Morphofunctional and ultrastructural evaluation of hip joint development in human fetuses / Evaluación morfofuncional y ultraestructural del desarrollo de la articulación de la cadera en fetos humanos

SUMMARY: The normal sequential development of the hip joint (HJ) was considered for the evaluation of the morphological and ultrastructural aspects of the joint cartilage of the proximal femoral head epiphysis in human fetuses between 16 to 31 weeks of intra uterine life (IUL). Twenty human fetuses were fixed in 10 % formalin solution. Fetuses were divided into 4 groups (n=5): Group 1 (G1): 16-19 weeks IUL; Group 2 (G2): 20-23 weeks IUL; Group 3 (G3): 24-27 weeks IUL and Group 4 (G4): 28-31 weeks IUL. The right moieties of the HJ were subjected to light microscopy to determine the chondrocyte area, volume, and density and the extracellular matrix (ECM) density. The collagen component in ECM was qualitatively evaluated using Safranin-O and picrosirius techniques under polarized light. The left portions were analyzed using scanning electron microscopy (SEM). The advance of age revealed a gradual increase in chondrocyte area and volume and in ECM density, and a decrease in chondrocyte density. The apparent prevalence of type II collagen fibers in G1 and type III collagen fibers in G4, as well as a balance between type I and III collagen fibers in G2 and G3 suggest a process of cartilaginous evolution and repair. The pantographic organization of the collagen fiber meshes from the depth to the cartilage surface of the femoral head suggests that the arcade collagen network architecture starts at the fetal stage, regardless of the compressive forces applied. The morphological data may contribute not only to a better understanding of the maturation and cartilage organization in this area but also to serve as a theoretical basis for aspects related to diseases and joint malformations.

RESUMEN: El desarrollo secuencial normal de la articulación de la cadera (AC) se consideró para la evaluación de los aspectos morfológicos y ultraestructurales del cartílago articular de la epífisis proximal y de la cabeza femoral en fetos humanos entre 16 y 31 semanas de vida intrauterina (SVIU). Veinte fetos humanos fueron fijados en solución de formalina al 10 %. Los fetos se dividieron en 4 grupos (n = 5): Grupo 1 (G1), 1619 semanas de IUL; Grupo 2 (G2), 20-23 semanas SVIU; Grupo 3 (G3), 24-27 semanas SVIU y Grupo 4 (G4), 28-31 semanas SVIU. Las muestras derechas de la AC se sometieron a microscopía óptica para determinar el área, el volumen y la densidad de los condrocitos y la densidad de la matriz extracelular (MEC). El componente de colágeno en la MEC se evaluó cualitativamente utilizando técnicas de safranina-O y picrosirius bajo luz polarizada. Las muestras de la AC izquierda se analizaron utilizando microscopía electrónica de barrido (MEB). El avance de la edad reveló un aumento gradual en el área y el volumen de los condrocitos y en la densidad de la MEB, y una disminución en la densidad de los condrocitos. La aparente prevalencia de las fibras de colágeno tipo II en G1 y tipo III en G4, así como el equilibrio entre las fibras de colágeno tipo I y III en G2 y G3 sugieren un proceso de evolución y reparación cartilaginosa. La organización pantográfica de las mallas de fibra de colágeno desde la profundidad a la superficie del cartílago de la cabeza femoral sugiere que la arquitectura de la red de colágeno comienza en la etapa fetal, independientemente de las fuerzas compresivas aplicadas. Los datos morfológicos pueden contribuir no solo a una mejor comprensión de la organización de la maduración y el cartílago en esta área, sino también servir de base teórica para los aspectos relacionados con enfermedades y malformaciones articulares.

