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Tendons and ligaments are fibrous connective tissues vital to the transmission of force and stabilization of the musculoskeletal system. Arising in precise regions of the embryo, tendons and ligaments share many properties and little is known about the molecular differences that differentiate them. Recent studies have revealed heterogeneity and plasticity within tendon and ligament cells, raising questions regarding the developmental mechanisms regulating tendon and ligament identity. Here, we discuss recent findings that contribute to our understanding of the mechanisms that establish and maintain tendon progenitors and their differentiated progeny in the head, trunk and limb. We also review the extent to which these findings are specific to certain anatomical regions and model organisms, and indicate which findings similarly apply to ligaments. Finally, we address current research regarding the cellular lineages that contribute to tendon and ligament repair, and to what extent their regulation is conserved within tendon and ligament development., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2021. Published by The Company of Biologists Ltd.)

Gubernaculum testes is the most important parameter in testicular migration. At the end of migration, it is described as scrotal ligament, which has implications in testicular torsion. The present study aims to examine the structure of scrotal ligament and compare it with gubernaculum. Sixteen adult cadaveric testicular specimens and fourteen fetal testicular specimens of different age groups were examined after getting ethical clearance from the institute ethics committee and consent from the parents. Meticulous dissection was done. The length, site of proximal, and distal attachment of scrotal ligament and gubernaculum were noted and histologically evaluated. A separate scrotal ligament could not be delineated in any adult specimens. It merged with testicular coverings. Histological examination showed the presence of patchy areas of dense collagen fibres of variable density amidst loose areolar connective tissue. In contrast, fetal specimens showed the presence of a definitive gubernaculum testes and revealed the presence of mesenchymal tissue, collagen, elastic fibres, and myocytes which varied according to gestational age of fetuses. Structure of scrotal ligament and gubernaculum testes is highly variable. Description of scrotal ligament as a firm attachment from lower pole of testes to scrotum is controversial, questioning its role as protective factor in testicular torsion.

Background: The Weberian apparatus enhances hearing in otophysan fishes, including Zebrafish (Danio rerio). Several studies have examined aspects of morphological development of the Weberian apparatus and hearing ability in Zebrafish. A comprehensive developmental description including both hard and soft tissues is lacking. This information is critical for both interpretation of genetic developmental analyses and to better understand the role of morphogenesis and integration on changes in hearing ability., Results: Histological development of hard and soft tissues of the Weberian apparatus, including ossicles, ear, swim bladder, and ligaments are described from early larval stages (3.8 mm notochord length) through adult. Results show a strong relationship in developmental timing and maturation across all regions. All required auditory elements are present and morphologically integrated early, by 6.5 mm SL. Dynamic ossification patterns and changes in shape continue throughout the examined developmental period., Conclusions: This study provides the first comprehensive histological description of Weberian apparatus development in Zebrafish. Morphological integration was found early, before increases in hearing ability were detected in functional studies (>10 mm total length), suggesting morphological integration precedes functional integration. Further research is needed to examine the nature of the functional delay, and how maturation of the Weberian apparatus influences functionality., (© 2020 Wiley Periodicals, Inc.)

Purpose: The aim was to clarify the topographical anatomy of the common tendinous ring for the four rectus muscles in both adults and fetuses., Methods: We histologically examined the annular ligament for a common origin of the extraocular rectus muscles using 10 specimens from elderly individuals and 31 embryonic and fetal specimens., Results: At 6 to 8 weeks, each rectus carried an independent long tendon, individually originating from the sphenoid. Notably, we found additional origins from the optic or oculomotor nerve sheath. At 12 to 15 weeks, the lateral, inferior, and medial recti muscles were united to provide a C-shaped musculofibrous mass that was separated from the superior rectus originating from the edge of the optic canal opening. Morphologic features at 31 to 38 weeks were almost the same as those at 12 to 15 weeks, but the long and thick common tendon of the three recti reached the sphenoid body in the parasellar area. In adults, a ring-like arrangement of the rectus muscles ended at a site 8.1 to 12.0 mm anterior to the optic canal opening and independent of the superior rectus origin, the lateral, inferior, and medial recti formed a C-shaped muscle mass. The united origins of the three recti changed to a fibrous band extending along the superomedial wall of the orbital fissure., Conclusions: Consequently, none of the specimens we examined exhibited an annular tendon representing a common origin of the four recti, suggesting that the common tendinous ring includes only medial, lateral, and inferior rectus muscles with the superior rectus taking its origin independently.

Bone protrusions provide stable anchoring sites for ligaments and tendons and define the unique morphology of each long bone. Despite their importance, the mechanism by which superstructures are patterned is unknown. Here, we identify components of the genetic program that control the patterning of Sox9 + / Scx + superstructure progenitors in mouse and show that this program includes both global and regional regulatory modules. Using light-sheet fluorescence microscopy combined with genetic lineage labeling, we mapped the broad contribution of the Sox9 + / Scx + progenitors to the formation of bone superstructures. Then, by combining literature-based evidence, comparative transcriptomic analysis and genetic mouse models, we identified Gli3 as a global regulator of superstructure patterning, whereas Pbx1 , Pbx2 , Hoxa11 and Hoxd11 act as proximal and distal regulators, respectively. Moreover, by demonstrating a dose-dependent pattern regulation in Gli3 and Pbx1 compound mutations, we show that the global and regional regulatory modules work in a coordinated manner. Collectively, our results provide strong evidence for genetic regulation of superstructure patterning, which further supports the notion that long bone development is a modular process.This article has an associated 'The people behind the papers' interview., Competing Interests: Competing interestsThe authors declare no competing or financial interests., (© 2019. Published by The Company of Biologists Ltd.)

Tendons and ligaments are connective tissues that have been comparatively less studied than muscle and cartilage/bone, even though they are crucial for proper function of the musculoskeletal system. In tendon biology, considerable progress has been made in identifying tendon-specific genes (Scleraxis, Mohawk, and Tenomodulin) in the past decade. However, besides tendon function and the knowledge of a small number of important players in tendon biology, neither the ontogeny of the tenogenic lineage nor signaling cascades have been fully understood. This results in major drawbacks in treatment and repair options following tendon degeneration. In this review, we have systematically evaluated publications describing tendon-related genes, which were studied in depth and characterized by using knockout technologies and the subsequently generated transgenic mouse models (Tg) (knockout mice, KO). We report in a tabular manner, that from a total of 24 tendon-related genes, in 22 of the respective knockout mouse models, phenotypic changes were detected. Additionally, in some of the models it was described at which developmental stages these changes appeared and progressed. To summarize, only loss of Scleraxis and TGFβ signaling led to severe tendon developmental phenotypes, while mice deficient for various proteoglycans, Mohawk, EGR1 and 2, and Tenomodulin presented mild phenotypes. These data suggest that the tendon developmental system is well organized, orchestrated, and backed up; this is even more evident among the members of the proteoglycan family, where the compensatory effects are much clearer. In future, it will be of great importance to discover additional master tendon transcription factors and the genes that play crucial roles in tendon development. This would improve our understanding of the genetic makeup of tendons, and will increase the chances of generating tendon-specific drugs to advance overall treatment strategies.

