bluuebunny/crossref_metadata_2025 huggingface dataset:
Comparison of osteopenic changes in cancellous bone induced by ovariectomy and/or immobilization in adult rats
Authors: Bagi C. M. , Miller S. C.
Published: February 2005

Background: Ovariectomy (OVX) and immobilization (IMM) in rats are useful models of osteopenia, replicating some aspects of osteoporosis in humans. The purpose of this study was to compare changes in cancellous bone after OVX and/or IMM.

Methods: Differences in cancellous bone were determined at 6 and 12 weeks after OVX or IMM. Comparisons were also made when rats were ovariectomized or immobilized for 6 weeks and then immobilized (OVX/IMM) and ovariectomized (IMM/OVX), respectively, for 6 more weeks. The femurs were used to determine bone mineral content (BMC) using single photon absorptiometry (SPA) and for scanning electron microscopy (SEM). Tibias were collected for microradiography, image analysis, and histomorphometry of metaphyseal cancellous bone.

Results: Six and 12 weeks after OVX, there was less cancellous bone mass, compared with controls, as indicated by SPA, SEM, microradiography, image analyses, and histomorphometry. Bone was lost primarily from the central metaphyseal regions in the OVX animals, whereas the loss occurred throughout the metaphyses in the IMM animals. There were more rodlike bone spicules and fewer platelike trabecule in the OVX and IMM groups compared with controls. Differences in the structural aspects of the cancellous bone, including differences in the types of bone struts and marrow star volumes, indicated less trabecular connectivity and greater trabecular separation in the OVX and IMM animals, compared with controls. Endochondral growth indices in the IMM groups tended to be less, whereas the OVX groups tended to be greater than controls. Cancellous bone formation rates were generally greater in the OVX groups but less in the IMM groups compared with controls. Osteoclastic resorption surfaces were substantially elevated in the IMM and OVX groups, particularly the IMM groups. Changes reflecting OVX and IMM, independently, were apparent in the OVX/IMM and IMM/OVX groups and indices of osteopenia were different from controls, including less bone mass, trabecular connectivity, and greater trabecular separation, bone turnover rates, and osteoclastic surface.

Conclusions: These results demonstrate differences in the osteopenic changes that occur in cancellous bone following OVX or IMM. The changes were generally more dramatic in the IMM than in the OVX animals. When OVX and IMM were applied in combination, the osteopenic changes are particularly severe, emphasizing the importance of mechanical usage even with a deficiency of gonadal hormones. © 1994 Wiley‐Liss, Inc.
Stereological and serial section analysis of chondrocytic enlargement in the proximal tibial growth plate of the rat
Authors: Breur G. J. , Turgai J. , Vanenkevort B. A. , Farnum C. E. , Wilsman N. J.
Published: February 2005

Background: It has been suggested that within the growth plate, the final volume and shape of hypertrophic chondrocytes are important variables in determining the rate of longitudinal bone growth. To better understand the organization and regulation of chondrocytic hypertrophy as related to longitudinal bone growth, the beginning and end, and the location and magnitude of chondrocytic volume and shape changes during the hypertrophic process were defined in the proximal tibial growth plate of 35‐day‐old rats.

Methods: In this study we used two different approaches, a stereological analysis of chondrocytes in unbiasedly defined, narrow growth plate strata, and a serial section reconstruction and measurement of individual cells. In both experiments chondrocytes were preserved using optimal chemical fixation. Proliferating chondrocytes were identified using bromodeoxyruidine labelling, and the rat of longitudinal bone growth was determined using oxytetracycline labelling.

Results: In both studies, immediately following cell division in the proliferative zone, chondrocytic volume gradually increased toward the midpoint of the growth plate. During this phase of about 30 hours, approximately 20% of the final cell volume was obtained. During the following 20 hours the remaining 80% was acquired. The estimated rate of cell volume increased changed from approximately 50 μm3/hr during the first 30 hours to about 800 μm3/hr during the last 20 hours. The increase in cell volume resulted in an increase in both the vertical and the horizontal chondrocytic diameters. Cell parameters did not change during the final five hours of the maturation process.

Conclusions: In this study we demonstrated that chondrocytic enlargement starts immediately following cell division in the proliferative zone, and that chondrocytic enlargement consists of two morphologically distinguishable phases. The transition point between the first and the second phase of chondrocytic enlargement corresponded with the junction between the proliferative zone and the maturation zone. © 1994 Wiley‐Liss, Inc.
Isolation of granulosal‐like cells from the bovine secretory corpus luteum and their characterization in long‐term culture
Authors: Spanel‐Borowski Katharina , Ricken Albert M. , Kress Annetrudi , Huber Peter R.
Published: February 2005

Background: The isolation of cells termed type 5 from the bovine corpus luteum was recently reported. Since these cells were reminiscent of immature granulosa cells, their morphological and functional relationship requires further investigation in view of the novel concept of corpus luteum growth. It suggests that putative stem cells of unknown origin supply the pool of small luteal cells.