Topographical variations of the incisive canal and nasopalatine duct in human fetuses / 대한해부학회지

Development of the pulmonary pleura with special reference to the lung surface morphology: a study using human fetuses / 대한해부학회지

In and after the third trimester, the lung surface is likely to become smooth to facilitate respiratory movements. However, there are no detailed descriptions as to when and how the lung surface becomes regular. According to our observations of 33 fetuses at 9–16 weeks of gestation (crown-rump length [CRL], 39–125 mm), the lung surface, especially its lateral (costal) surface, was comparatively rough due to rapid branching and outward growing of bronchioli at the pseudoglandular phase of lung development. The pulmonary pleura was thin and, beneath the surface mesothelium, no or little mesenchymal tissue was detectable. Veins and lymphatic vessels reached the lung surface until 9 weeks and 16 weeks, respectively. In contrast, in 8 fetuses at 26–34 weeks of gestation (CRL, 210–290 mm), the lung surface was almost smooth because, instead of bronchioli, the developing alveoli faced the external surfaces of the lung. Moreover, the submesothelial tissue became thick due to large numbers of dilated veins connected to deep intersegmental veins. CD34-positive, multilayered fibrous tissue was also evident beneath the mesothelium in these stages. The submesothelial tissue was much thicker at the basal and mediastinal surfaces compared to apical and costal surfaces. Overall, rather than by a mechanical stress from the thoracic wall and diaphragm, a smooth lung surface seemed to be established largely by the thick submesothelial tissue including veins and lymphatic vessels until 26 weeks.

CD57 (Leu-7, HNK-1) immunoreactivity seen in thin arteries in the human fetal lung / 대한해부학회지

CD57 (synonyms: Leu-7, HNK-1) is a well-known marker of nerve elements including the conductive system of the heart, as well as natural killer cells. In lung specimens from 12 human fetuses at 10–34 weeks of gestation, we have found incidentally that segmental, subsegmental, and more peripheral arteries strongly expressed CD57. Capillaries near developing alveoli were often or sometimes positive. The CD57-positive tissue elements within intrapulmonary arteries seemed to be the endothelium, internal elastic lamina, and smooth muscle layer, which corresponded to tissue positive for a DAKO antibody reactive with smooth muscle actin we used. However, the lobar artery and pulmonary arterial trunk as well as bronchial arteries were negative. Likewise, arteries in and along any abdominal viscera, as well as the heart, thymus, and thyroid, did not express CD57. Thus, the lung-specific CD57 reactivity was not connected with either of an endodermal- or a branchial arch-origin. CD57 antigen is a sugar chain characterized by a sulfated glucuronic acid residue that is likely to exist in some glycosphingolipids. Therefore, a chemical affinity or an interaction might exist between CD57-positive arterioles and glycosphingolipids originating from alveoli, resulting in acceleration of capillary budding to make contact with the alveolar wall. CD57 might therefore be a functional marker of the developing air-blood interface that characterizes the fetal lung at the canalicular stage.

Pacinian corpuscle-like structure in the digital tendon sheath and nail bed: a study using late-stage human fetuses / 대한해부학회지

Pacinian corpuscle-like structures were identified in the digital tendon sheaths and nail beds of hands obtained from eight of 12 human fetuses of gestational age 20–34 weeks (crown-rump length, 150–290 mm). The aberrant corpuscles were present in tight fibrous tissue connecting the flexor tendon sheath to the dorsal aponeurosis (138 corpuscles in the thumbs and all fingers of eight fetuses); loose fibrous tissue inside the sheath on the dorsal side of the tendon (37 corpuscles in the thumbs and all fingers of four fetuses); and the nail bed (10 clusters in the thumbs and second fingers of four smaller fetuses). The aberrant corpuscles in the tendon sheath were classified into two types: thin and short, with tightly packed lamellae, of diameter 20–40 µm and length 20–200 µm; and thick and long, with loosely packed lamellae, of diameter 70–150 µm and length 0.5–1.5 mm. The small corpuscles tended to form clusters, each containing 5–10 structures. Their similarity indicated that the tight and loose lamellae in these two types of corpuscles corresponded to typical immature and mature corpuscles, respectively, usually distributed along the palmar digital nerve. However, mature, large corpuscles were absent from the nail bed, and most aberrant corpuscles were smaller than typical corpuscles along the nerve. The aberrant corpuscles were apparently incorporated into the tendon sheath or nail bed during fetal vascular development, but they appeared to degenerate after birth due to mechanical stress from the tendon or nail.