The goal of this study is to clarify the development of the long head of the biceps brachii tendon (LHBT) and to verify the existence and development of the coracoglenoid ligament. Histological preparations of 22 human embryos (7-8 weeks of development) and 43 human fetuses (9-12 weeks of development) were studied bilaterally using a conventional optical microscope. The articular interzone gives rise to the LHBT, glenoid labrum, and articular capsule. During the fetal period, it was observed that in 50 cases (58%), the LHBT originated from both the glenoid labrum and the scapula, while in 36 cases (42%), it originated only from the glenoid labrum. The coracoglenoid ligament, first described by Sappey in 1867, is a constant structure that originates at the base of the coracoid process and projects toward the glenoid labrum zone, which is related to the origin of the LHBT. The coracoglenoid ligament was more easily identifiable in the 36 cases in which the LHBT originated only from the glenoid labrum. We suggest that the coracoglenoid ligament is a constant anatomical structure, is not derived from the articular interzone unlike the LHBT, and contributes to the fixation of the glenoid labrum in the scapula in cases in which the LHBT originated only from the glenoid labrum. We postulate that, when the LHBT is fixed only at the glenoid labrum, alterations in the coracoglenoid ligament could lead to a less sufficient attachment of the glenoid labrum to the scapula which could predispose to a superior labral lesion., (© 2017 S. Karger AG, Basel.)

Despite the common structure of vertebrates, the development of the vertebral column differs widely between teleosts and tetrapods in several respects, including the ossification of the centrum and the function of the notochord. In contrast to tetrapods, vertebral development in teleosts is not fully understood, particularly for large fish with highly ossified bones. We therefore examined the histology and gene expression profile of vertebral development in fugu, Takifugu rubripes, a model organism for genomic research. Ossification of the fugu centrum is carried out by outer osteoblasts expressing col1a1, col2a1, and sparc, and the growing centra completely divide the notochord into double cone-shaped segments that function as intercentral joints. In this process, the notochord basal cells produce a thick notochord sheath exhibiting Alcian-blue-reactive cartilaginous properties and composing the intercentral ligament in cooperation with the external ligament connective tissue. Synthesis of the matrix by the basal cells was ascertained by an in vitro test. Expression of twist2 indicates that this connective tissue is descended from the embryonic sclerotome. Notochord basal cells express sox9, ihhb, shh, and col2a1a, suggesting that the signaling system involved in chondrocyte proliferation and matrix production also functions in notochord cells for notochord sheath formation. We further found that the notochord expression of both ntla and shh is maintained in the fugu vertebral column, whereas it is turned off after embryogenesis in zebrafish. Thus, our results demonstrate that, in contrast to zebrafish, a dynamic morphogenesis and molecular network continues to function in fugu until the establishment of the adult vertebral column.

The segmental organization of the vertebrate body is most obviously visible in the vertebral column, which consists of a series of vertebral bones and interconnecting joints and ligaments. During embryogenesis, the vertebral column derives from the somites, which are the primary segments of the embryonic paraxial mesoderm. Anatomical, cellular and molecular aspects of vertebral column development have been of interest to developmental biologists for more than 150 years. This review briefly summarizes the present knowledge on early steps of vertebral column development in amniotes, starting from sclerotome formation and leading to the establishment of the anatomical bauplan of the spine composed of vertebral bodies, vertebral arches, intervertebral discs and ribs, and their specific axial identities along the body axis., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

The term 'inconspicuous penis' refers to a group of anatomical abnormalities in which the penis looks smaller than is expected. Micropenis can be defined as 'true micropenis'--which results from a defect in the hypothalamic-pituitary-gonadal axis--and 'micropenis secondary to congenital anatomical anomalies of the surrounding and overlying structures'--also known as 'concealed penis'. The different forms of concealed penis include webbed penis, congenital megaprepuce and partially hidden penis caused by prepubic adiposity. This disorder can also have iatrogenic causes resulting from adhesions that are secondary to circumcision--this type of concealed penis is known as 'trapped penis'. However, in both groups, micropenis is defined as a stretched penile length that is at least 2.5 SD below the mean for the patient's age, but without any other penile defects. Patients with true micropenis can be managed with testosterone, which has demonstrated good penile elongation results in the long term. Surgery also has a pivotal role in reconstruction for elongating the penis and for correction of anatomical abnormalities in concealed penis.

The gastric ligaments are superficial cord-like structures, located on the lesser curvature of the stomach, that extend from the pylorus to the esophagus. These ligaments have been documented in a wide variety of mammalian species, including humans, but their composition and ontogeny is unexplored. Here, we demonstrate that, during ontogeny, the gastric ligaments are first visible as extensions from a C-shaped domain of Gata3-expressing cells that surround the future pylorus; this domain will later give rise to the pyloric outer longitudinal muscle (OLM). The open ends of the C are located ventrally, and, beginning at embryonic day (E) 13.5, the ligaments grow anteriorly from these points. Whereas most other ligaments of the stomach are neurovascular in nature, the gastric ligaments are composed of smooth muscle cells that mature between E14.5 and E16.5. The gastric ligaments coexpress the transcription factors Gata3, Nkx2-5, and Sox9, and germline loss of Gata3 or conditional deletion of Nkx2-5 abrogates Sox9 expression and impairs gastric ligament smooth muscle development; similar phenotypes were previously seen in the OLM. In accord with this molecular contiguity between the OLM and gastric ligaments, three-dimensional image reconstruction highlights physical contiguity between these smooth muscle structures, suggesting that they may work together as a unit to control flexure of the pyloric region, a function similar to the ligament of Treitz at the duodenojejunal junction. These findings may have implications for our understanding of normal pyloric sphincter function, as well as the common human congenital pathology idiopathic hypertrophic pyloric stenosis., (Copyright © 2014 the American Physiological Society.)

Despite the importance of tendons and ligaments for transmitting movement and providing stability to the musculoskeletal system, their development is considerably less well understood than that of the tissues they serve to connect. Zebrafish have been widely used to address questions in muscle and skeletal development, yet few studies describe their tendon and ligament tissues. We have analyzed in zebrafish the expression of several genes known to be enriched in mammalian tendons and ligaments, including scleraxis (scx), collagen 1a2 (col1a2) and tenomodulin (tnmd), or in the tendon-like myosepta of the zebrafish (xirp2a). Co-expression studies with muscle and cartilage markers demonstrate the presence of scxa, col1a2 and tnmd at sites between the developing muscle and cartilage, and xirp2a at the myotendinous junctions. We determined that the zebrafish craniofacial tendon and ligament progenitors are neural crest derived, as in mammals. Cranial and fin tendon progenitors can be induced in the absence of differentiated muscle or cartilage, although neighboring muscle and cartilage are required for tendon cell maintenance and organization, respectively. By contrast, myoseptal scxa expression requires muscle for its initiation. Together, these data suggest a conserved role for muscle in tendon development. Based on the similarities in gene expression, morphology, collagen ultrastructural arrangement and developmental regulation with that of mammalian tendons, we conclude that the zebrafish tendon populations are homologous to their force-transmitting counterparts in higher vertebrates. Within this context, the zebrafish model can be used to provide new avenues for studying tendon biology in a vertebrate genetic system.