Methods: Bovine corpora lutea were mechanically dispersed, cell suspensions separated over a Percoll® density gradient, and type 5 cells purified by colony transfer. Granulosa cells were harvested from small‐sized antral follicles. Observations were carried out at the light and electron microscopical level. 3β‐Hydroxy‐steroid‐dehydrogenase was localized histochemically in addition to intracellular lipid droplets stained with nile red. Immunolocalization was used to study Factor VIII antigen presence, the architecture of the cytoskeleton, as well as the occurrence of neuronal cell adhesion molecules, and of neuronal cadherin‐like molecules. The uptake of acetylated low density lipoprotein was examined. As for progesterone concentration, cells were seeded at low density on day zero. Cell numbers and progesterone levels of supernatants were determined on day 10 in culture.

Results and Conclusions: Type 5 cells behaved morphologically like immature granulosa cells, yet the total cell number and the progesterone concentration differed for type 5 cells compared to granulosa cells. The addition of LH had no influence on the progesterone concentration as seen for either type 5 cells or for granulosa cells. It is concluded that type 5 cells, which were originally mistaken for microvascular endothelial cells, display similarities with immature granulosa cells. Type 5 cells may play a role in renewal of luteal cells. © 1994 Wiley‐Liss, Inc.
Morphological and histochemical study of human submucosal laryngeal glands
Authors: Pastor Luis M. , Ferran Antonio , Calvo Alfonso , Sprekelsen Carlos , Horn Ramon , Marin Juan A.
Published: February 2005

Background: The respiratory submucosal glands are a major source of secretions in the airway. Human submucosal laryngeal glands have been scarcely studied, with no works existing about their ultrastructure and histochemistry.

Methods: Samples of epiglottis, ventricle, false vocal folds and true vocal folds were fixed in 10% buffered formalin for histochemical study with conventional and carbohydrate lectin histochemistry. Other samples were fixed in 2.5% glutaraldehyde and conventionally processed for transmission electron microscopy.

Results: The human submucosal laryngeal glands are composed of serous tubules; mucous tubules; collector duct; and final portion of this duct. The serous cells showed sialosulphomucins and affinity for WGA and Con‐A lectins. With a previous treatment with neuraminidase, they also labelled with PNA. The mucous cells contained sialosulphomucins and showed affinity for WGA and DBA lectins in the samples proceeding from blood group A, and for WGA, UEA‐I and LTA with those from blood group O. Ultrastructurally, the serous cells presented a wide variety of granules, cells in which seromucous granules predominated. The mucous cells presented larger‐sized granules which were very electron‐lucent. The collector duct was composed of mitochondria‐rich cells and basal cells. A cell which we have termed “intermediate” was identified in the transition zone between the mucous tubules and the collector duct, and in the final portion of the collector duct. It had morphological characteristics as if it were a transition between a goblet cell and collector duct cell. Some nerve endings with cholinergic and peptidergic vesicles were found among the myoepithelial cells.

Conclusions: These glands presented some histological differences from the bronchial glands, the mucous secretion was related to the blood group antigens, and the serous cells showed a wide variability in their secretory granules, many of them being of a seromucous type. © 1994 Wiley‐Liss, Inc.
Liver of juvenile atlantic salmon, Salmo salar L.: A light, transmission, and scanning electron microscopic study, with special reference to the sinusoid
Authors: Speilberg Lars , Evensen Øystein , Nafstad Per
Published: February 2005

Background: This report provides a detailed description of sinusoidal and perisinusoidal structures in the normal liver of the juvenile Atlantic salmon (Salmo salar L.), a teleost species.

Methods: The liver was studied by light, transmission, and scanning electron microscopy, and organ specimens were sampled after retrograde, whole‐body perfusion through the dorsal aorta using 3% glutaraldehyde. Detailed characterization of perisinusoidal stellate cells also included immunohistochemical staining for desmin and evaluation of autofluorescence of the same cells upon excitation in ultraviolet (UV) light.

Results: The sinusoid is lined by one cell type only: the endothelial cell. No intraluminal pit cells or Kupffer cells are present. The space of Disse contains reticulin fibres, visualized by Gomori's silver stain, and perisinusoidal stellate cells (PSC). PSC exhibited autofluorescence in UV light, indicating that these cells store vitamin A in cytoplasmic lipid droplets. Immunohistochemically, PSC were found negative for desmin. The space of Disse, extending deep down between adjacent hepatocytes, receives long, slender microvilli from parenchymal cells. In addition to scattered macrophages, interhepatocytic cells (IHC) are found perisinusoidally. Hepatocytes of Atlantic salmon form branching and anastomosing tubules.

Conclusions: The sinusoids of Atlantic salmon liver are lined by a fenestrated endothelium, with PSC located in the space of Disse, with macrophages and IHC as inhabitants of the interhepatocytic space. IHC show ultrastructural similarities to mammalian pit cells and teleostean large granular lymphocytes, as well as to piscine monocytes. PSC might be storage cells for vitamin A in Atlantic salmon as shown by autofluorescence in these cells, while immunohistochemical studies indicate that desmin does not seem to be an adequate immunohistochemical marker for PSC in the juvenile Atlantic salmon. Methodologically, fixation for electron microscopy was performed by a new and convenient perfusion method: arterial retrograde whole body perfusion. Liver specimens intended for scanning electron microscopy were fractured at room temperature after prolonged osmium postfixation, leaving hepatocytes intact and producing images well suited to document the three‐dimensional structure of cells and tissue. © 1994 Wiley‐Liss, Inc.
Perspectives: A vital biomechanical model of the endochondral ossification mechanism
Authors: Frost Harold M. , Jee Webster S. S.
Published: February 2005

Background: Mechanical usage effects could explain many features of endochondral ossification and related processes. Mineralization of growth plate cartilage could reduce its mechanical strains enough to make its resorption begin and to guide it in space. By removing most of its mineralized vertical septae, resorption could overload the remainder enough to increase woven bone formation on them and construct the primary spongiosa. After it finishes mineralizing, the primary spongiosa could become stiff enough to begin partial disuse in strain terms, so BMU‐based remodeling would begin replacing it with lamellar bone. This would construct the secondary spongiosa. In transferring loads from the growth plate to the cortex, the central metaphyseal spongiosa becomes deloaded. This disuse would make remodeling remove it in the diaphyseal marrow space.

