regulated by members of the Polycomb group (PcG) of transcriptional repressors. Here we show that signaling from oncogenic RAS overrides PcG-mediated repression of INK4a by activating the H3K27 demethylase JMJD3 and down-regulating the methyltransferase EZH2. In human fibroblasts, JMJD3 activates INK4a, but not ARF, and causes p16(INK4a)-dependent arrest. In mouse embryo fibroblasts, Jmjd3 activates both Ink4a and Arf and elicits a p53-dependent arrest, echoing the effects of RAS in this system. Our findings directly implicate JMJD3 in the regulation of INK4a/ARF during oncogene-induced senescence and suggest that JMJD3 has the capacity to act as a tumor suppressor.

Ulnar nerve innervation of the triceps muscle: real or apparent? An anatomic study.

BACKGROUND: Since the 18th century, the existence of ulnar nerve innervation of the medial head of the triceps brachii muscle has been controversial. The evidence for or against such innervation has been based on macroscopic dissection, an unsuitable method for studying intraneural topography or intramuscular branching. The study of smaller specimens (embryos or fetuses) by means of serial histologic sections may resolve the controversy. QUESTIONS/PURPOSES: Using fetal specimens and histology we determined the contributions of the ulnar and radial nerves to innervation of the triceps brachii muscle. METHODS: We histologically examined 15 embryonic and fetal arms. Radial nerve branches obtained from six adult arms were analyzed immunohistochemically to determine motor fiber content. RESULTS: The medial head of the triceps brachii muscle was always innervated by the radial nerve (ulnar collateral branch). The branches seeming to leave the ulnar nerve at elbow level were the continuation of the radial nerve that had joined the ulnar nerve sheath via a connection in the axillary region. Immunohistochemistry revealed motor and nonmotor fibers in this radial nerve branch. CONCLUSIONS: A connection between the radial and ulnar nerves sometimes may exist, resulting in an apparent ulnar nerve origin of muscular branches to the medial head of the triceps, even though in all our specimens the fibers could be traced back to the radial nerve. CLINICAL RELEVANCE: Before performing or suggesting new muscle and nerve transpositions using this apparent ulnar innervation, the real origin should be confirmed to avoid failure.

Obturator artery revisited.

INTRODUCTION AND HYPOTHESIS: The aim of this work is to analyse the variability of the obturator artery (oa), unify previous criteria and propose a simple classification for clinical use. METHODS: A sample of 119 adult human embalmed cadavers was used. Origin and course of the oa in relation with the external iliac artery, internal iliac artery and inferior epigastric artery were studied. Chi-squared and t test were used for statistical comparison, and p < 0.05 was considered significant. RESULTS: Based on the number of roots of origin, three different situations were observed. The oa shows a single origin (96.55%). The oa presents a double origin (3.02%), or the oa arises from three roots (0.43%). The first situation was subclassified into six types according to the oa origin. Equal vascular pattern in both hemi-pelvises was observed in 58.93%. CONCLUSIONS: Almost 31% of oa passes over the superior pubic ramus implying an increased risk during some procedures.

The clinical interest of the ary-thyro-cricoid fascicle.

The aim of this work was to study the prevalence and form of the ary-thyro-cricoid (ATC) muscular fascicle, a variable muscular slip connecting the oblique and/or transverse arytenoid muscles with the thyroarytenoid (TA) and/or lateral cricoarytenoid (LCA) muscles resembling a sphincter encircling the glottis. Thirty larynges obtained from necropsies of individuals with no known laryngeal pathology were dissected. The ATC fascicle was observed in 96.7% of the larynges. It appeared bilaterally in 60% of subjects and unilaterally in 36.7%. The posterior attachment of the muscular slip was observed to be in common with either the transverse arytenoid (34%), or the oblique arytenoid (28%) or both muscles (38%). Its fibers terminated by intermingling with either those of the LCA muscle (10.6%), or the TA muscle (38.3%) or both (51.1%). These variable attachments mean that there are nine possible variants of this muscular fascicle. The ATC fascicle was supplied by branches originating bilaterally from the recurrent laryngeal (RLN) and internal laryngeal nerves. The existence of this ATC fascicle could explain the variable position (intermediate, paramedian or lateral) adopted by the vocal folds after lesion of the RLN. The bilateral disposition and innervation of the fascicle could also complicate the interpretation of electromyographic techniques used for testing laryngeal nerve function.

Popliteal artery: Anatomical study and review of the literature.

