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PubMed1987

Recovery of free muscle grafts in rat: improvement is associated with an increase in cyclic adenosine monophosphate concentration or use of the condition/test paradigm.

The muscle fibers in freely grafted skeletal muscles degenerate and are replaced by new fibers which develop within the graft. Myogenesis in regenerating muscle recapitulates, to a large extent, developmental myogenesis and may depend on similar modulating influences. In addition to the generation of new fibers, functional recovery of free muscle grafts also requires reinnervation and revascularization of the new fibers. Recovery of function should be improved by enhancing either myogenesis or reinnervation and revascularization. We have used two procedures, shown previously to stimulate peripheral nerve regeneration, to improve the morphologic and functional recovery of free, orthotopic grafts of rat extensor digitorum longus muscle. Each of the procedures was effective, but had potentially different sites of action. The first procedure, the condition/test paradigm, presumably increases the rate and extent of graft reinnervation. The second procedure, continuous infusion of the adenylate cyclase activator forskolin during the first 21 days after grafting, may influence both myogenesis and nerve regeneration. Each procedure increased regenerating muscle fiber size and functional capacity, and forskolin also significantly increased capillary density and fatigue resistance.

Animals↗

PubMed1996

The prepubertal ontogeny of luteinizing hormone releasing hormone-like immunoreactivity in the diencephalon and telencephalon of the male Meishan pig brain.

Luteinizing hormone releasing hormone (LHRH) is a decapeptide that regulates reproductive function and behaviors in mammalian species. Because of the importance of proper reproductive function and efficiency in agricultural species, we have investigated the prepubertal ontogeny of LHRH-like immunoreactivity (IR) in the male Meishan pig. The Meishan is a Chinese breed known for reproductive traits including increased litter size and precocious puberty, but slow growth and obesity. Brains of animals from gestational day (g) 30, 50, 70, 90 and 110 and postnatal day (pn) 1, 10, 20 and 50 (duration of pregnancy averaged 114 days) were processed using a standard immunohistochemical technique utilizing a commercially available rabbit anti-LHRH antibody. LHRH-IR in cell bodies and fibers was detected at g30 entering the brain via the terminal nerve and in the septal region of the basal telencephalon. The number of immunoreactive cells increased at g50 and cells were localized primarily to the septum, organum vasculosum of the lamina terminalis, preoptic area and lateral hypothalamus, whereas immunoreactive fibers were present throughout the septum and hypothalamus and had reached the median eminence. The density and distribution of immunoreactive fibers increased by g70 and g90, but did not change dramatically from g90 to pn50. These results indicate that LHRH may be present in the Meishan brain earlier during development and fibers containing LHRH-IR appear to reach the median eminence earlier than previously reported for the domestic pig. These results suggest a breed difference in the ontogeny of reproductive control systems in the pig. Future studies utilizing direct comparisons between domestic and Chinese breeds will be required to investigate the apparent differences in the ontogeny of LHRH-containing systems in the pig.

Animals↗

PubMed1984

Elastin metabolism and chemistry: potential roles in lung development and structure.

Elastic fibers are important for elasticity and extensibility of lung tissue. In the developing lung, elastic fibers appear in greatest numbers during the process or period of alveolarization . A variety of mesenchymal cells in lung appear responsible for elastin synthesis. Elastin is a novel protein both from the standpoint of its processing into elastic fibers and chemical properties. For example, elastin undergoes posttranslational modification before its assembly into fibers. These steps include limited proteolysis, hydroxylation of prolyl residues and the oxidative deamination of lysyl residues prior to their incorporation into the crosslinks that covalently bond together polypeptide chains of elastin. The crosslinking amino acids include lysinonorleucine , merodesmosine and desmosine isomers. A key enzyme that controls this process is lysyl oxidase. Lysyl oxidase is a copper metalloprotein whose activity is responsive to and modulated by environmental insults, nutrition deficiencies and the administration of various pharmacological agents. Regarding chemical properties, elastin is one of the most apolar proteins secreted by mammalian cells. Moreover, elastin is one of the most long-lived proteins secreted into the extracellular matrix. In relationship to its processing into elastic fibers and chemical properties, details related to major aspects of elastin metabolism as well as speculation on its potential as a factor in lung development and disease are discussed.