Methods: The slow growth of epiphyses and apophyses gives their spongiosas more time to adapt to their loads than the metaphyseal spongiosa beneath faster growing growth plates. Compared to metaphyseal trabeculae, this leads to fewer and thicker epiphyseal trabeculae that turn over more slowly and should persist for life because they carry loads for life.

Results: Rapid turnover of metaphyseal cortex in very young subjects could let it strain enough to form woven bone. Increased thickness and slower turnover of this cortex in older subjects could reduce its strains enough to make lamellar bone form there instead. This would compose this cortex mostly of woven bone in the very young and of lamellar bone in adults.

Conclusions: This model assigns particular importance to the stiffness and strains of tissues (as distinguished from their strength and stresses), to the relative rates of some processes, and to responses of the skeleton's biologic mechanisms to a tissue's typical largest mechanical strains (as distinguished from their stresses). © 1994 Wiley‐Liss, Inc.
Perspectives: Applications of a biomechanical model of the endochondral ossification mechanism
Authors: Frost H. M. , Jee W. S. S.
Published: February 2005

A biomechanical model of endochondral ossification (Frost and Jee, 1994. Anat. Rec., 240:435–446) can help to explain: (1) some differences in fracture patterns in children and adults, (2) increased fractures during the human adolescent growth spurt, (3) localization of stress fractures and pseudofractures to cortical instead of trabecular bone, (4) increased bone mass in adult‐acquired and childhood obesity, (5) subchondral bone densification and osteopenia in some arthroses, (6) why and where mammals lose spongiosa with aging, (7) why, as percents of the original bone stock, metaphyseal trabecular bone losses with aging usually exceed cortical bone losses, (8) why osteochondritis dissecans and aseptic necroses of bone localize in epiphyses instead of metaphyses, (9) some features of growth plate histology in rickets and the chondrodystrophies, (10) why spontaneous fractures in osteoporotic patients affect vertebral more than metaphyseal spongiosa, (11) why osteopenias develop in most chronic, debilitating diseases, and (12) why histomorphometric values can differ in iliac bone biopsies obtained by the “vertical” Jamshidi and “horizontal” Bordier‐Meunier techniques. © 1994 Wiley‐Liss, Inc.
Connections between the various elements of the Cis‐ and mid‐compartments of the Golgi apparatus of early rat spermatids
Authors: Clermont Y. , Rambourg A. , Hermo L.
Published: February 2005

Background: The exact structural relationships of the saccules, membranous tubules, and vesicles that compose the cis‐ and midcompartments of the Golgi cortex of rat spermatids was investigated to determine the relationship of these elements to each other.

Methods: Tissues fixed with glutaraldehyde and buffered in sodium cacodylate were examined with the electron microscope. Electron micrographs, including stereopairs, were analyzed to determine the three‐dimensional organization of the Golgi elements.

Results: The deeper layer of the Golgi cortex was composed of stacks of saccules connected to each other either by saccules or membranous tubules. The peripheral region of the Golgi cortex, located between the cisside of the stacks and a network of overlying ER cisternae contained numerous membranous tubules and vesicles of two class sizes: 50–100 nm vesicles and microvesicles 5–10 nm in diameter. The tubules formed tight networks, known as cis‐elements or cis‐Golgi networks (CGN), which were strictly parallel and next to the first or cis‐saccule of the stack. The cis‐elements were continuous with more loosely arranged peripheral tubules which formed elaborate, intertwined and interconnected networks. These peripheral tubules closely approximated the overlying ER cisternae in areas often showing fuzz‐coated finger‐like projections. Occasionally such peripheral tubules were continuous with ER cisternae. The saccules forming the stacks were continuous with membranous tubules which not only connected saccules of adjacent stacks, but also saccules of the same stack. These tubules were also connected with the tight tubular networks forming the cis‐elements and the broad networks formed by the peripheral membranous tubules. Vesicles (50–100 nm) and microvesicles (5–10 nm) frequently formed aggregates in the peripheral Golgi region next to areas of ER membrane free of fuzz‐coated projections. The microvesicles, embedded in a denser cytoplasmic matrix, had a more or less distinct delimiting membrane suggestive of their disintegration in this juxta‐ER location. The 50–100 nm vesicles that were seen at the periphery of the vesicular aggregates appeared to form mainly from the membranous tubules of the Golgi cortex.

Conclusions: Thus the saccules and membranous tubules of the spermatid's Golgi cortex formed a single continuous membranous system connected to ER cisternae. The vesicles, seemingly arising from the membranous tubules, appear to follow a retrograde pathway and undergo dissolution next to ER cisternae. © 1994 Wiley‐Liss, Inc.
Type and regional diversity in the distribution of myosin heavy chains in chicken intrafusal muscle fibers
Authors: Maier Alfred
Published: February 2005

Background: Chicken intrafusal fibers were classifed on the basis of their myosin heavy chain (MHC) composition, which was compared to that of mammalian nuclear bag and nuclear chain types.