Through a histological examination of eight mid-term human fetuses (10-15 weeks) and seven late-stage fetuses (30-34 weeks), we attempted to determine how and when fetal ligaments around the tarsal bones form the regular arrangement seen in adults. Ligaments along the dorsal aspect of the tarsal bones developed early as an elongation of the perichondrium, in contrast to the late development of the plantar-sided ligaments. In contrast, a distal elongation of the tibialis posterior tendon was a limited plantar ligament in the early stage; finally, it extended from the navicular, ran obliquely to cross the dorsal side of the fibularis longus tendon, and inserted to the lateral cuneiform and fourth metatarsal. In the late stage, the adductor hallucis muscle origin provided multiple ligamentous structures along the cuneiforms and metatarsals. The tarsal sinus contained multiple fibrous bundles (possibly, the putative interosseous talocalcanean ligaments) that were derived from (1) insertion tendons of the extensor digitorus brevis muscle and (2) the fibrous sheath of the extensor digitorus longus tendon. The aponeurotic origin of the quadratus plantae muscle seemed to contribute to formation of the long plantar ligament. Therefore, tarsal ligaments appeared likely to develop from the long tendons, their fibrous sheaths and aponeuroses and intramuscular tendons of the proper foot muscles. Under in utero conditions with little or no stress from the plantar side of the foot, the muscle-associated connective tissue seems to play a crucial role in providing a regular arrangement of the ligaments in accordance with tensile stress from muscle contraction., (Copyright © 2013 Wiley Periodicals, Inc.)

In the human middle ear, the annular ligament of the incudostapedial joint and the insertions of the tensor tympani and stapedius muscles contain abundant elastic fibers; i.e., the elastic-fiber-mediated entheses. Hyaluronan also coexists with the elastic fibers. In the present study using immunohistochemistry, we demonstrated the distribution of elastin not only in the incudostapedial joint but also in the other two joints of the middle ear in adults and fetuses. In adults, the expression of elastin did not extend out of the annular ligament composed of mature elastic fibers but clearly overlapped with it. Electron microscopic observations of the annular ligament demonstrated a few microfibrils along the elastic fibers. Thus, in contrast to the vocal cord, the middle ear entheses seemed not to contain elaunin and oxytalan fibers. In mid-term fetuses (at approximately 15-16 weeks of gestation) before opening of the external acoustic meatus, the incudostapedial joint showed abundant elastic fibers, but the incudomalleolar and stapediovestibular joints did not. At this stage, hyaluronan was not colocalized, but distributed diffusely in loose mesenchymal tissues surrounding the ear ossicles. Therefore, fetal development of elastin and elastic fibers in the middle ear entheses is unlikely to require acoustic oscillation. In late-stage fetuses (25-30 weeks), whose ear ossicles were almost the same size as those in adults, we observed bundling and branching of elastic fibers. However, hyaluronan expression was not as strong as in adults. Colocalization between elastic fibers and hyaluronan appeared to be a result of postnatal maturation of the entheses., (Copyright © 2013 Elsevier GmbH. All rights reserved.)

Purpose: Here, we examined the development, composition, and structural organization of the ciliary zonule of the mouse. Fibrillin 1, a large glycoprotein enriched in force-bearing tissues, is a prominent constituent of the mouse zonule. In humans, mutations in the gene for fibrillin 1 (FBN1) underlie Marfan syndrome (MS), a disorder characterized by lens dislocation and other ocular symptoms., Methods: Fibrillin expression was analyzed by in situ hybridization. The organization of the zonule was visualized using antibodies to Fbn1, Fbn2, and microfibril-associated glycoprotein-1 (Magp1) in conjunction with 5-ethynyl-2'-deoxyuridine (EdU), an S-phase marker., Results: Microfibrils, enriched in Fbn2 and Magp1, were prominent components of the temporary vascular tunic of the embryonic lens. Fbn2 expression by nonpigmented ciliary epithelial cells diminished postnatally and there was a concomitant increase in Fbn1 expression, especially in cells located in valleys between the ciliary folds. Zonular fibers projected from the posterior pars plicata to the lens in anterior, equatorial, and posterior groupings. The attachment point of the posterior zonular fibers consisted of a dense meshwork of radially oriented microfibrils that we termed the fibrillar girdle. The fibrillar girdle was located directly above the transition zone, a region of the lens epithelium in which cells commit to terminal differentiation., Conclusions: The development and arrangement of the murine ciliary zonule are similar to those of humans, and consequently the mouse eye may be a useful model in which to study ocular complications of MS.

To investigate the factors affecting the development of the ligament of Treitz, we examined sagittal and frontal histological sections of 35 human fetuses with a crown-rump length of 100-300 mm (approximately 16-38 weeks of gestation). The retropancreatic fascia consistently extended in a layer behind the pancreatic body and the splenic artery and vein, and also in front of the left renal vein and left adrenal. In 18 specimens, a connective tissue band was seen originating from the diaphragmatic crus around the esophageal opening and ending at the retropancreatic fascia to the left of the origin of the celiac artery. In 10 of these 18 specimens, these putative upper parts of the ligament contained striated muscles, or so-called Hilfsmuskel. Although most of other 17 specimens were larger fetuses, the left adrenal, the liver caudate lobe and the celiac ganglion made space for the ligament very limited. In 22 specimens including the above 18, the retropancreatic fascia extended inferiorly to approach the fourth portion of the duodenum (D4) or the duodenojejunal junction (DJJ). However, in 11 of the 22 examples of the putative lower part of the ligament, the connection between the duodenal muscle coat and the fascia was interrupted by developing lymphatic tissues. Consequently, the ligament of Treitz seemed to develop from both pleuroperitoneal membrane-derived cells and the retropancreatic fusion fascia, although the morphology was markedly modified by adjacent structures such as the adrenal gland. The ligament may "recover" after the adrenal becomes reduced in size after birth.