BACKGROUND: The frequency of appearance of anatomical variability in the terminal division of the popliteal artery (PA) is different according to the type of sample used, and ranges from 2% to 21%. The PA locates 1,01 cm behind to the lateral meniscus, which makes it vulnerable during surgical procedures. Iatrogenic injury of the PA or its terminal branches increases if anatomical variables are present. Our aim was to describe and review the branching pattern of the PA in a body-donors to science sample to determine the influence of the sample used (body-donors vs imaging test). METHODS: A sample consisting of 260 popliteal regions, corresponding to 130 corpses (66 women, 64 men), have been dissected. Multivariate analysis was carried out. RESULTS: The terminal division of the PA was classified as follows: Pattern 1: the PA divided into the anterior tibial (ATA) and the posterior tibial arteries (PTA) at the level or distal to the lower border of the popliteal muscle (PM) (94.7%). Pattern 2: the PA bifurcated into the ATA and PTA, proximal to the lower border of the PM (3.3%). Pattern 3: the PA divided at the same level into the ATA, PTA and PEA. (2%). No significant differences between gender and side of the limb could be find. CONCLUSIONS: We propose a classification that encloses three identifiable groups only. This will allow clinicians to bear in mind these variables easily, at the same time avoiding injuries during surgical procedures such as lateral meniscus repair.

Anatomic study of blood supply of the dorsum of the foot and ankle.

PURPOSE: This project was to study the different patterns of the anterior tibal and dorsalis pedis arteries in relation to the blood supply of the dorsum of the foot and ankle. METHODS: A reliable sample of 150 human embalmed cadavers was dissected. RESULTS: Four different patterns were identified. The dorsalis pedis artery was most frequently (287 cases, 95.7%) found to be the continuation of the anterior tibial artery distal to the ankle, and lay between the tendon of extensor hallucis and the first tendon of extensor digitorum longus. The other 13 cases (4.3%) showed 3 variant patterns of the anterior tibial-dorsalis pedis vascular axis: the anterior tibial artery took a more lateral course, passing in front of the lateral malleolus (6 cases, 2%); the perforating branch of the peroneal artery assumed the expected course of the dorsalis pedis artery (4 cases, 1.3%); the anterior tibal artery gave a lateral branch that replaced the perforating branch of the peroneal artery to supply the lateral aspect of the ankle (3 cases, 1%). CONCLUSION: Arterial variations of the anterior tibial-dorsalis pedis axis occurred in almost 5% of cases. CLINICAL RELEVANCE: An awareness of the existence of such variations is helpful during a preoperative assessment and could prevent injury during surgery.

Expression and regulation of ROR-1 during early avian limb development.

ROR-1 is a member of the ROR family of tyrosine kinase like orphan receptors and is highly conserved among various species. We have isolated the chick ROR-1 ( cROR-1) and show that cROR-1 expression is high and restricted to the proximal limb region until HH-stage 25. At later stages, expression spreads towards the distal limb region. In order to determine the signals that control cROR-1 expression, factors known to be involved in limb patterning (FGFs, BMPs, SHH, retinoic acid) were applied to the developing limb. Whereas neither FGFs, BMPs, nor SHH affected cROR-1 expression, upregulation could be achieved by ectopic application of retinoic acid to the distal limb region. As retinoic acid also upregulated retinoic acid receptor beta ( Rar-beta), we assume that cROR-1 upregulation is mediated by Rar-beta. We conclude that ROR-1 signaling is an independently regulated pathway, which is involved in late rather than early limb development.

Spatial and temporal pattern of Wnt-6 expression during chick development.

The WNT family of proteins is composed of several members. In the present study we isolated the full length chick Wnt-6 cDNA and analyzed its expression pattern by in situ hybridization during chick development. Wnt-6 expression is observed in the ectoderm from HH-stage 4 onwards. At HH-stages, 7-16 expression can be seen in the ectoderm overlying the segmental plate and the epithelial somite, while the ectoderm overlying the compartmentalized somite is Wnt-6 negative. Expression is also observed at the heart outflow tract and in the ectoderm overlying the pharyngeal arches. From HH-stages 17 to 27, expression is also observed at limb level, both in the dorsal and ventral ectoderm and a stronger expression in the dorsoventral boundary. Furthermore, expression in the ectoderm delimiting the somitic boundaries in the anteroposterior and mediolateral axis at limb level was observed, as well as in the ventral body wall. Expression becomes evident in the inner ear. From HH-stage 30 onwards, expression is restricted to the feather buds and to the gastrointestinal tract.

Arthroscopic anatomy of the posterior ankle ligaments.