Methods: Immunoreactivities of intrafusal fibers from leg muscles of 8‐week‐old chickens were evaluated in serial cross‐sections after incubation with monoclonal antibodies against slow‐twitch, slow‐tonic, or fast‐twitch MHC and fast muscle C‐protein.

Results: Four categories of slow intrafusal fiber could be distinguished on the basis of differential expression of slow‐twitch and slow‐tonic MHC. Segregation into types was most evident at the motor axon supplied pole, followed by the sensory region of the equator. Fiber types were least distinct at the juxtaequator where sensory and motor axons meet. Intrafusal fibers negative for slow myosins reacted with anti‐fast myosins. Fast fibers were best viewed as a single group without subdivisions. Immunostaining for fast muscle C‐protein paralleled in large part reactivities for neonatal/fast MHC, indicating that proteins other than MHC are useful fiber type markers.

Conclusions: Despite regional changes along the length of intrafusal fibers and some variation within fiber types, the concept of separate MHC‐based fiber types was valid as long as typing of fibers was restricted to the proximal polar region. Comparisons of MHC profiles revealed similarities between chicken fast intrafusal fibers and mammalian nuclear chain fibers and between some chicken slow intrafusal fibers and mammalian nuclear bag fibers. © 1994 Wiley‐Liss, Inc.
Immunocytochemical detection of actin and 53 kDa polypeptide in the epididymal spermatozoa of rat and mouse
Authors: Paranko Jorma , Yagi Ahmed , Kuusisto Mari
Published: February 2005

Background: Presence of immunocytochemically detectable actin in the rat and mouse sperm head has been enigmatic for years. In this study, we demonstrate actin in the perinuclear theca and show that the detection of actin epitopes in the rat and mouse epididymal spermatozoa can effectively be enhanced by pre‐extraction of sperm cells with SDS.

Methods: The study with one monoclonal and one polyclonal anti‐actin antibody was carried out at conventional and confocal fluorescence and electron microscope level, and by immunoblotting of proteins isolated from the head and tail fractions.

Results: In the head of the control methanol‐acetone fixed rat spermatozoa, the polyclonal antibody gave a stronger immunostaining in the postacrosomal area and in the perforatorium than the monoclonal antibody. In the mouse sperm head, the monoclonal antibody labeled the ventral edge of the postacrosomal area and slightly the perforatorium, whereas the polyclonal antibody stained the entire perinuclear space. In the SDS‐extracted spermatozoa, an intense postacrosomal and perforatorial labeling was obtained with both antibodies but, in particular in the rat spermatozoa, the middle lateral portion of the postacrosomal segment remained unlabeled. Sonication seemed to cause structural modifications which specifically impeded staining with the monoclonal antibody. Both antibodies detected actin in the basal plate and the monoclonal antibody in the neck. Amorphous matrix of the connecting piece showed immunogold labeling. In the tail, the monoclonal antibody recognized actin and a relatively basic 53 kDa polypeptide, whereas the polyclonal antibody reacted with several protein bands. SDS‐soluble actin of the tail was addressed to the midpiece and the SDS‐insoluble 53 kDa protein profoundly to the outer dense fibers of the principal piece.

Conclusions: Intense labeling of actin in the SDS‐extracted rat and mouse spermatozoa was presumably due to the generated demasking of actin epitopes embedded in the perinuclear cytoplasm. The results are important in confirming that actin in the rat and mouse sperm head is not lost during spermiogenesis but apparently contributes to the three‐dimensional packing of the mature perinuclear cytoplasm. This study further demonstrates the importance of the methods used in sample preparation and advantages of conofocal microscopy when attempting to detect cytoskeletal proteins which, as in spermatozoa, may occur in small quantities. © 1994 Wiley‐Liss, Inc.
Maturation of undifferentiated lung epithelial cells into type II cells in vitro: A temporal process that parallels cell differentiation in vivo
Authors: Chinoy Mala R. , Antonio‐Santiago Maria T. , Scarpelli Emile M.
Published: February 2005

Background: Formation of alveolar‐like structures (ALS) by mature fetal rabbit type II pneumocytes (day 29 gestation) and long‐term differentiation on Engelbreth‐Holms‐Swarm mouse tumor extract or EHS gel (Matrigel) were reported by our group (Blau et al., 1988. J. Cell Physiol., 136:203–214). We now describe structural organization and differentiation of immature lung epithelial cells, isolated at day 22 gestation, into mature type II cells in vitro.

Methods: Peripheral pulmonary tissue was pooled and undifferentiated epithelial cells isolated for primary culture on Matrigel. Cells were examined 12–16 h after plating and on days 1,3,5, and 7 of culture and assessed by phase contrast and by transmission electron microscopy after fixation in situ.

Results: Cells formed ALS 12–16 h after plating. Spherule diameter increased about four to eight times from day 1–7 in culture. There was rapid transformation of tall columnar cells to cuboidal, normal polarization of cells with respect to cell‐free lumen of ALS, progressive reduction of glycogen zones, apparent gradual increase of cell organelles such as Golgi apparatus, rough endoplasmic reticulum and mitochondria, and apparent extrusion of lipidic figures into the lumen. These morphologic transformations in vitro temporally paralleled cell differentiation in vivo. The relative increase of 14C‐acetate precursor into phosphatidylcholine in contrast to cardiolipin was consistent with these transformations.