Amino Acids↗

PubMed1985

Influence of genetic type, slaughter weight and sex on ovine muscle fiber and fat-cell development.

Histological properties from the longissimus (LD) and semimembranosus (SM) muscles of 51 wether and ewe lambs from Hampshire rams and two ewe genetic types (SR, 1/2 Suffolk and 1/2 Rambouillet, and FS, 1/2 Finnish Landrace and 1/2 Southdown) and three slaughter live weights (32, 41 and 50 kg) were compared. Fibers in both muscles were classified as beta R (red), alpha R (intermediate) or alpha W (white). All LD muscle fiber types from FS ewe lambs increased in diameter from 32 to 41 kg, but decreased in diameter from 41 to 50 kg. Also, this quadratic effect with slaughter weight was found in alpha R and alpha W fibers from the SM muscle of FS wether and ewe lambs. However, diameters of alpha R and alpha W fibers from SR wether and ewe lambs and beta R fibers from SR ewe lambs increased linearly in the SM muscle with increasing slaughter weight. As slaughter weight increased, the proportion of alpha R fibers decreased in both the LD and SM muscles for SR wether and FS wether and ewe lambs. Concurrent with the increase in slaughter weight and decrease in alpha R fiber percentage, the proportion of alpha W fibers increased in the LD muscle of SR wether lambs, the SM muscle of SR ewe lambs and both muscles of FS wether and ewe lambs. Genetic type and sex groups, except SR ewe lambs, support the theory of transformation of alpha R to alpha W fibers with increasing slaughter weights. Fat-cell diameters increased in both muscles of SR wether and FS ewe lambs with increasing slaughter weights. Increases in fat traits of lamb carcasses were related to increases in red-fiber size. Alpha red fiber numbers were inversely related (P less than .05) to alpha W fiber numbers in both the LD and SM muscles (r = -.83 and -.79). The proportion of alpha R to alpha W fibers might be used as an indicator of physiological maturity for lambs.

Adipose Tissue↗

PubMed1987

Elastic fibers in tunica propria of undescended and contralateral scrotal testes from cryptorchid patients.

Elastic fibers in the tunica propria of the testes from cryptorchid patients and normal fertile adults were examined by light (Weigert resorcin-fuchsin stain) and electron microscopic techniques. In the testes from normal fertile adults, elastic fibers were proved to exist in the tunica propria by light and electron microscopy, and were located in the fibrous and cellular layers of the tunica propria. In the undescended testes from the cryptorchid patients, during prepubertal and pubertal periods, elastic fibers could not be visualized in the tunica propria, but were found after puberty. The positive Weigert reaction of the tunica propria in the undescended testes from postpubertal cryptorchid patients, however, was markedly weaker than that in normal control patients, suggesting diminution of elastic fibers. This diminution of elastic fibers in the undescended testes from postpubertal cryptorchid patients was also substantiated by electron microscopy. However, in the contralateral scrotal testes, elastic fibers appeared during puberty and were observed after puberty in the same manner as in normal testes. Thus, the present study suggested retarded appearance of elastic fibers in puberty and impaired development of those fibers after puberty, in the undescended testes of cryptorchid patients.

Adolescent↗

PubMed2002

13C CP/MAS NMR study on structural heterogeneity in Bombyx mori silk fiber and their generation by stretching.