Purpose: Cryptorchidism is the most common male congenital abnormality. The rodent gubernaculum steers the testis from abdomen to scrotum postnatally by eversion and migration through the developing inguinal fat pad (IFP). We hypothesize that extracellular matrix remodeling in/around the gubernaculum is necessary for eversion and migration and is permitted by timed IFP maturation and aimed to examine regional development and matrix metalloproteinase (MMP) content., Methods: Embryonic day 19 (E19) and postnatal days 0 and 2 (P0, P2) wild-type Sprague-Dawley rats (n = 10) were prepared for histologic examination (trichrome) and immunohistochemistry (membrane-type MMP-1 [MT1-MMP], MMP2) and analyzed using light/confocal microscopy., Results: At E19, IFP contained fibroblasts and immature cells in an extensive collagenous extracellular matrix. Cells in the gubernaculum base were cytoplasmic-MT1-MMP-positive (inactive). At P0, the gubernaculum had everted, and adjacent cells were membranous-MT1-MMP-positive (active). At P2, the gubernaculum was migrating through the IFP, and adjacent cells were membranous-MT1-MMP-positive. Adipocyte maturation began cranially in the IFP and proceeded in a craniocaudal gradient until more uniformly mature at P2., Conclusion: The MT1-MMP-positive cells may remodel the gubernaculum for eversion and provide the collagenolysis necessary for migration, like an icebreaking ship, through the IFP, which matures to permit migration through collagen-rich tissue. Disruption of these processes may cause cryptorchidism., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Background/purpose: Inguinoscrotal testicular descent is controlled by androgens between embryonic days E16-19, but androgen receptor (AR) and estrogen receptor (ER) locations are unknown. We aimed to find AR, ERα, and ERβ in the gubernaculum and inguinal fat pad (IFP) in normal rats and after flutamide treatment., Methods: Sprague-Dawley timed-mated rats were injected with flutamide (75 mg/kg body weight/5% ethanol + oil) on E16-19 or vehicle alone. Male fetuses or pups (5-10/group) were collected at E16; E19; and postnatal (P) days 0, 2, 4, 8. Sections were prepared for hematoxylin and eosin or immunohistochemistry for AR, ERα, and ERβ. Receptor labeling was quantitated as distinct nuclear labeling/100 μm(2) in gubernaculum and IFP., Results: There was minimal gubernacular AR-labeling until E19, dramatically increasing postnatally. By contrast, at E16-E19 there was significant IFP AR immunoreactivity suppressed by flutamide (P < .05). No ERα expression was observed, but ERβ was expressed in both gubernaculum and IFP, maximally at E16, but unchanged by flutamide., Conclusions: During the androgen sensitivity window (E16-19), the gubernaculum contains ERβ but minimal ERα or AR, while the IFP, which is supplied by the genitofemoral nerve, contains abundant AR that are flutamide-sensitive. These results suggest that the IFP could be the site of androgenic action controlling gubernacular development., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Background/aims: How androgens control inguinoscrotal descent remains controversial but may include canonical Wnt signaling via the transcriptional co-activator β-catenin. The canonical Wnt pathway transcribes genes regulating mesenchymal cell migration, fate, extracellular matrix remodeling, and in addition Axin2, a feedback product that reliably identifies Wnt activation. The relationship between β-catenin and androgen receptor warranted investigation into the involvement of the canonical Wnt pathway in testicular descent., Methods: Gubernacula from male Sprague-Dawley control (n = 22) and flutamide-treated (n = 18) rats at E17, E19, and D0 time-points were processed for immunohistochemistry. Sagittal sections stained for presence of androgen receptor, Axin2, and β-catenin were analyzed by fluorescent confocal microscopy., Results: At E19, β-catenin was strongly expressed in the membrane of developing cremaster muscle cells and the cytoplasm of gubernacular core cells. Axin2 expression was ubiquitous in nuclei of gubernacular mesenchymal cells, representing canonical Wnt signaling. After androgen blockade, Axin2 was conspicuously absent in the fibroblasts of the gubernacular core while remaining unaffected elsewhere. Reduced staining of Axin2 in E17 and D0 gubernacula suggests that Wnt signaling coincides with androgen programming., Conclusion: Axin2 expression in the E19 gubernaculum confirms canonical Wnt pathway activation. Its absence in the core of flutamide-treated gubernacula indicates Wnt down-regulation. As androgen is required for inguinoscrotal descent, downstream Wnt signaling may control initial gubernacular remodeling. Defects in this complex molecular process may play a role in cryptorchidism., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Purpose: There seems to be little or no information about the morphology of the fetal eye check ligament., Methods: The authors examined longitudinal and cross-histologic sections from the large collection of human fetuses at Universidad Complutense, Madrid., Results: In longitudinal sections from 20 fetuses (four at each of 12, 15, 20, 25, and 30 weeks of gestation), a distinct connective tissue band was found connecting the medial and lateral recti and the limbus of the conjunctiva (sites at and around the lateral and medial angles of the conjunctival space). Silver impregnation revealed that the muscle endomysium (type 4 collagen dominant) was connected with composite fibers of the band (type 1 collagen). The cross-sections from three fetuses (20 weeks) exhibited a site-dependent difference in the rectus sheaths: the orbital-sided sheath suddenly increased in thickness when it tightly attached to the muscle bundles. The attaching orbital-sided muscle bundles reached 14% to 15% (or 18%-20%) in the cross-sectional area of the MR (or the LR)., Conclusions: Taken together, the distinct connective tissue band extending to the conjunctiva was "originated from" the MR and LR rather than from a part of the muscles inserting into the connective tissue band. This band was most likely a primitive form of the check ligament. The authors hypothesize that the primitive check ligament conducts muscle tension to the conjunctiva to coordinate growth patterns between the anterior and posterior sides of the eyeball. This hypothesis may support en bloc recession for infantile esotropia.

Gubernacular elongation during inguinoscrotal testicular descent and cremaster muscle development remains poorly described in mammals. The role of the genitofemoral nerve (GFN) remains elusive. We performed detailed histological analysis of testicular descent in normal rats to provide a comprehensive anatomical description for molecular studies. Fetuses and neonatal male offspring (5-10 per group) from time-mated Sprague-Dawley dams (embryonic days 15, 16, and 19; postnatal days 0, 2, and 8) were prepared for histology. Immunohistochemistry was performed for nerves (Class III tubulin, Tuj1) and muscle (desmin). At embryonic days 15 and 16, the gubernaculum and breast bud are adjacent and both supplied by the GFN. By embryonic day 19, the breast bud has regressed and the gubernacular swelling reaction is completed. Postnatally, the gubernacular core regresses, except for a cranial proliferative zone. The cremaster is continuous with internal oblique and transversus abdominis. By postnatal day 2 (P2), the gubernaculum has everted, locating the proliferative zone caudally and the residual mesenchymal core externally. Eversion creates the processus vaginalis, with the everted gubernaculum loose in subcutaneous tissue but still remote from the scrotum. By P8, the gubernaculum has nearly reached the scrotum with fibrous connections attaching the gubernaculum to the scrotal skin. A direct link between GFN, gubernaculum, and breast bud suggests that the latter may be involved in gubernacular development. Second, the cremaster muscle is continuous with abdominal wall muscles, but most of its growth occurs in the distal gubernacular tip. Finally, gubernacular eversion at birth brings the cranial proliferative zone to the external distal tip, enabling gubernacular elongation similar to a limb bud., (Copyright © 2011 Wiley-Liss, Inc.)