Conclusions: Under the conditions of our culture system, maturation of undifferentiated pulmonary epithelial cells is reproduced in vitro along the same time course and according to the same developmental sequence of fetal lungs in vivo. © 1994 Wiley‐Liss, Inc.
Formation of alternating tiers in the optic chiasm of the chick embryo
Authors: Drenhaus Ulrich , Rager Günter
Published: February 2005

Background: When the fibers of the two optic nerves of the chick cross to the contralateral side at the prospective chiasmatic region, they segregate into clearly defined bundles. These bundles form horizontally oriented tiers which alternate between the right and the left optic nerve.

Methods: We have analyzed the development of these tiers qualitatively and quantitatively using light and electron microscopy between embryonic days (E) 4 and E19.

Results: The formation of the chiasm begins on E4. In the course of E4, tiers become visible for the first time. Their number increases rapidly until E7. Then the increase is slowed down and the final value (32 ± 1) is approximated by E18/19. Growing axons allow one to distinguish three different segments: the growth cone, the distal, and the proximal segment. The latter originates in the perikaryon. Growth cones and distal segments are found predominantly in the ventralmost tiers. Their frequency decreases from ventral to dorsal. Proximal segments which indicate the presence of older axons appear first in the dorsal tiers and later also in more ventrally located tiers.

Conclusion: Based on these criteria it is concluded that newly formed axons contribute primarily but not exclusively to the ventral tiers. There is a gradient of maturity of axons from ventral to dorsal whose slope becomes steeper with age until the last growth cones have arrived by E18. Thus, the formation of the chiasm corresponds to the spatiotemporal pattern of ganglion cell formation in the retina. The process of cell death of retinal ganglion cells is also seen in the chiasm but probably does not lead to a transitory diminution in the number of tiers. © 1994 Wiley‐Liss, Inc.
Immunohistochemical localization of the high‐affinity NGF receptor (gp 140‐trkA) in the adult human dorsal root and sympathetic ganglia and in the nerves and sensory corpuscles supplying digital skin
Authors: Vega J. A. , Vazquez E. , Naves F. J. , Del Valle M. E. , Calzada B. , Represa J. J.
Published: February 2005

Background: Nerve growth factor (NGF) is produced in target issues of sympathetic and neural‐crest derived sensory neurons, including skin, to provide them trophic support. The biological effects of NGF on responsive cells are mediated by specific high‐affinity receptors. Recently, a protein tyrosine kinase of ≃ 140 kDa molecular weight, encoded by the proto‐oncogene trkA, has been identified as the high‐affinity NGF receptor (gp140‐trkA). The present work was undertaken to study the localization of gp140‐trkA‐like immunoreactivity (IR) in human peripheral ganglia (sympathetic and dorsal root ganglia), and in glabrous skin.

Methods: Lumbar dorsal root ganglia, para‐ and prevertebral sympathetic ganglia, and digital glabrous skin were studied immunohistochemically using a rabbit anti‐gp140‐trkA polyclonal antibody. In order to accurately establish the localization of gp140‐trkA IR, the neurofilament proteins and S‐100 protein were studied in parallel in: (1) sensory and sympathetic ganglia, to label neuron cell bodies and satellite or supporting cells, respectively; (2) human skin, to label axons, Schwann and related cells within nerves and sensory corpuscles. Moreover, a quantitative study (neuron size, intensity of immunostaining) was carried out on sympathetic and dorsal root ganglia neuron cell bodies.

Results: A specific gp140‐trkA‐like IR was found in: (1) a subpopulation (65%) of primary sensory neuron cell bodies, including most of the largesized ones but also small‐ and intermediate‐sized ones; (2) most of sympathetic neuron cell bodies (82%); (3) theineurial cell, Schwann cells, and large axons of the nerve trunks supplying digital skin; (4) the lamellar cells of Meissner corpuscles; (5) the central axon, inner‐core, outer‐core, and capsule of Pacinian corpuscles. In addition, the occurrence of gp140‐trkA‐like IR was observed in some non‐nervous tissues of the skin, including epidermis (mainly in the basal layer), sweat glands, and arterial blood vessels.

Conclusions: Present results provide evidence for the localization of gp140‐trkA‐like IR in: (1) nerve cells which are known to be NGF‐responsive, and (2) non‐nervous cutaneous tissues which are innervated by NGF‐dependent peripheral neruons. These findings suggest that, in addition to the well‐established role of NGF on sensory and sympathetic neurons, this neurotrophin may be able to regulate some other functions on non‐nervous cell which are targets for NGF‐dependent peripheral neurons. © 1994 Wiley‐Liss, Inc.
Phagocytic defence mechanism in sea bass (Dicentrarchus labrax L.): An ultrastructural study
Authors: Angeles Esteban M. , Meseguer José
Published: February 2005

Background: The ultrastructure of the phagocytic process in fish has not been established in spite of the significant morphofunctional differences detected in the fish immune system with respect to the basic immunological pattern in vertebrates. We report the ultrastructure of the bacterial phagocytic defence mechanism in sea bass (Dicentrarchus labrax L.).