PURPOSE: Our objective was to define the arthroscopic anatomy of the posterior ankle ligaments. TYPE OF STUDY: Anatomic study. METHODS: Twenty fresh-frozen feet were dissected in order to examine the morphology of the posterior ankle ligaments and their variations. In addition, arthroscopic examination was performed in 8 feet with standard anteromedial and anterolateral portals. Posterior ligamentous structures were identified and marked with suture. Following arthroscopy, careful dissection was performed and previously marked anatomic structures were identified. RESULTS: At the time of anatomic dissection, the posteroinferior tibiofibular ligament, also called the tibial slip, was found to be a constant structure with varying size and shape, and well evident during arthroscopy due to its oblique course. The deep component of the posteroinferior tibiofibular ligament, also called the transverse ligament, is well defined during arthroscopy. The 2 ligaments not well visualized during arthroscopy are the superficial component of the posteroinferior tibiofibular ligament and the posterior talofibular ligament. CONCLUSIONS: These findings should be of interest in interpreting findings encountered during arthroscopic examination of ankle, and when a posterior soft-tissue impingement syndrome must be ruled out.

Genome-wide co-localization of Polycomb orthologs and their effects on gene expression in human fibroblasts.

BACKGROUND: Polycomb group proteins form multicomponent complexes that are important for establishing lineage-specific patterns of gene expression. Mammalian cells encode multiple permutations of the prototypic Polycomb repressive complex 1 (PRC1) with little evidence for functional specialization. An aim of this study is to determine whether the multiple orthologs that are co-expressed in human fibroblasts act on different target genes and whether their genomic location changes during cellular senescence. RESULTS: Deep sequencing of chromatin immunoprecipitated with antibodies against CBX6, CBX7, CBX8, RING1 and RING2 reveals that the orthologs co-localize at multiple sites. PCR-based validation at representative loci suggests that a further six PRC1 proteins have similar binding patterns. Importantly, sequential chromatin immunoprecipitation with antibodies against different orthologs implies that multiple variants of PRC1 associate with the same DNA. At many loci, the binding profiles have a distinctive architecture that is preserved in two different types of fibroblast. Conversely, there are several hundred loci at which PRC1 binding is cell type-specific and, contrary to expectations, the presence of PRC1 does not necessarily equate with transcriptional silencing. Interestingly, the PRC1 binding profiles are preserved in senescent cells despite changes in gene expression. CONCLUSIONS: The multiple permutations of PRC1 in human fibroblasts congregate at common rather than specific sites in the genome and with overlapping but distinctive binding profiles in different fibroblasts. The data imply that the effects of PRC1 complexes on gene expression are more subtle than simply repressing the loci at which they bind.

Morphogenesis of the human laryngeal ventricles.

BACKGROUND: Two theories explain the origin of human laryngeal ventricles: (1) ventricles derive from the fifth pharyngeal pouches; (2) development independent from the pouches. METHODS: In all, there were 21 serially sectioned human embryos from stages 15 to 23, and 11 fetuses of 9 to 18 weeks. Computer-aided 3-dimentional reconstructions were made. RESULTS: The cranial part of the laryngeal sulcus and future vestibule expands from the pharyngeal floor between the third and fourth pharyngeal pouches during stages 15 and 16 (33-37 days). The primordia of the ventricles appeared at stage 18 (44 days) as 2 lateral swellings in the caudal end of the future vestibule, limited by the third pharyngeal pouches. Active epithelial expansion and subsequent canalization during late embryonic and early fetal periods finalizes their development. CONCLUSIONS: The laryngeal ventricles do not derive from the pharyngeal pouches but the median region of the pharyngeal floor between the third and fourth pharyngeal pouches and arches at the caudal end of the future vestibule.

Recurrent laryngeal nerve landmarks revisited.

BACKGROUND: The aim of this work was to evaluate, to prove their reliability, the different surgical landmarks previously proposed as a mean to locate the recurrent laryngeal nerve (RLN). METHODS: The necks of 143 (68 male and 76 female) human adult embalmed cadavers were examined. RLN origin and length and its relationship to different landmarks were recorded and results compared with those previously reported. Statistical comparisons were performed using the chi-square test (significance, p ≤ .05). RESULTS: Mostly, RLN is located anterior to the tracheoesophageal sulcus (41.6%), posterior to the inferior thyroid artery (35.8%), lateral to Berry's ligament (88.1%), below the inferior rim of the inferior constrictor muscle (90.4%), and entering the larynx before its terminal division (54.6%). CONCLUSIONS: The position of the RLN in relation to those structures classically considered as landmarks is highly variable. The most reliable relationships are those with Berry's ligament or the inferior constrictor muscle.