Background/aims: Recent work both from our laboratory and in marsupial models of testicular descent suggests a strong connection between the mammary line and gubernacular migration. This study investigated the relationship between the mammary fat pad (MFP) that underlies the mammary line and the developing gubernaculum by fluorescent immunohistochemistry., Methods: Rats at E17 and E19 were fixed and processed for immunohistochemistry. Sagittal sections of male fetuses were stained with antibodies against androgen receptor (AR), prolyl-4 hydroxylase β, Desmin, activated Notch-1, Jagged-1, and Ki-67. These were analyzed by fluorescent confocal microscopy., Results: At E17 and E19, the MFP anlage forms a continuous distribution of fibroblasts passing immediately adjacent the gubernaculum to the future scrotum. Within this exists a distinct subpopulation of fibroblasts expressing AR distributed over the path of inguinoscrotal descent of the gubernaculum. Proliferation and Notch-1 signaling were similar throughout the MFP with differential Notch-1 signaling in the E19 gubernaculum., Conclusion: This investigation has identified the presence of a distinct AR-expressing subpopulation of MFP fibroblasts over the path of inguinoscrotal descent during the key androgenic programming window of this phase. This unique developmental pattern is consistent with a prime role for the MFP in testicular descent., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Background: Knowledge of biomechanics and the cervical spine's anatomy has become more topical as the incidence of whiplash neck disorders has increased. Unfortunately, injuries after traffic accidents are often brought to court, where the medical expert's knowledge is of utmost importance to ensure a correct medical evaluation., Material and Methods: The article is based on information identified through non-systematic searches of PubMed and on the author's experience as a professor of anatomy., Results: The cervical spine is particularly vulnerable to forces perpendicular to the length axis. Stability depends largely on the soft tissue. Injuries of soft tissue (especially in ligaments and intervertebral discs) may lead to instability and periosteal reaction with subsequent new formation of bone., Interpretation: The cervical spine is a relatively weak and vulnerable part of the body. One should consider locally restricted new formations of tissue with corresponding height reduction of the intervertebral disc as a sign of genuine injury.

Objective: To define the vascular and nervous relationships of the uterosacral ligament and to analyze histologically its content for a better description of this structure., Materials and Methods: Three fresh fetal cadavers, three embalmed and one fresh adult cadavers were used. The anatomical relationships of the uterosacral ligament were studied by dissecting one fresh fetal pelvis and two embalmed adult pelves. By histological and immunohistological examinations, eight biopsies of the cervical origin of the complexe ligamentaire utérosacral (USLC) were analyzed: four from fresh fetuses, two from a fresh adult cadaver and two from an embalmed adult cadaver. The specimens were stained with haematoxylin eosin safran (HES) coloration, with antinervous cell specific antibodies (PS100) and with antismooth muscle actine antibodies (to visualize vessel walls) before examination under optical microscope., Results: On anatomic examination, the uterosacral ligament was covered by the visceral pelvic fascia. By removing this fascia, the uterosacral ligament appeared to be a condensation of nervous fibers made up of hypogastric and pelvic nerves forming the hypogastric plexus. Histologically, the uterosacral ligament contained connective tissue, nervous fibers, sympathetic nodes, vessels and fatty tissue. No structured ligamentous organization was identified., Conclusion: The uterosacral "ligament" is in fact a "ligament complex" integrating connective tissue as well as nervous and vascular elements. Radical wide excisions of the USLC during cancer or endometriosis surgery and uterosacral suspension during pelvic floor reconstructive surgery should be performed with caution in order to preserve pelvic innervation.

Joint formation is a developmental process regulated by various factors including bone morphogenetic proteins, transforming and growth factors, etc. Recently, a high expression of cyclooxygenase (COX) isoforms in the foetal cartilaginous elements was also revealed. On the other hand, various joint and skeletal abnormalities were seen in laboratory animal and human offspring, exposed in utero to several COX inhibitors. Immunoexpression of constitutive (COX-1) and inducible (COX-2) cyclooxygenase isoforms was evaluated in various articular structures of untreated and unfamiliar 21-day-old male rat foetuses. Both COX isoforms were detected in the articular cartilage and joint capsule, as well as in the intra-articular disc of the temporomandibular joint and meniscus of the knee joint. COX-1 immunostaining was revealed in the anterior and posterior cruciate ligament of the knee joint and the labrum of the hip and shoulder, whereas COX-2 immunoreactivity in those structures was not found. It could be concluded that both constitutive and inducible COX isoforms are physiologically expressed in various structures of synovial joints in rat foetuses at the end of prenatal development.

Background: The control of the left hepatic vein (LHV) and the common trunk of the middle hepatic vein (MHV) and LHV (CT) is considered difficult during liver resection and could be improved by detailed knowledge on the ligamentum venosum Arantii (LV)., Aim: The aim of this study was to describe the LV and its connections to the LHV and the CT and to present surgical relevance of the obtained data., Material and Methods: During autopsy of 50 cadavers of both sexes, the LV was exposed, measured and then dissected, simulating a surgical maneuver to facilitate the approach to the LHV and CT. The extrahepatic parts of the LHV, MHV and CT were measured., Results: The LV was 52-70 mm long and 5-8 mm thick. It had a fibrotic structure and was not patent in 96% of the cases. The extrahepatic part of the LHV measured 3-19 mm, that of the MHV 3-18 mm and that of the CT 4-15 mm., Conclusion: LV dissection facilitated extraparenchymatous clamping of the hepatic veins: the extrahepatic parts of the LHV and CT measured > 3 mm in 86 and 84% of the cases, respectively., (2009 S. Karger AG, Basel.)

The aim of this study was to define the anatomical relationships of the uterosacral ligament complex (USLC) and to analyze histologically its content. Three fetal and four adult cadavers were used. Anatomical dissections were carried out. Eight fresh biopsies (four fetal and four adult) of the USLC were analyzed histologically and immunohistochemically. Specimens were stained with hematoxylin eosin safran coloration, with anti-nervous cell antibodies (PS 100) and with anti-smooth muscle antibodies (to visualize vessel walls). By removing the visceral pelvic fascia, nervous fibers were found within the USLC forming the hypogastric plexus. Histologically, the USLC contained connective tissue, nervous fibers, sympathetic nodes, vessels, and fatty tissue. No structured ligamentous organization was identified. The uterosacral "ligament" is a "complex" integrating connective tissue as well as nervous and vascular elements. Radical excisions and USLC suspension during pelvic floor reconstructive surgery should be performed with caution in order to preserve pelvic innervation.

Chondromodulin-I (ChM-I) and tenomodulin (TeM) are homologous angiogenesis inhibitors. We have analyzed the spatial relationships between capillary networks and the localization of these molecules during mouse and chick development. ChM-I and TeM proteins have been localized to the PECAM-1-negative avascular region: ChM-I is expressed in the avascular cartilage, whereas TeM is detectable in dense connective tissues, including tendons and ligaments. We have also examined the vasculature of chick embryos by injection with India ink and have performed in situ hybridization of the ChM-I and TeM genes. The onset of ChM-I expression is associated with chondrogenesis during mouse embryonic development. ChM-I expression is also detectable in precartilaginous or noncartilaginous avascular mesenchyme in chick embryos, including the somite, sclerotome, and heart. Hence, the expression domains of ChM-I and TeM during vertebrate development incorporate the typical avascular regions of the mesenchymal tissues.