Methods: Head‐kidney, blood, and peritoneal exudate leukocytes were challenged with Aeromonas salmonicida and Escherichia coli and processed for transmission electron microscopic study.

Results: Macrophages challenged with bacteria showed changes in the cell outline, in the chromatin pattern, and in the ultrastructural features of the cytoplasm as a consequence of an activation process. The phagocytic process consists of the following: (1) Bacteria‐macrophage contact. One or more spot contacts between the bacterial wall and the phagocyte membrane are observed. (2) Bacteria engulfment. Slight depressions, membrane invaginations, or cytoplasmic processes are formed at the phagocyte surface. Macrophage processes occasionally surround the bacteria, overlaping and roaming parallel, or a single, long pseudopod encircles a bacterium several times. (3) Endocytic vesicle formation. Macrophages show one or more bacteria inside membrane‐bound cytoplasmic vesicles. (4) Phagolysosome formation. Some dense granules (lysosomes) fuse with the endocytic vesicle. (5) Intracelular killing/digestion. Bacteria inside the endocytic vesicles are observed both virtually intact or damaged at different digestion stages.

Conclusions: Sea bass macrophages possess the mechanisms necessary to both engulf and kill bacteria. Cellular and subcellular events in the morphology of phagocytosis and lysosomal dissolution of bacteria fit the general pattern described for mammals. © 1994 Wiley‐Liss, Inc.
Anatomy of the hyoid apparatus in Odontoceti (toothed whales): Specializations of their skeleton and musculature compared with those of terrestrial mammals
Authors: Reidenberg Joy S. , Laitman Jeffrey T.
Published: February 2005

Background: The hyoid apparatus of odontocetes (toothed whales) serves as a major attachment point for many of the muscles and ligaments subserving breathing, swallowing, and sound production.

Methods: This study examines the hyoid apparatus in 48 specimens of ten odontocete genera (Phocoena, Lagenorhynchus, Stenella, Delphinus, Tursiops, Grampus, Globicephala, Mesoplodon, Physeter, and Kogia) collected post mortem from beach strandings.

Results: The odontocete hyoid apparatus, as that of their closest terrestrial relatives, the artiodactyls, is divisible into a basal portion (bashyal, paired thyrohyals) and a suspensory portion (paired ceratohyals, epihyals, stylohyals, and tympanohyals) connecting the basal portion to the skull base. Unlike other terrestrial mammals, the basal portion lies inferior to the laryngeal aditus, is flattened dorso‐ventrally, and is relatively large, thus providing a broad surface area for muscle attachments. The suspensory elements are not as flattened and are joined by synovial joints (except for epihyal‐stylohyal fusion). Muscular specializations include enlargement of those which retract the hyoid apparatus (e.g., sternohyoid) or control the tongue (e.g., styloglossus, hyoglossus). These muscles may be particularly important in a specialized prey capture behavior called suction feeding. In addition, the hyoid apparatus has a tilted placement, which allows asymmetrical enlargement of the piriform sinuses. Asymmetry is also seen in the muscular attachment between the larynx and the hyoid apparatus. The most pronounced differences from the basic pattern are observed in two families: Physeteridae and Ziphiidae.

Conclusions: The derived position and shape of the odontocete hyoid apparatus may have evolved to subserve several specialized upper respiratory/digestive tract functions, such as simultaneous feeding (suction and swallowing) and sound production. © 1994 Wiley‐Liss, Inc.
Variation in articular cartilage in rabbits between weeks six and eight
Authors: Oreja Maria Teresa Castaño , Rodriguez Maximino Quintáns , Abelleira Antonio Crespo , García Manuel Avelino Giráldez , García Miguel Angel Saavedra , Barreiro Francisco Javier Jorge
Published: February 2005

Background: Previous studies of variation in cartilage characteristics with age have involved comparison of young and adult individuals, but no data on short‐term age‐related change are available. Such data are important for studies of the response of cartilage to experimental stimuli in young rabbits, to distinguish the response to the stimuli from accompanying age‐related changes.

Methods: We used light microscopy to study the thickness, cell density, and degree of histological definition of articular cartilage on the femoral trochlea of 6‐, 7‐, and 8‐week‐old rabbits.

Results: Thickness and cell density both decline significantly with age. The decline in cell density is more marked in surface layers of cartilage and is accompanied by an increase in the safranin O‐staining affinity of the extracellular matrix and an extension of this affinity towards the surface.

Conclusions: Our results indicate that the synthesis of matrix components becomes more important relative to proliferative activity. The traditionally defined histological layers (zones I, II, III, and IV) are not clearly distinguishable in rabbits of this age. In 6‐ and 7‐week old animals only a “surface” (I/II) and a “deep” layer (III) can be distinguished. By 8 weeks, zones I and II are well defined but the mineralization front (marking the boundary between zones III and IV) is still absent. © 1995 Wiley‐Liss, Inc.
Integrin and phosphotyrosine expression in normal and migrating newt keratinocytes
Authors: Donaldson Donald J. , Mahan James T. , Yang Hui , Yamada Kenneth M.
Published: February 2005

Background: Cells interact with the extracellular matrix through a family of cell surface receptors known as integrias. Ligand specificity of a given integrin is determined in part by the type of α and the type of β subunit comprising it. Accumulating evidence suggests that integrinligand binding in some systems influences cell behavior through tyrosine phosphorylation of intracellular proteins.