Screening and Analysis of Proteins Interacting with HCMV UL145 from a Human Fetal Brain cDNA Library / 中国医科大学学报

Objective To screen a human fetal brain cDNA library for proteins that can interact with HCMV UL145 using a yeast two-hybrid system.Methods A bait plasmid (pGBKT7-UL145) was constructed.Using HCMV UL145 as bait,a human fetal brain cDNA library was screened and proteins interacting with UL145 were identified using bioinformatic methods to sequence and analyze the positive clones.Results Three clones interacting with HCMV UL145 were found,and identified as FOXG1.Conclusion Several proteins interacting with HCMV UL145 in the human fetal brain cDNA library were identified as FOXG1,indicating that this protein may play an important role in the course of HCMV infection.

Screening and identification of proteins interacting with HCMV UL132 protein / 吉林大学学报(医学版)

Objective:To screen the proteins interacting with the human cytomegalovirus(HCMV)UL132 protein from the human fetus brain cDNA library by using Yeast Two-Hybrid System, and to elucidate the possible mechanism of UL132 protein in congenital cytomegalovirus infection.Methods:The HCMV UL132 fragment was amplified by polymerase chain reaction,the amplified HCMV UL132 fragment and expression vector pGBKT7 were digested and purified,and the HCMV UL132 fragment was linked to the vector pGBKT7.The pGBKT7-UL132 was constructed and transformed to yeast AH109, then the Human Fetal Brain DNA Library DNA was transformed into AH109 yeast.Using HCMV UL132 as abait, a human fetus brain cDNA was screened and the proteins interacting with UL132 protein were searched, the positive clone was sequenced and analyzed by bioinformatics methods.Results:The bait expression vector pGBKT7-UL132 was successfully constructed.The results of double enzyme digestion showed that there were two visible bands of 800 and 7 000 bp, respectively.After transformation of library plasmid, the transformation efficiency was calculated, and the transformation efficiency was 6.6×103 cfu· μg-1.There were 95 blue clones by X-gal coloration reactionsequencing and there were 10 clones interacting with the protein encoded by UL141 protein.The BLAST analysis showed that 7 of them were highly homologous with CAML.Conclusion:CAML might be one interaction protein with HCMV UL132 in Human Fetus Brain cDNA Library,suggesting that the interaction may be associated with the invasion and proliferation of the HCMV.

An anomalous portal vein crossing the lesser sac and ending at the upper part of ductus venosus / 대한해부학회지

In serial sagittal sections of a fetus on week 9 (crown-rump length, 36 mm), we incidentally found absence of the usual portal vein through the hepatoduodenal ligament. Instead, an anomalous portal vein originated behind the pancreatic body, crossed the lesser sac and merged with the upper part of the ductus venosus. During the course across the lesser sac, the vein provided a deep notch of the liver caudate lobe (Spiegel's lobe). The hepatoduodenal ligament contained the hepatic artery, the common bile duct and, at the right posterior margin of the ligament, and a branch of the anomalous portal vein which communicated with the usual right branch of the portal vein at the hepatic hilum. The umbilical portion of the portal vein took a usual morphology and received the umbilical vein and gave off the ductus venosus. Although it seemed not to be described yet, the present anomalous portal vein was likely to be a persistent left vitelline vein. The hepatoduodenal ligament was unlikely to include the left vitelline vein in contrast to the usual concept.

Distribution of CD10-positive epithelial and mesenchymal cells in human mid-term fetuses: a comparison with CD34 expression / 대한해부학회지

CD10, a marker of immature B lymphocytes, is expressed in the developing epithelium of mammary glands, hair follicles, and renal tubules of human fetuses. To assess mesenchymal and stromal expression of CD10, we performed immunohistochemical assays in whole body sections from eight fetuses of gestational ages 15-20 weeks. In addition to expression in urinary tract and intestinal epithelium, CD10 was strongly expressed at both gestational ages in fibrous tissues surrounding the airways from the larynx to lung alveoli, in the periosteum and ossification center, and in the glans of external genitalia. CD10 was not expressed, however, in other cavernous tissues. These findings suggest that mesenchymal, in addition to epithelial cells at specific sites, are likely to express CD10. The glomeruli, alveoli, and glans are all end products of budding or outgrowth processes in the epithelium or skin. However, in contrast to the CD34 marker of stromal stem cells, CD10 was not expressed in vascular progenitor cells and in differentiated vascular endothelium. The alternating pattern of CD10 and CD34 expression suggests that these factors play different roles in cellular differentiation and proliferation of the kidneys, airway and external genitalia.