Functional role of human laryngeal nerve connections.

OBJECTIVES/HYPOTHESIS: Current knowledge of the functional role of human laryngeal nerves is based on traditional laryngeal neuroanatomic descriptions or contradictory electromyographic studies. The aim of this study was to clarify the functional role of neural connections between laryngeal nerves by correlating the different electromyographic patterns observed after laryngeal stimulation and the existence of different neural connections. STUDY DESIGN: Descriptive. METHODS: Electromyographic and morphologic study in 13 patients during total laryngectomy procedure. RESULTS: Seven patients showed an additional evoked response from the cricothyroid muscle after recurrent laryngeal nerve stimulation. External laryngeal nerve stimulation resulted in additional responses from the posterior cricoarytenoid muscle in three cases and from the arytenoid muscle in one. The presence of a neural connection was confirmed in all patients who showed an unexpected electromyographic response. CONCLUSIONS: The different connections between laryngeal nerves are at least partially of motor nature and play a role in the mobility of vocal folds.

Several distinct polycomb complexes regulate and co-localize on the INK4a tumor suppressor locus.

Misexpression of Polycomb repressive complex 1 (PRC1) components in human cells profoundly influences the onset of cellular senescence by modulating transcription of the INK4a tumor suppressor gene. Using tandem affinity purification, we find that CBX7 and CBX8, two Polycomb (Pc) homologs that repress INK4a, both participate in PRC1-like complexes with at least two Posterior sex combs (Psc) proteins, MEL18 and BMI1. Each complex contains a single representative of the Pc and Psc families. In primary human fibroblasts, CBX7, CBX8, MEL18 and BMI1 are present at the INK4a locus and shRNA-mediated knockdown of any one of these components results in de-repression of INK4a and proliferative arrest. Sequential chromatin immunoprecipitation (ChIP) reveals that CBX7 and CBX8 bind simultaneously to the same region of chromatin and knockdown of one of the Pc or Psc proteins results in release of the other, suggesting that the binding of PRC1 complexes is interdependent. Our findings provide the first evidence that a single gene can be regulated by several distinct PRC1 complexes and raise important questions about their configuration and relative functions.

Tbx1 regulation of myogenic differentiation in the limb and cranial mesoderm.

The T-box transcription factor Tbx1 has been implicated in DiGeorge syndrome, the most frequent syndrome due to a chromosomal deletion. Gene inactivation of Tbx1 in mice results in craniofacial and branchial arch defects, including myogenic defects in the first and second branchial arches. A T-box binding site has been identified in the Xenopus Myf5 promoter, and in other species, T-box genes have been implicated in myogenic fate. Here we analyze Tbx1 expression in the developing chick embryo relating its expression to the onset of myogenic differentiation and cellular fate within the craniofacial mesoderm. We show that Tbx1 is expressed before capsulin, the first known marker of branchial arch 1 and 2 muscles. We also show that, as in the mouse, Tbx1 is expressed in endothelial cells, another mesodermal derivative, and, therefore, Tbx1 alone cannot specify the myogenic lineage. In addition, Tbx1 expression was identified in both chick and mouse limb myogenic cells, initially being restricted to the dorsal muscle mass, but in contrast, to the head, here Tbx1 is expressed after the onset of myogenic commitment. Functional studies revealed that loss of Tbx1 function reduces the number of myocytes in the head and limb, whereas increasing Tbx1 activity has the converse effect. Finally, analysis of the Tbx1-mesoderm-specific knockout mouse demonstrated the cell autonomous requirement for Tbx1 during myocyte development in the cranial mesoderm.

BMPs restrict the position of premuscle masses in the limb buds by influencing Tcf4 expression.

Previous studies have shown the distally retreating source of Scatter factor/Hepatocyte growth factor (SF/HGF) can account for the distal migration of myogenic precursor cells in the limb bud mesenchyme. However, the normal expression pattern of Sf/Hgf alone does not explain the distribution of muscle precursor cells. Hence, the position of the dorsal and ventral premuscle masses suggests the presence of additional patterning factors. We present evidence that BMP2 and 4 can act as such factors by inhibiting the expression of Tcf4, a downstream element of the canonical Wnt pathway. The normal position of muscle cells depends on the correct distribution of BMP and SF/HGF throughout the limb bud mesenchyme. Removal or inhibition of the BMP signals within the limb margins leads to a shift in position resulting in the fusion of the dorsal and ventral premuscle masses towards the manipulated areas. In the absence of BMPs, mispositioning requires the presence of SF/HGF. Consequently, ectopic application of exogenous SF/HGF in the presence of BMP signals does not change muscle positioning. We conclude that correct positioning of the premuscle masses in the limb buds is controlled by the combined influence of SF/HGF signals--guiding cells mainly in the proximo-distal axis--and BMP signals that restrict the positioning to the dorsal and ventral central portions of the limb buds.