Methods: In this study, we utilized immunohistochemistry to examine the expression of β1 and β4 integrin subunits as well as tyrosine phosphorylation in normal keratinocytes and in keratinocytes migrating to form a wound epithelium. An adhesion assay was used to determine if freshly isolated keratinocytes could interact with fibronectin and collagen. Polyacrylamide gel electrophoresis followed by immunoblotting was employed to compare β1 integrins in migrating and nonmigrating keratinocytes.

Results: In normal epidermis, β1 and β4 localized primarily to basal cells, where both subunits were generally distributed over all parts of the cell periphery. Except for a modest presence in suprabasal cells and a minimal presence adjacent to the epidermal basement membrane, phosphotyrosine (ptyr) had a similar distribution. In migrating keratinocytes, β1, β4, and ptyr localized most heavily at the interface between the forming wound epithelium and the wound bed. Adhesion assays using keratinocytes from normal epidermis revealed a population of cells that could specifically adhere and spread on fibronectin and type I collagen. Immunoblots of β1 subunits from normal and migrating keratinocytes showed no increase in amount of β1, nor did the apparent size of β1 change in migrating compared to normal cells.

Conclusions: The heavy accumulation of β1 and β4 at the wound bed interface in migrating cells suggests that these subunits may be involved in attachments of migrating cells to extracellular matrix proteins in the wound. The accumulation of ptyr in the same region further suggests that integrin‐ligand interaction in keratinocytes modulates cell behavior through phosphorylated proteins. The fact that freshly isolated newt keratinocytes could adhere and spread on fibronectin or collagen shows that these cells are constitutively activated. This view is supported by the absence of any evidence that the β1 in migrating keratinocytes is larger and therefore more mature than β1 in normal keratinocytes. By comparison, β1 integrins on human keratinocytes are not constitutively activated (Takashima and Grinnell, 1985; Toda et al., 1987; Guo et al., 1990, 1991), a difference that may explain why epidermal wound healing is faster in newts than in humans. © 1995 Wiley‐Liss, Inc.
Changes in the arrangement of actin filaments in myoid cells and sertoli cells of rat testes during postnatal development and after experimental cryptorchidism
Authors: Maekawa Mamiko , Kazama Hirohito , Kamimura Kyoko , Nagano Toshio
Published: February 2005

Background: Abundant actin filaments are present in myoid cells and Sertoli cells in the testis. In the adult rat, the filaments form a lattice arrangement within the myoid cell, and show a hexagonal pattern in the basal junctional regions of Sertoli cells.

Methods: Isolated seminiferous tubules and frozen sections were prepared from juvenile to adult Wistar rat testes, stained with FITC‐conjugated phalloidin, and observed by confocal microscopy. Unilateral cryptorchidism was induced in adult rats, and seven days later, their testes were also examined.

Results: In the myoid cell, parallel actin filaments running circularly around the seminiferous tubules were observed at 15 and 20 days of age. Then, at 30 days, actin filaments arranged longitudinally along the tubular long axis appeared in addition to the circular bundles. A lattice arrangement of actin‐filament bundles in myoid cells became obvious at 40 days, when elongated spermatids are found in the tubule. Actin filaments in the basal junctional regions of Sertoli cells did not acquire the hexagonal pattern seen in the adult testis until 30 days of age. In the cryptorchid testes, the arrangement of actin filaments in the both cells showed a remarkable change compared to the control testis; the filaments became thinner and disrupted.

Conclusions: A lattice arrangement of the actin filaments in the myoid cell appear at around 30 days, before the completion of spermatogenesis. A hexagonal pattern of the filaments in the junctional regions of Sertoli cells has already developed at this age. Cryptorchidism affects the actin filaments of the both cells. © 1995 Wiley‐Liss, Inc.
Actin, α‐actinin, and spectrin with specific associations with the postacrosomal and acrosomal domains of bovine spermatozoa
Authors: Yagi Ahmed , Paranko Jorma
Published: February 2005

Background: Characteristic membrane changes in spermatozoa culminating in acrosome reaction and sperm‐egg fusion, and suspected involvement of actin‐containing cytoskeleton in membrane changes in general, prompted us to investigate subcellular distribution of actin and actin‐binding proteins in bovine spermatozoa subjected to various extractions which sequentially denude the sperm investments.

Methods: Spermatozoa were treated with either 1% SDS, 0.1% Triton X‐100, 0.1% Hyamine, or 1 M MgCl2 or were sonicated. Immunostaining of actin, α‐actinin, spectrin, and acrosin as well as electron microscopic analysis of extracted spermatozoa were carried out.

Results: Extractions caused evagination of the acrosomal lamina which retained focal contacts with the inner acrosomal membrane. Extractions further revealed lateral prongs at the anterior border of the postacrosomal sheath. Labeling for α‐actinin and spectrin was localized in the acrosinpositive acrosomal lamina, neck, and principal piece, the latter containing also relatively extraction‐resistant oligomeric or polymerized actin. In the postacrosomal area, actin was accumulated in the extraction‐resistant posterior ring structure and anteriorly at the sites apparently related to the lateral prongs. Notably, spectrin reactivity was enhanced by MgCl2 in head, neck, and principal piece, and sonication abolished cytoskeletal immunoreactivity in the head.