Aim/background: How the gubernaculum guides the testis into the scrotum remains controversial, with various proposals from passive inversion to active growth. We aimed to determine if the gubernaculum contains an area of active proliferation, such as a "progress zone" in a growing embryonic limb bud, using a fluorescent cell membrane marker, 1,1'-didodecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate [DiIC12(3)], to trace cell migration, and 5-bromodeoxyuridine (BUDR) (a thymidine analogue) as a mitotic marker., Methods: Gubernacula were collected from neonatal male rats (n = 42, day 1-2, Sprague-Dawley) and cultured with calcitonin gene-related peptide (CGRP; 714 nmol/L). 1,1'-didodecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate-coated glass beads (diameter, 150-212 microm) were placed next to the bulb for the first 3 hours. Gubernacula were cultured for 3, 18, and 24 hours, then frozen sections cut and examined by confocal microscopy (wavelength, 549 nm). In a second experiment, pups not exposed to exogenous CGRP (n = 53, day 0, Sprague-Dawley) were injected intraperitoneally with BUDR (50 mg/kg of body weight); gubernacula were collected at 2, 48, 72, and 96 hours postinjection (PI), sectioned, and stained using immunohistochemistry to count the number of BUDR-positive cells per 100 cells (labeling index) in the bulb, cremaster, cord, and epididymis., Results: After 24 hours' culture with CGRP, the bulb showed an oval region (diameter, 300 microm) of high fluorescence, and the cremaster region showed elongated cells migrating out of the bulb. When cultured without CGRP, the same oval region contained no fluorescence. In vivo BUDR labeling index increased in all areas until 48 hours postinjection and then decreased most rapidly in the bulb (P < .05), in the presence of endogenous CGRP from the genitofemoral nerve., Conclusions: The rat gubernaculum contains a putative progress zone, such as in a growing limb bud, in the presence of CGRP. Cells migrate out of this zone to form cremaster muscle. We hypothesize that proliferation in the bulb elongates the gubernaculum, whereas proliferation of cremaster cells would increase gubernacular diameter. This brings to "life" the gubernaculum as an actively growing organ in contrast to the inert ligament connecting the testis to the scrotum portrayed in most anatomy textbooks.

Connective tissue mechanical behavior is primarily determined by the composition and organization of collagen. In ligaments and tendons, type I collagen is the principal structural element of the extracellular matrix, which acts to transmit force between bones or bone and muscle, respectively. Therefore, characterization of collagen fibril morphology and organization in fetal and skeletally mature animals is essential to understanding how tissues develop and obtain their mechanical attributes. In this study, tendons and ligaments from fetal rat, bovine, and feline, and mature rat were examined with scanning electron microscopy. At early fetal developmental stages, collagen fibrils show fibril overlap and interweaving, apparent fibril ends, and numerous bifurcating/fusing fibrils. Late in fetal development, collagen fibril ends are still present and fibril bundles (fibers) are clearly visible. Examination of collagen fibrils from skeletally mature tissues, reveals highly organized regions but still include fibril interweaving, and regions that are more randomly organized. Fibril bifurcations/fusions are still present in mature tissues but are less numerous than in fetal tissue. To address the continuity of fibrils in mature tissues, fibrils were examined in individual micrographs and consecutive overlaid micrographs. Extensive microscopic analysis of mature tendons and ligaments detected no fibril ends. These data strongly suggest that fibrils in mature ligament and tendon are either continuous or functionally continuous. Based upon this information and published data, we conclude that force within these tissues is directly transferred through collagen fibrils and not through an interfibrillar coupling, such as a proteoglycan bridge.

Tendons and ligaments (T/L) are very similar fibrous tissues that respectively connect muscle to bone and bone to bone. They are comprised of fibroblasts that produce large amounts of extra-cellular matrix, resulting in a dense and hypocellular structure. The complex molecular organization of T/L, together with high water content, are responsible for their viscoelastic properties, hence insuring their mechanical function. We will first review recent work on tendon embryology and discuss ligament formation, which has been less documented. We will next summarize our current knowledge of T/L molecular architecture, alterations of which are a major cause for disease. We will finally focus on T/L repair after injury and on genetic diseases responsible for T/L defects., (Copyright (c) 2005 Wiley-Liss, Inc.)

The avian lumbosacral vertebral column and spinal cord show a number of specializations which have recently been interpreted as a sense organ of equilibrium. This sense organ is thought to support balanced walking on the ground. Although most of the peculiar structures have been described previously, there was a need to reevaluate the specializations with regard to the possible function as a sense organ. Specializations were studied in detail in the adult pigeon. The development of the system was studied both in the pigeon (semiprecocial at hatching) and in the chicken (precocial). Specializations in the vertebral canal consist of a considerable enlargement, which is not due to an increase in the size of the spinal nervous tissue, but to a large glycogen body embedded in a dorsal rhomboid sinus. The dorsal wall of the vertebral canal shows segmented bilateral dorsal grooves, which are covered by the meninges towards the lumen of the vertebral canal leaving openings in the midline and laterally. This results in a system of lumbosacral canals which look and may function similar to the semicircular canals in the inner ear. Laterally these canals open above ventrolateral protrusions or accessory lobes of the spinal cord which contain neurons. There are large subarachnoidal cerebrospinal fluid spaces, lateral and ventral to the accessory lobes. Movement of this fluid is thought to stimulate the lobes mechanically. As to the development of avian lumbosacral specializations, main attention was given to the organization of the lobes and the adjacent fluid spaces including the dorsal canals. In the pigeon the system is far from being adult-like at hatching but maturates rapidly after hatching. In the chicken the system looks already adult-like at hatching. The implications derived from the structural findings are discussed with regard to a possible function of the lumbosacral specializations as a sense organ of equilibrium. The adult-like organization in the newly hatched chickens, which walk around immediately after hatching, supports the assumed function as a sense organ involved in the control of locomotion on the ground.

The teleost backbone consists of amphicoelous vertebrae and intervertebral ligaments, both of which include notochord-derived structures. On the basis of a sequential developmental study of the vertebral column of Atlantic salmon (Salmo salar L.) from the egg stage up to early fry stage (300-2500 day-degrees) we show that the vertebral body consists of four layers or compartments, two of which are formed through mineralization of preformed collagenous tissue (the notochordal sheath and the intervertebral ligament) and two of which are formed through ossification. The three inner layers have ordered lamellar collagen matrixes, which alternate perpendicularly from layer to layer, whereas the outer layer consists of cancellous bone with a woven matrix. The bone layers also differ in osteocyte content. In this study we describe the structural details of the layers, and their modes of formation. The results are compared with previous descriptions, and possible phylogenetic implications are discussed.

Background/purpose: We aimed to clarify the morphogenesis of an anomalous ligamentum venosum terminating in the trunk of the superior left hepatic vein, because the ligamentum venosum ordinarily terminates into the root of the left hepatic vein or directly into the inferior vena cava., Methods: We examined an anomalous ligamentum venosum found in the cadaveric liver of an 84-year-old Japanese woman., Results: The ligamentum venosum in this liver was not found in the usual course, the fissure for the ligamentum venosum. It lay on the posterior surface of the liver, connecting the left branch of the portal vein and the trunk of a small left hepatic vein. The small left hepatic vein draining the cranio-dorsal part of the lateral segment of the liver was revealed to be a superior left hepatic vein. This type of anomaly was found only in this 1 liver, among 125 cadaveric livers that were dissected., Conclusions: Taking previous reports into consideration, the morphogenesis of the anomalous ligamentum venosum in the present case may be explained as being due to the persistence of the right half of the subdiaphragmatic anastomosis, which receives the blood from the ductus venosus in the embryonal period.