It is important to resolve the structure of Bombyx mori silk fibroin before spinning (silk I) and after spinning (silk II), and the mechanism of the structural transition during fiber formation in developing new silk-like fiber. The silk I structure has been recently resolved by (13)C solid-state NMR as a "repeated beta-turn type II structure." Here, we used (13)C solid-state NMR to clarify the heterogeneous structure of the natural fiber from Bombyx mori silk fibroin in the silk II form. Interestingly, the (13)C CP/MAS NMR revealed a broad and asymmetric peak for the Ala Cbeta carbon. The relative proportions of the various heterogeneous components were determined from their relative peak intensities after line shape deconvolution. Namely, for 56% crystalline fraction (mainly repeated Ala-Gly-Ser-Gly-Ala-Gly sequences), 18% distorted beta-turn, 13% beta-sheet (parallel Ala residues), and 25% beta-sheet (alternating Ala residues). The remaining fraction of 44% amorphous Tyr-rich region, 22% in both distorted beta-turn and distorted beta-sheet. Such a heterogeneous structure including distorted beta-turn can be observed for the peptides (AG)(n) (n > 9 ). The structural change from silk I to silk II occurs exclusively for the sequence (Ala-Gly-Ser-Gly-Ala-Gly)(n) in B. mori silk fibroin. The generation of the heterogeneous structure can be studied by change in the Ala Cbeta peak of (13)C CP/MAS NMR spectra of the silk fibroin samples with different stretching ratios.

Alanine↗

PubMed2003

Subgroup B and F fiber chimeras eliminate normal adenovirus type 5 vector transduction in vitro and in vivo.

Altering adenovirus vector (Ad vector) targeting is an important goal for a variety of gene therapy applications and involves eliminating or reducing the normal tropism of a vector and retargeting through a distinct receptor-ligand pathway. The first step of Ad vector infection is high-affinity binding to a target cellular receptor. For the majority of adenoviruses and Ad vectors, the fiber capsid protein serves this purpose, binding to the coxsackievirus and adenovirus receptor (CAR) present on a variety of cell types. In this study we have explored a novel approach to altering Ad type 5 (Ad5) vector targeting based on serotypic differences in fiber function. The subgroup B viruses bind to an unidentified receptor that is distinct from CAR. The subgroup F viruses are the only adenoviruses that express two distinct terminal exons encoding fiber open reading frames. We have constructed chimeric fiber adenoviruses that utilize the tandem fiber arrangement of the subgroup F genome configuration. By taking advantage of serotypic differences in fiber expression, fiber shaft length, and fiber binding efficiency, we have developed a tandem fiber vector that has low binding efficiency for the known fiber binding sites, does not rely on an Ad5-based fiber, and can be grown to high titer using conventional cell lines. Importantly, when characterizing these vectors in vivo, we find the subgroup B system and our optimal tandem fiber system demonstrate reduced liver transduction by over 2 logs compared to an Ad5 fiber vector. These attributes make the tandem fiber vector a useful alternative to conventional strategies for fiber manipulation of adenovirus vectors.

Adenoviridae↗

PubMed1988

Tissue interactions with dentin specimens after demineralization using tetracycline.

Greater cell attachment to demineralized dentin has been associated with subsequent development of a fiber attachment system. The purpose of this study was to evaluate tissue interactions to dentin demineralized with different concentrations of tetracycline solution. Dentin specimens were obtained from beneath root surfaces covered by periodontal ligament. Each rectangular specimen had a face of root surface dentin and an opposite surface of pulpal dentin. Experimental specimens were treated with a tetracycline solution of either 200 mg/cc or 100 mg/cc for five minutes. The remaining group of specimens served as untreated controls. Specimens were implanted transcutaneously into incisional wounds on the dorsal surface of rats with one end protruding through the skin. Four specimens in each group were available for examination one and ten days after implantation. Histologic and histometric analysis of both root and pulpal surface of implants included counts of adhering cells, assessment of implant length within the connective tissue, and evaluation of connective tissue fiber relationships. In each group, specimens became severely extruded between days one and ten, the number of attached cells decreased, and a fiber attachment system did not develop. Tetracycline-treated surfaces had greater numbers of attached cells at both time points compared to untreated controls. No differences were discernible relating to different tetracycline concentrations. It was concluded that tetracycline-demineralized dentin provided a substrate that increased cell attachment; however, this enhanced response did not result in a connective tissue attachment.

Animals↗

PubMed2002

Characterization of a new in vivo hollow fiber model for the study of progression of prostate cancer to androgen independence.