Supernumerary humeral heads of the biceps brachii muscle revisited.

Supernumerary humeral heads of the biceps brachii muscle were found in 27 (15.4%) of 175 cadavers. They were bilateral in five cadavers and unilateral in 22 (8 left, 14 right), giving a total of 32 examples in 350 arms (9.1%). Depending on their origin and location, the supernumerary heads were classified as superior, infero-medial, and infero-lateral humeral heads. Previous studies were reviewed using this classification. The infero-medial humeral head was observed in 31 of 350 (9%) arms and was therefore the most common variation. The superior humeral head was observed in five (1.5%). The infero-lateral humeral head was the least common variation, observed only in one (0.3%) of 350 arms. A biceps brachii with three heads was observed in 27 of 350 (7.7%) arms and with four heads in five (1.4%) arms.

Patterns of connections between the musculocutaneous and median nerves in the axilla and arm.

Variations in connections between the musculocutaneous and median nerves in the arm are not as uncommon as was once thought. This opinion led us to perform a study in 138 cadavers (66 male, 72 female). These variations were seen in 64 cadavers (46.4%), 9 bilaterally and 55 unilaterally (26 right and 29 left); in total, therefore, variations were observed in 73 out of 276 arms (26.4%), 42 male and 31 female. No statistically significant differences by gender and side were observed. We classify the variations in three main patterns: Pattern 1, fusion of both nerves (14 arms, 19.2%); Pattern 2, presence of one supplementary branch between both nerves (53 arms, 72.6%); and Pattern 3, two branches (5 arms, 6.8%). Pattern 2 was further subdivided into a sub-group 2a when a single root from the musculocutaneous nerve contributed to the connection (51 arms, 69.9%), and 2b when there were two roots from the musculocutaneous nerve (2 arms, 2.7%). A combination of Patterns 1 and 2a was observed in one case (1.4%). Further variations are described, published classification systems are reviewed and a meta-analysis of previous results is presented. An overall incidence of 33% of variant arms was observed. Of these variant arms, Pattern 1 represented 13.1%, Pattern 2 represented 75.4%, and Pattern 3, 8.5%, similar to our figures.

Coexistence of a pectoralis quartus muscle and an unusual axillary arch: case report and review.

A pectoralis quartus muscle and an unusual axillary arch were found on the left side of a female cadaver. The axillary arch was a musculoaponeurotic complex continuous with the iliacal fibers of the latissimus dorsi. The muscular part, together with the tendon of pectoralis major, inserted into the lateral lip of the bicipital groove of the humerus, whereas the aponeurotic part was formed by a fibrous band that extended deep to the pectoralis major to insert into the coracoid process between the attachments of the coracobrachialis and pectoralis minor. The pectoralis quartus originated from the rectus sheath, and joined the inferior medial border of the fibrous band of the axillary arch, at the lateral edge of the pectoralis major. The axillary arch muscle crossed anteriorly the axillary vessels and the brachial plexus. The clinical importance of these muscles is reviewed.

Extensor digitorum brevis manus: anatomical, radiological and clinical relevance. A review.

The extensor digitorum brevis manus (EDBM) is a supernumerary muscle in the dorsum of the hand frequently misdiagnosed as a dorsal wrist ganglion, exostosis, tendon sheath cyst or synovitis. Its presence in a living subject, confirmed by magnetic resonance imaging (MRI), is presented together with a review of the hitherto reported cases and the results of an anatomical study on 128 adult human cadavers (59 males and 69 females). The EDBM was found in three (2.3%) of the 128 cadavers. It occurred in two (3.4%) of the 59 male cadavers (one bilateral and one unilateral on the right side) and in one (1.5%) of the 69 female cadavers (unilateral on the left side). Consequently, the EDBM was found in four (1.6%) of the 256 upper limbs. It originated from the dorsal wrist capsule within the compartment deep to the extensor retinaculum for the extensor digitorum and inserted into the extensor hood of the index finger in one case and into that of the middle finger in three cases. In both hands of the living subject, the origin was similar but the insertion was into the index and middle fingers. In all cases, it was innervated by the posterior interosseous nerve and its blood supply was provided by the posterior interosseous artery.