Fetal development of the minor lung segment / 대한해부학회지

The mediobasal segment (S7) of the right lung has been considered to correspond to the cardiac lobe generally seen in mammals. To investigate fetal development of the right mediobasal segmental bronchus (B7), we examined paraffin-embedded serial sections of 15 embrynic and fetal lungs at 7-8 weeks (serial sections) as well as semiserial sections of 8 fetuses at 15-18 weeks (semiserial sections). All of the smaller specimens did not contain B7, but 2 of the 8 larger specimens carried B7: one was found in the immediately anterior side of the inferior pulmonary vein, while in the other, the subdivisions (B7a, B7b) were overriding the vein. Although the incidence might be underestimated because of observations using semiserial sections, the B7 was most likely to develop secondarily during a period from 8 to 15 weeks. Fetal topographical changes (mainly, the descent) of the middle lobe and the inferior pulmonary vein might relate with the secondarily budding of B7. The present result does not reduce a clinical relevance of B7 as a segmental bronchus of the lung segment system.

Distribution of elastic fibers in the head and neck: a histological study using late-stage human fetuses / 대한해부학회지

There is little or no information about the distribution of elastic fibers in the human fetal head. We examined this issue in 15 late-stage fetuses (crown-rump length, 220-320 mm) using aldehyde-fuchsin and elastica-Masson staining, and we used the arterial wall elastic laminae and external ear cartilages as positive staining controls. The posterior pharyngeal wall, as well as the ligaments connecting the laryngeal cartilages, contained abundant elastic fibers. In contrast with the sphenomandibular ligament and the temporomandibular joint disk, in which elastic fibers were partly present, the discomalleolar ligament and the fascial structures around the pterygoid muscles did not have any elastic fibers. In addition, the posterior marginal fascia of the prestyloid space did contain such fibers. Notably, in the middle ear, elastic fibers accumulated along the tendons of the tensor tympani and stapedius muscles and in the joint capsules of the ear ossicle articulations. Elastic fibers were not seen in any other muscle tendons or vertebral facet capsules in the head and neck. Despite being composed of smooth muscle, the orbitalis muscle did not contain any elastic fibers. The elastic fibers in the sphenomandibular ligament seemed to correspond to an intermediate step of development between Meckel's cartilage and the final ligament. Overall, there seemed to be a mini-version of elastic fiber distribution compared to that in adults and a different specific developmental pattern of connective tissues. The latter morphology might be a result of an adaptation to hypoxic conditions during development.

Spatial relationship between expression of cytokeratin-19 and that of connexin-43 in human fetal kidney / 대한해부학회지

Connexin-43, a major gap junction protein, and cytokeratin-19, one of the intermediate filament keratins, are known to be markers of well-differentiated epithelium. In this study, we investigated the expression of these markers in the head region, lungs, and abdominal organs of 10 human mid-term fetuses. The expression of connexin-43 was found to be restricted to the dura mater, kidney, and adrenal cortex. In the kidney, we found a clear site-dependent difference in the expression pattern of these markers: connexin-43 expression was observed in the tubules of the renal cortex whereas cytokeratin-19 was strongly expressed in the collecting ducts and renal pelvis. This difference remained unchanged throughout the fetal stages examined. Immunoreactivity was not observed for either of the markers in the intrarenal vessels, including the glomeruli, and mesangial cells. Connexin-43 expression seemed to be restricted to the metanephric vesicle-derived structures that differentiate in the urogenital ridge of the splanchnic mesoderm. The adrenal cortex also originates from the same para-aortic mesoderm. In contrast, in the urogenital organs, cytokeratin-19 seemed to be expressed in ducts derived from the urogenital sinus.