Research investigating the molecular events underlying progression of prostate cancer to androgen independence has been impeded by the lack of an appropriate in vivo model that yields "pure" populations of prostate cancer cells that are not contaminated with host cells. Here we characterize a new in vivo model that uses hollow fibers and allows for the retrieval of uncontaminated prostate cancer cells during various stages of endocrine progression to androgen independence in male immunocompromised mice. Prostate-specific antigen (PSA) gene expression, proliferation of cells, and histology were examined in these mice before and after castration. LNCaP cells seeded at a density of 1 x 10(7) cells/ml, or a total of approximately 4.8 x 10(6) cells/animal, provided measurable serum PSA levels that increased in intact (noncastrated) animals, decreased by 80% to a nadir after castration, and subsequently increased by 4 weeks after castration, indicating progression to androgen independence. In vivo proliferation of LNCaP cells inside the fibers continued in the presence of androgens and continued to increase, albeit at a slower rate, in the castrated animals. Histology of cells cultivated in hollow fibers demonstrated that initially the cells grew along the wall of the fiber and tended to stack up, forming layers and scaffold structures resembling a solid tumor. Fibers removed from castrated animals with elevated levels of serum PSA contained spheroids of cells that had detached from the fiber wall. The development of the LNCaP hollow fiber model described here provides a reproducible means of obtaining "pure" populations of LNCaP cells during different stages of progression to androgen independence for molecular analysis requiring RNA and protein extracts free of host cell contamination.

Androgens↗

PubMed1983

Optical fiber fluoroprobes in clinical analysis.

We used quartz optical fibers and laser excitation in developing single-fiber fluoroprobes, to be used to measure molecular fluorescence in minute volumes of body fluids. We illustrate and discuss the analytical capabilities of these fluoroprobes. Limits of detection for the anti-tumor drug doxorubicin are about 10(-7) mol/L, by either conventional fluorescence or sequentially excited fluoroscence modes of detection. We also report the results of preliminary in vivo measurements of doxorubicin in the interstitial fluids of human tumors, heterotransplanted in immune-deficient laboratory mice.

Animals↗

PubMed1993

[Pulmonary and pleural reaction patterns to artificial mineral fibers --rockwool].

Epidemiological and animal studies about the fibrogenic and carcinogenic properties of natural mineral fibers required the development of man made vitreous fibers as substitutes for asbestos containing material. The question about the possible fibrogenic and carcinogenic properties of man-made vitreous fibers is not yet answered. By means of different experimental animal studies we tried to investigate the man made vitreous fibers-related pulmonary and pleural diseases. The experimental administration of rockwool induce lesions in the lung of the rats. The lung showed an extensive granulomatous inflammation. In the observation time of 10 months we did not find any malignant tumor of the lung and the pleura.

Animals↗

PubMed1995

Origin of intrafusal fibers from a subset of primary myotubes in the rat.

S46, a monoclonal antibody (mAb) specific for the SM-1 and SM-2 isoforms of avian slow myosin heavy chains (MHC), was used to study the earliest stages of development of intrafusal fibers in muscle spindles of the rat hindlimb. Spindles formed only in the regions of fetal muscles that contained primary myotubes reactive to mAb S46, such as the axial region of the tibialis anterior muscle. The first intrafusal fiber to form, the nuclear bag2 fiber, originated from within the population of S46-reactive primary myotubes. Binding of mAb S46 by myotubes giving rise to the bag2 fibers preceded the appearance of encapsulated spindles in the muscles by electron microscopy. However, reactivity to S46 intensified in the myotubes transforming into bag2 fibers after the innervation of the fibers by afferents, and dissipated in myotubes differentiating into slow-twitch (type I) extrafusal fibers. Thus, afferents may enhance intrafusal expression of the MHC isoform reactive to mAb S46. The pattern of S46 binding to nuclear bag and chain intrafusal fibers in both developing and adult spindles was the same as that reported for the mAb ALD19, suggesting that both antibodies bind to the same MHC isoform. This isoform is probably a developmental form of slow myosin, because it was transiently expressed during the development of type I extrafusal fibers. The origin of bag2 intrafusal and type I extrafusal fibers from a bipotential subpopulation of primary myotubes reactive to mAb S46 correlates with the location of muscle spindles in the slow regions of muscles in adult rat hindlimbs.

Animals↗

PubMed1999

An improved method for exchanging troponin subunits in detergent skinned rat cardiac fiber bundles.