Conclusions: Destabilization of membranes with selected extractions induces changes in the acrosomal lamina mimicking acrosomal vesicle formation. The lateral prongs and posterior ring structure, respectively, may serve as anterior and posterior anchors for the extraction‐resistant post‐acrosomal sheath. The lateral prongs may also be a merger zone for actin, α‐actinin, and spectrin with important implication on sperm function. The latter two proteins may be involved in acrosomal vesicle formation. It is apparent that extractions have a significant effect on the detectability of sperm cytoskeletal elements. © 1995 Wiley‐Liss, Inc.
Isolation of intramitochondrial helical filaments appearing in outer compartment of mitochondria
Authors: Sasaki Hiroyuki , Kurioka Susumu , Fukata Hiroyuki , Ohoki Takao , Arai Hisako , Suzuki Teruo
Published: February 2005

Background: Packets of helical filaments have been observed in the outer compartment of occasional mitochondria in many cell types in a variety of animals. The compositon and funcation of these intramitochondrial helical filaments (IMHF) are unknown.

Methods: IMHF wre induced in a hepatic mitochondria by administration of ethanol in the drinking water of rats. Hepatic mitochondria were isolated and ruptured by osmotic shock, relasing their IMHF. To purify these structures, the IMHF‐containing supernatant was further fractionated by ammonium sulfate precipitation, a 50–60% solution of this reagent being the most effective in this regard. Isolated IMHF were examined by electron microcopy and were analyzed by SDS‐PAGE.

Result Isolated IMHF closely resembled their in situ counterparts: they had a right‐handed helical structure with a 16 nm pitch. SDS‐PAGE analysis revealed that they contained three polypeptides with weigh tmolecular of 135, 98, and 53 kD, respectively.

Conculusions: These observations will stand as a aseline for comparisons with IMHF that occur naturally or that are induced in other cell types by other kinds of experimental manipulation. © 1995 Wiley‐Liss, Inc.
Effects of lateral pterygoid muscle hyperactivity on differentiation of mandibular condyles in rats
Authors: Takahashi Ichiro , Mizoguchi Itaru , Nakamura Masanori , Kagayama Manabu , Mitani Hideo
Published: February 2005

Background: The effects of biomechanical stress on the growth and development of the mandibular condyle have been studied by many investigators. However, the role of the lateral pterygoid muscle in this development is not clear.

Methods: Hyperfunction of the lateral pterygoid muscles of male 3‐weekold Sprague‐Dawley rats was induced by electrical stimulation, and the responses of the mandibular condyles were compared to control tissues by a double‐fluorescent staining technique using polyclonal antibodies against type I and type II collagen. Electrical stimulation consisted of repeated application (5 seconds on/5 seconds off) of a Hz current for up to 7 days.

Results: In the first 2 days, cartilaginous tissues rich in type II collagen disappeared in the anterior and posterior areas, which were loaded by tensional force due to direct and indirect attachment of the lateral pterygoid muscles. Tissues in these areas were replaced by intramembranous bone that was reactive for type I collagen at 7 days. By the end of the experiment, the trabecula of the condyle was remodled more perpendicularly, thus resisting the compressive force due to hyperfunction of the lateral pterygoid muscles.

Conclusions: These results suggest that the activity of the lateral pterygoid muscle might play a significant role in the differentiation of progenitor cells and in the maturation and calcification of chondrocytes in mandibular condyles. © 1995 Wiley‐Liss, Inc.
Glycoconjugate expression in the chick embryonic chorioallantoic membrane: Comparisons of the chorionic ectoderm and allantoic endoderm
Authors: Henry Charmaine B. S. , Kleinstein Eric , Shum Wesley , DeFouw David O.
Published: February 2005

Background: The chorioallantoic membrne (CAM) of the chick embryo expands during embryogenesis to meet the increased oxygen demands during growth and differentiation. Temporal and spatial glycosylation patterns of CAM ectodermal and endodermal proteins likely contribute to differentiation of the functional attributes of the CAM.

Methods: Using lectins for light and electron microscopic observations, we studied the patterns of glycoconjugate expression on the ectoderm and endoderm of the chorioallantoic membrane (CAM) of the chick at days 4.5, 5.0, 6.0, and 10 of morphogenesis. For light microscopy, samples of unfixed CAM were incubated with the following FITC lectins: Con A, DBA, GSA‐I, GSA‐II, PNA, SBA, UEA‐I, and WGA.

Results: All lectins, except GSA‐I and ‐II, gave positive results. The positive lectins, labeled with HRP, served to ultrastructurally localize PNA, SBA, and WGA, but not DBA binding to the luminal surface of the endoderm. UEA‐I and Con A bound similarly except on day 10 when UEA‐I no longer bound. On the ectodermal surface, only WGA bound at all times studied. PNA and SBA binding were present from days 5.0 to 6.0 but absent at days 4.5 and 10. DBA binding occurred through day 5.0 but was absent thereafter. UEA‐I bound to the ectoderm at days 4.5, 5.0, and 10 but not days 5.5 and 6.0 Con A bound only on days 5.0 and 10.

Conclusion: That the ultrastructurally similar ectoderm and endoderm of the CAM display functional differences conforms to the hypothesis that differential expression of glycoconjugate microdoains likely contributes to such functional specialization. © 1995 Wiley‐Liss, Inc.