Influence of developing ligaments on the muscles in contact with them: a study of the annular ligament of the radius and the sacrospinous ligament in mid-term human fetuses / 대한해부학회지

The supinator muscle originates from the annular ligament of the radius, and the muscle fibers and ligament take a similar winding course. Likewise, the coccygeus muscle and the sacrospinous ligament are attached together, and show a similar fiber orientation. During dissection of adult cadavers for our educational curriculum, we had the impression that these ligaments grow in combination with degeneration of parts of the muscles. In histological sections of 25 human fetuses at 10-32 weeks of gestation, we found that the proximal parts of the supinator muscle were embedded in collagenous tissue when the developing annular ligament of the radius joined the thick intermuscular connecting band extending between the extensor carpi radialis and anconeus muscles at 18-22 weeks of gestation, and the anterior parts of the coccygeus muscle were surrounded by collagenous tissue when the intramuscular tendon became the sacrospinous ligament at 28-32 weeks. Parts of these two muscles each seemed to provide a mold for the ligament, and finally became involved with it. This may be the first report to indicate that a growing ligament has potential to injure parts of the "mother muscle," and that this process may be involved in the initial development of the ligament.

Fetal anatomy of the upper pharyngeal muscles with special reference to the nerve supply: is it an enteric plexus or simply an intramuscular nerve? / 대한해부학회지

We examined pharyngeal nerve courses in paraffin-embedded sagittal sections from 10 human fetuses, at 25-35 weeks of gestation, by using S100 protein immunohistochemical analysis. After diverging from the glossopharyngeal and vagus nerves at the level of the hyoid bone, the pharyngeal nerves entered the constrictor pharyngis medius muscle, then turned upward and ran superiorly and medially through the constrictor pharyngis superior muscle, to reach either the levator veli palatini muscle or the palatopharyngeus muscle. None of the nerves showed a tendency to run along the posterior surface of the pharyngeal muscles. Therefore, the pharyngeal nerve plexus in adults may become established by exposure of the fetal intramuscular nerves to the posterior aspect of the pharyngeal wall because of muscle degeneration and the subsequent rearrangement of the topographical relationship between the muscles that occurs after birth.

Expression of carbonic anhydrase IX in human fetal joints, ligaments and tendons: a potential marker of mechanical stress in fetal development? / 대한해부학회지

Carbonic anhydrase type IX (CA9) is known to express in the fetal joint cartilage to maintain pH against hypoxia. Using paraffin-embedded histology of 10 human fetuses at 10-16 weeks of gestation with an aid of immunohistochemistry of the intermediate filaments, matrix components (collagen types I and II, aggrecan, versican, fibronectin, tenascin, and hyaluronan) and CA9, we observed all joints and most of the entheses in the body. At any stages examined, CA9-poisitive cells were seen in the intervertebral disk and all joint cartilages including those of the facet joint of the vertebral column, but the accumulation area was reduced in the larger specimens. Glial fibrillary acidic protein (GFAP), one of the intermediate filaments, expressed in a part of the CA9-positive cartilages. Developing elastic cartilages were positive both of CA9 and GFAP. Notably, parts of the tendon or ligament facing to the joint, such as the joint surface of the annular ligament of the radius, were also positive for CA9. A distribution of each matrix components examined was not same as CA9. The bone-tendon and bone-ligament interface expressed CA9, but the duration at a site was limited to 3-4 weeks because the positive site was changed between stages. Thus, in the fetal entheses, CA9 expression displayed highly stage-dependent and site-dependent manners. CA9 in the fetal entheses seemed to play an additional role, but it was most likely to be useful as an excellent marker of mechanical stress at the start of enthesis development.

Reappraisal of intergender differences in the urethral striated sphincter explains why a completely circular arrangement is difficult in females: a histological study using human fetuses / 대한해부학회지

To investigate why the development of a completely circular striated sphincter is so rare, we examined histological sections of 11 female and 11 male mid-term human fetuses. In male fetuses, the striated muscle initially extended in the frontal, rather than in the horizontal plane. However, a knee-like portion was absent in the female fetal urethra because, on the inferior side of the vaginal end, a wide groove for the future vestibule opened inferiorly. Accordingly, it was difficult for the developing striated muscle to surround the groove, even though there was not a great difference in width or thickness between the female vestibule and the male urethra. The development of a completely circular striated sphincter seems to be impossible in females because of interruption of the frontal plane by the groove-like vestibule. However, we cannot rule out the possibility that before descent of the vagina, the urethral striated muscle extends posteriorly.