We describe a method for the removal of endogenous troponin (Tn) complex from bundles of detergent-treated cardiac fibers. After 70 min treatment with cTnT-cTnI most of the endogenous Tn complex was removed from fiber bundles. Complete reconstitution of the Tn complex was achieved by reconstituting with cardiac troponin C (cTnC) in fully relaxing conditions. Ca(2+)-dependent maximum force of the fibers treated with cTnT-cTnI or cTnT-cTnI(33-211), which was used to aid in the visualization of the troponin exchange, decreased to 85-90% of the force developed by fibers before the treatment. SDS-PAGE analysis of the cTnT-cTnI(33-211) and the cTnT(77-289)-cTnI(33-211) treated fiber bundles demonstrated that 70-80% of the endogenous Tn subunits were removed. After reconstitution with cTnC, approximately 80-85% of the Ca(2+)-regulated force was restored in cTnT-cTnI/cTnI(33-211) treated fibers. Our results demonstrate that by minimizing the prolonged exposure of skinned cardiac fiber bundles to rigor conditions, successful exchange of all three subunits of the Tn complex can be accomplished with minimal loss of function.

Animals↗

PubMed1991

Expression of stress fibers in bullfrog mesothelial cells in response to tension.

The relationship between stress fibers and tension in mesothelial cells of the bullfrog small intestine was examined by fluorescence cytochemistry using en face mesothelial cell preparations. In nontreated controls, actin revealed by rhodamine-phalloidin staining was localized only along the margins of the mesothelial cells. On the other hand, many stress fibers were formed in the mesothelial cells within 5-7 min after stretching of the intestinal wall in a given direction. The orientation of stress fibers within the cells was coincident with the direction of the tension applied. These cytoplasmic fibers disappeared almost completely from the mesothelial cells within 30 min after the release of tension. According to a difference in the intensity of tension necessary for stress fiber expression, the intestinal mesothelial cells were classified into two groups. Furthermore, cells containing stress fibers in each group showed a rapid increase in number once a given value of tension was applied. The present results indicate that the mesothelial cells of bullfrog small intestine may develop stress fibers to counteract tension exerted on the intestinal wall. Such stress fibers may serve to maintain cellular integrity by strengthening the cellular attachment to subepithelial tissue.

Animals↗

PubMed1998

Fluorescence polarization transients from rhodamine isomers on the myosin regulatory light chain in skeletal muscle fibers.

Fluorescence polarization was used to examine orientation changes of two rhodamine probes bound to myosin heads in skeletal muscle fibers. Chicken gizzard myosin regulatory light chain (RLC) was labeled at Cys108 with either the 5- or the 6-isomer of iodoacetamidotetramethylrhodamine (IATR). Labeled RLC (termed Cys108-5 or Cys108-6) was exchanged for the endogenous RLC in single, skinned fibers from rabbit psoas muscle. Three independent fluorescence polarization ratios were used to determine the static angular distribution of the probe dipoles with respect to the fiber axis and the extent of probe motions on the nanosecond time scale of the fluorescence lifetime. We used step changes in fiber length to partially synchronize the transitions between biochemical, structural, and mechanical states of the myosin cross-bridges. Releases during active contraction tilted the Cys108-6 dipoles away from the fiber axis. This response saturated for releases beyond 3 nm/half-sarcomere (h.s.). Stretches in active contraction caused the dipoles to tilt toward the fiber axis, with no evidence of saturation for stretches up to 7 nm/h.s. These nonlinearities of the response to length changes are consistent with a partition of approximately 90% of the probes that did not tilt when length changes were applied and 10% of the probes that tilted. The responding fraction tilted approximately 30 degrees for a 7.5 nm/h.s. release and traversed the plane perpendicular to the fiber axis for larger releases. Stretches in rigor tilted Cys108-6 dipoles away from the fiber axis, which was the opposite of the response in active contraction. The transition from the rigor-type to the active-type response to stretch preceded the main force development when fibers were activated from rigor by photolysis of caged ATP in the presence of Ca2+. Polarization ratios for Cys108-6 in low ionic strength (20 mM) relaxing solution were compatible with a combination of the relaxed (200 mM ionic strength) and rigor intensities, but the response to length changes was of the active type. The nanosecond motions of the Cys108-6 dipole were restricted to a cone of approximately 20 degrees half-angle, and those of Cys108-5 dipole to a cone of approximately 25 degrees half-angle. These values changed little between relaxation, active contraction, and rigor. Cys108-5 showed very small-amplitude tilting toward the fiber axis for both stretches and releases in active contraction, but much larger amplitude tilting in rigor. The marked differences in these responses to length steps between the two probe isomers and between active contraction and rigor suggest that the RLC undergoes a large angle change (approximately 60 degrees) between these two states. This motion is likely to be a combination of tilting of the RLC relative to the fiber axis and twisting of the RLC about its own axis.