The gubernaculum is a mesenchymal tissue that connects the gonads to the inguinal abdominal wall in the mammalian embryo. During gestation in the male, differential development of the gubernaculum and regression of the cranial suspensory ligament coordinate the first phase of testicular descent. As many as 1-3% newborn boys show impaired testicular descent, in part due to problems in gubernacular development. Little is known about the signaling molecules and cascades that are required for the development and differentiation of the gubernaculum. Protein tyrosine kinases comprise a large class of proteins that play important roles in proliferation, differentiation, and many aspects of cell-cell signaling in tissues. To date, no information on the existence of members of tyrosine kinase family in gubernaculum is available. We used a reverse transcription-polymerase chain reaction approach to identify 25 individual members of cytoplasmic and receptor tyrosine kinase classes in the developing male gubernaculum. The analysis of cellular distribution suggests that each tyrosine kinase examined may play a unique role in gubernacular development and differentiation., (Copyright 2004 Wiley-Liss, Inc.)

The gubernaculum plays an essential role in the complex mechanism of testicular descent and inguinal hernia closure. Understanding this complex developmental process is gradually allowing us insight into how to regulate normal descent and also treat maldescended testes.

Purpose: One of the most accepted theories for explaining testicular ectopia is the existence of multiple distal insertions of the gubernaculum testis. We studied the distal insertions of the gubernaculum in human fetuses and patients with cryptorchidism., Materials and Methods: We studied 326 gubernacula in 163 human fetuses at 10 to 35 weeks of gestation and 133 gubernacula in 101 patients with cryptorchidism (mean age 6.4 years). With the aid of a stereoscopic microscope (2.5x magnification), the existence of distal insertions of the gubernaculum was investigated. Fisher's exact test was used to compare the 2 study populations., Results: Of 326 fetal testes 224 (68.7%) were abdominal, 45 (13.8%) inguinal and 55 (16.9%) scrotal. In 1 fetus (0.6%) at 23 weeks of gestation both testes and both gubernacula were absent. Among 133 cryptorchid testes 17 (12.8%) were abdominal, 92 (69.2%) inguinal, 24 (18%) high scrotal and 3 (2.3%) evanescent. Of 324 fetal gubernacula present we found only 2 anomalous distal insertions in the pubopenile region (0.6%). All other distal insertions (99.4%) were at the usual location. Of 133 cryptorchid testes the 3 evanescent testes did not present visible gubernacula. Among 130 gubernacula analyzed all were inserted in the scrotal region., Conclusions: The existence of anomalous insertions at the distal portion of the gubernaculum is rare in human fetuses and patients with cryptorchidism.

Peritoneal attachments of the liver to the diaphragm include the falciform ligament, the left triangular ligament, and the coronary ligament with the right triangular ligament. It is often surgically convenient when the gastroesophageal junction is explored to refer to a "right" and a "left" coronary ligament, but this is anatomically unjustified. To reassess the issue there are no "right" and "left" coronary ligaments; there are only the left triangular ligament and the complex of coronary and right triangular ligament. The latter is the lateral unification of the layers of the coronary ligament. The liver is posteriorly connected to the diaphragm, and therefore the coronary ligament has superior and inferior layers rather than anterior and posterior layers. Embryologically these ligaments reflect the remnants of the developmental pathways of the liver in the septum transversum, and in the mature organism they are peritoneal connections. Besides the liver is suspended in the bare area mostly by fibrous attachments and by the hepatic veins. Consequently the term "ligament" for these structures has to be revised. Therefore we suggest the terms "left triangular peritoneal attachment" of the liver and "coronary peritoneal attachment" instead of "left triangular ligament" and "coronary ligament."

Members of the cysteine-rich protein (CRP) family are evolutionary conserved proteins that have been implicated in the processes of cell proliferation and differentiation via the cytoskeletal proteins. In this paper, we present the dynamic expression pattern of CPR2 transcripts during chick limb bud development. CRP2 transcripts are located in various tissues, including muscle, arteries, cartilage, ligaments and digit tendons and also in the apical ectodermal ridge and feather buds.

Deletion of the relaxin-like factor (RLF) gene in mice causes retention of testicles and infertility. The development of a synthetic RLF has made it possible to investigate the events that connect the genomic event and the basic biological responses that cause gonadal positioning. Anti-RLF antibodies were raised against synthetic RLF, allowing determination of RLF concentrations during the critical period, testing for RLF receptors on the gubernaculum and exploration of the temporal relationship between receptor display and migration of the testes in developing rats. In male rat pups, serum RLF concentrations were high at day 2 before parturition (2.4 ng ml(-1)) and decreased sharply just before parturition. Thereafter, males and females had the same low serum concentrations until RLF concen-trations began to increase in males only, starting at day 10 after parturition and continuing until adult RLF concentrations (0.6 ng ml(-1)) were reached on day 39 after parturition. The testicles are descending into the scrotum during this phase of increasing RLF concentrations and are descended fully by day 19-21 after parturition, before adult hormone concentrations are established. The high prenatal serum RLF concentration coincides with high expression of RLF receptors in the gubernaculum tissue. Competitive binding of RLF per mg of membrane protein prepared from rat gubernacula at various developmental stages showed no increase in receptor density as sexual maturity was reached. Gubernaculum cells in primary culture showed an increased uptake of 5-bromo-2'-deoxyuridine in the presence of RLF compared with controls. These studies demonstrate that the synthetic RLF is biologically active and indicate that the cryptorchid phenotype INSL3(-/-) is a direct consequence of defective gubernaculum growth, caused by the absence of RLF during early phases of development.

Chondromodulin-I (CHM1) was identified recently as an angiogenesis inhibitor in cartilage. It is highly expressed in the avascular zones of cartilage but is absent in the late hypertrophic region, which is invaded by blood vessels during enchondral ossification. Blast searches with the C-terminal part of CHM1 in available databases led to the identification of human and mouse cDNAs encoding a new protein, Tendin, that shares high homology with CHM1. Based on computer predictions, Tendin is a type II transmembrane protein containing a putative proteinase cleavage and two glycosylation sites. Northern assays with mouse RNAs demonstrated strong expression of a 1.5-kb tendin transcript in the diaphragm, skeletal muscle, and the eye and low levels of expression in all other tissues investigated. In 17.5-day-old mouse embryos, in situ hybridization revealed high levels of tendin transcript in tendons and ligaments. Additional signals were detected in brain and spinal cord, liver, lung, bowels, thymus, and eye. Cartilage, where CHM1 is found, revealed low levels of tendin m-RNA. In adult mice, tendin is expressed in neurons of all brain regions and the spinal cord. The tendin gene is localized in the human Xq22 region, to which several human diseases have been mapped., (Copyright 2001 Wiley-Liss, Inc.)