Reappraisal of intergender differences in the urethral striated sphincter explains why a completely circular arrangement is difficult in females: a histological study using human fetuses / 대한해부학회지

To investigate why the development of a completely circular striated sphincter is so rare, we examined histological sections of 11 female and 11 male mid-term human fetuses. In male fetuses, the striated muscle initially extended in the frontal, rather than in the horizontal plane. However, a knee-like portion was absent in the female fetal urethra because, on the inferior side of the vaginal end, a wide groove for the future vestibule opened inferiorly. Accordingly, it was difficult for the developing striated muscle to surround the groove, even though there was not a great difference in width or thickness between the female vestibule and the male urethra. The development of a completely circular striated sphincter seems to be impossible in females because of interruption of the frontal plane by the groove-like vestibule. However, we cannot rule out the possibility that before descent of the vagina, the urethral striated muscle extends posteriorly.

Dense distribution of macrophages in flexor aspects of the hand and foot of mid-term human fetuses / 대한해부학회지

In the developing human musculoskeletal system, cell death with macrophage accumulation occurs in the thigh muscle and interdigital area. To comprehensively clarify the distribution of macrophages, we immunohistochemically examined 16 pairs of upper and lower extremities without the hip joint (left and right sides) obtained from 8 human fetuses at approximately 10-15 weeks of gestation. Rather than in muscles, CD68-positive macrophages were densely distributed in loose connective tissues of the flexor aspects of the extremities, especially in the wrist, hand and foot. In contrast, no or fewer macrophages were evident in the shoulder and the extensor aspects of the extremities. The macrophages were not concentrated at the enthesis of the tendon and ligament, but tended to be arranged along other connective tissue fibers. Deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick end-labeling revealed apoptosis in the hand lumbricalis muscles, but not in the area of macrophage accumulation. Likewise, podoplanin-positive lymphatic vessels were not localized to areas of macrophage accumulation. Re-organization of the connective tissue along and around the flexor tendons of the hand and foot, such as development of the bursa or tendon sheath at 10-15 weeks, might require the phagocytotic function of macrophages, although details of the mechanism remain unknown.

Screening of protein interacting with the transcript of UL128 gene showed two protein patterns by yeast two-hybrid from human fetus brain cDNA library / 中华微生物学和免疫学杂志

Objective Using yeast two-hybrid system to screen the proteins which can interact with the human cytomegalovirus (HCMV) UL128 which have two difference transcription structure from human fetus brain cDNA library, and compare the difference with structure and function of interacting proteins. Methods Two fragments of UL128 were amplified by 3'RACE and 5'RACE technology, the length are 519 bp and 642 bp, respectively. The "bait plasmid" (named as pGBKT7-UL128-519 bp and pGBKT7-UL128-642 bp) was constructed successfully. Using pGBKT7-UL128-519 bp and pGBKT7-UL128-642 bp as a bait, a human fetus brain cDNA was screened and the proteins interacting with UL128-519 bp and UL128-642 bp encoded protein were searched, and the positive clones were sequenced and analyzed by bioinformatic methods. Results EFEMP2 interacting with HCMV UL128-519 bp were identified, THY-1 interacting with HCMV UL128-642 bp were identified. Conclusion EFEMP2 and THY-1 proteins interacting with HCMV UL128-519 bp and UL128-642 bp in human fetus brain cDNA library were successfully screened, but same proteins weren't found from the proteins interacting with UL128-519 bp and UL128-642 bp protein, UL128-519 bp and UL128-642 bp protein may be play an different effect in the process of infect by HCMV.

Screening of Protein Interacting with HCMV UL130 Protein by Yeast Two Hybrid from Human Fetus Brain cDNA Library / 中国医科大学学报

Objective To screen the human proteins interacting with human cytomegalovirus（HCMV）UL130 from human fetus brain cDNA library by GAL4 two-hybrid system 3 technique and analyze the corresponding coding sequences.Methods The ＂bait plasmid＂（named as pGBKT7-UL130）was constructed.By using HCMV UL130 as the bait,a human fetus brain cDNA library was screened and the proteins interacting with UL130 protein were searched.The positive clones were sequenced and analyzed by bioinformatic methods.Results Nine clones interacting with HCMV UL130 were identified,two of them were synaptosome-associated protein（SNAP）.Conclusion Some proteins interacting with HCMV UL130 in human fetus brain cDNA library were successfully screened.SNAP might play an important role in HCMV infection pathogenesis.