Animals↗

PubMed2003

Ultrahigh-resolution optical coherence tomography by broadband continuum generation from a photonic crystal fiber.

We have developed an ultrahigh-resolution optical coherence tomographic system in which broadband continuum generation from a photonic crystal fiber is used to produce high longitudinal resolution. Longitudinal resolution of 1.3-microm has been achieved in a biological tissue by use of continuum light from 800 to 1400 nm. The system employed a dynamic-focusing tracking method to maintain high lateral resolution over a large imaging depth. Subcellular imaging is demonstrated.

Animals↗

PubMed1990

Eliminating afferent impulse activity does not alter the dendritic branching of the amphibian Mauthner cell.

In the developing amphibian, the formation of extra vestibular contacts on the Mauthner cell (M-cell) enhances dendritic branching, while deprivation reduces it (Goodman and Model, 1988a). The mechanism underlying the interaction between afferent fibers and developing dendritic branches is not known; neural activity may be an essential component of the stimulating effect. We examined the role of afferent impulse activity in the regulation of M-cell dendritic branching in the axolotl (Ambystoma mexicanum) embryo. M-cells occur as a pair of large, uniquely identifiable neurons in the axolotl medulla. Synapses from the ipsilateral vestibular nerve (nVIII) are restricted to a highly branched region of the M-cell lateral dendrite. We varied the amount of nVIII innervation and eliminated neural activity. First, unilateral transplantation of a vestibular primordium deprived some M-cells of nVIII innervation and superinnervated others. Second, surgical fusion of axolotls to TTX-harboring California newt (Taricha torosa) embryos paralyzed the Ambystoma twin: voltage-sensitive Na+ channel blockade by TTX eliminated action potential propagation. Reconstruction of M-cells in 18 mm larvae revealed that dendritic growth was influenced by in-growing axons even in the absence of incoming impulses: impulse blockade had no effect on the stimulation of dendritic growth by the afferent fibers.

Action Potentials↗

PubMed2000

Development of the cavernous sinus in the fetal period: a morphological study.

The development and morphological structure of the lateral sellar compartment (LSC), an interdural space containing the cavernous sinus (CS), cranial nerves, and internal carotid artery (ICA), was investigated by histological examination of sections of the LSC and cerebral venograms from human fetal specimens. Twenty-eight LSC coronal sections were obtained from 14 fetuses of 13-32 weeks' gestation. Venograms of 11 other fetuses of 13-32 weeks' gestation were studied to observe changes in venous drainage. The CS appeared as a collection of small venous canals with an endothelial layer. These venous canals gradually became much larger through expansion and unification. The CS and basilar venous plexus were demonstrated as a faint cluster of small vessels on venograms obtained after 13 weeks' gestation. The dura mater increased in thickness and collagen fiber networks developed around all the components in the LSC after 23 weeks' gestation. The LSC lateral wall could not be histologically differentiated as separate multiple layers. Branching and joining of the cranial nerve fascicles were completed with the envelopment of collagen fibers after 23 weeks' gestation. The ICA at 13-15 weeks' gestation ran straight within the LSC, becoming tortuous before birth. CS formation occurs through the development of venous canals without smooth muscle layers, followed by web-formation by collagen fibers in the mesenchymal interstices. LSC formation, including the dense dura mater and an internal structure like that seen in the adult, is largely completed before birth.

Adult↗