Purpose: Testicular descent is controlled by 2 morphological and hormonal steps. Transabdominal testicular descent is mediated by gubernacular swelling and regression of the cranial suspensory ligament. Müllerian inhibiting substance (MIS) has been proposed to stimulate the swelling but this remains controversial. Recently, a mouse mutant for Leydig insulin-like hormone (Insl3) was found to have undescended testis and deficient gubernaculum. We examine the testicular position of Insl3 mutant mice and the development of gubernacula., Materials and Methods: Mice with Insl3 homozygotes (-/-), heterozygotes (+/-) and wild-types (+/+) were examined at embryonic day 16.5 and birth. Macroscopic dissections and measurements of the testicular position, as well as microscopic analysis (hematoxylin and eosin, and Masson's trichrome) were performed., Results: Of the mice 11 Insl3 homozygote males had significantly impaired testicular descent at embryonic day 16.5 and birth (p <0.01), and the cord was thin and elongated, while 14 heterozygotes and 7 wild-types had normal testicular descent. Microscopically, the gubernaculum of Insl3 homozygotes was small with some muscle development but no central core of mesenchyme at embryonic day 16.5. On the other hand, heterozygotes and wild-types had normal gubernacular development with a swelling reaction., Conclusions: Insl3 mutants show feminized gubernaculum with deficient mesenchymal core. Insl3 appears to have some role in the gubernacular swelling reaction in mice.

Purpose: In this study, the development of human embryonic temporomandibular joint (TMJ) and masticatory muscles were investigated by using computed 3-dimensional reconstructions., Materials and Methods: Sixteen human embryos and fetuses, ranging from 6.5 to 107 mm crown-rump length, were examined., Results: At 10 weeks, a band of mesenchyme extending from the attachment of the lateral pterygoid muscle to the condylar process was observed to pass through the medial side of the condylar process to attach to the malleus. The temporal, masseter, and pterygoid muscles develop from the so called "temporal muscle" primordium, and the temporal muscle was in continuity with the masseter muscle until 14 weeks of fetal life., Conclusions: The study shows that the muscles of mastication arise from a single primordium. It also confirms the presence of a ligamentous attachment between the lateral pterygoid muscle and the malleus.

Many studies have been published on the development of the human elbow joint, but authors disagree on its morphogenetic timetable. Most discrepancies center on the cavitation of the elbow joint (including the humeroradial, humeroulnar, and superior radioulnar joints), and the organization of the tunnel of the ulnar nerve. We summarize our observations on the development of the elbow joint in 49 serially sectioned human embryonic (n = 28) and fetal (n = 21) upper limbs. During week 12, ossification begins in the epiphyses of the elements comprising the elbow joint. At the end of the embryonic period, the shallow groove between the posterior aspect of the medial epicondyle and the olecranon process, begins to be visible. The elbow joint cavity appears in O'Rahilly stage 21 (51 days) at the level of the humeroulnar and humeroradial interzones. Formation of the cavity begins at the medialmost portion of the humeroradial interzone and the lateralmost portion of the humeroulnar interzone. The annular ligament begins to develop in O'Rahilly stage 21 (51 days), and the superior radioulnar joint cavity appears between this ligament and the lateral aspect of the head of the radius during O'Rahilly stage 23 (56 days). We established the morphogenetic timetable of the human elbow joint., (Copyright 2000 Wiley-Liss, Inc.)

The major, central portion of Meckel's cartilage undergoes fibrous transformation and contributes to the sphenomandibular ligament, whereas its distal end undergoes endochondral ossification ultimately giving rise to inner-ear ossicles. This regional histodifferentiation of Meckel's cartilage is known to be associated with the spatially restricted expression of type X collagen. The objective of this study was to determine if this unique histodifferentiation is regulated by local environmental factors or by a preprogrammed genetic mechanism. Meckel's cartilage, and condylar cartilage used for comparison, were isolated from 17-day-old rat embryos and from newborn rats, respectively. The cartilage explants were maintained in vitro for 50 days with or without supplementation with 10% fetal bovine serum. When the explants were cultured under serum-free conditions, well-regulated cartilage development was observed. Expression of type X collagen, a differentiation marker for hypertrophic cartilage, was restricted to the distal end of Meckel's cartilage, whereas type II and IX collagens were found uniformly along the entire explant. Matrix calcification was examined histochemically using alizarin red S staining and found to be restricted to the distal end of Meckel's cartilage. Both Meckel's and condylar cartilage cultured with 10% fetal bovine serum developed unregulated dysmorphogenesis. These data suggest that, although Meckel's cartilage has an intrinsic potential to differentiate to its terminal stage, external regulatory factors can significantly influence its normal development at the molecular level.

Introduction: Three-dimensional reconstruction of histologic structures is still a real challenge. This is the reason why very few specimens are published, especially for the head., Material and Methods: The chondrocranium of an 18-mm human embryo was reconstructed from serial histologic sections. The three-dimensional reconstruction was based on a software used in geology and adapted to medical applications. This software was able to smooth the reconstructed model, i.e. to correct the distortions due to the histologic preparation., Results: The chondrocranium model is presented. To facilitate spatial orientation, we added the reconstruction of Merkel cartilage, of Reichert cartilage, of the vertebrae, and of the cerebrum. The different portions of the chondrocranium are described., Discussion: The three-dimensional result is described and advantageously compared to the rare wax models available. The morphological differences are detailed., Conclusion: This technique of three-dimensional reconstruction with its smoothing procedure is a very well adapted method for reconstruction of histologic structures. The results presented confirm the educational value of this tool, which is otherwise a powerful instrument of morphogenic study.

The cranial suspensory ligament is located on the border of the cranial (mesonephric) mesentery in adult female mammals, which runs between the cranial pole of the internal genitalia and the dorsal abdominal wall. Absence of the cranial suspensory ligament in male mammals depends upon exposure of its primordium to fetal testicular androgens and is a prerequisite for testis descent. Female rats were exposed to 5alpha-dihydrotestosterone propionate at different stages of genital development, and cranial suspensory ligament development was studied in neonatal and in adult animals. Androgens suppressed cranial suspensory ligament development when exposure started during the early stages of genital development, until day 19 postconception (pc). Androgen receptor expression was immunohistochemically detected in the cranial mesentery of both sexes from day 16 pc onwards. A decrease of androgen receptor expression in female fetuses from day 18 pc onwards coincided with the appearance of a differentiated cranial suspensory ligament, as evidenced by the expression of two cell differentiation markers: alpha-smooth muscle (alpha-SM) actin and desmin. alpha-SM actin was located on the outer border of the cranial mesentery of both sexes at day 17 pc, and expression increased only in female fetuses. On day 19 pc, desmin expression was also detectable in the a-SM actin-positive cells. Proliferation and apoptosis indices of cells in the cranial mesentery, as analysed by 5'-bromodeoxyuridine incorporation and by detection of DNA strand breaks (TUNEL method) respectively, did not show any difference between the sexes, neither on day 17 nor on day 18 pc. Since primordial cells of the cranial suspensory ligament highly express the androgen receptor during the period of gestation when androgens can suppress cranial suspensory development, altered morphogenesis of these cells may be a direct consequence of androgen action.