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Transformation of 3DP gypsum model to HA by treating in ammonium phosphate solution.

Three-dimensional printing (3DP) is a CAD/CAM built-up using ink-jet printing technique. Commercially available 3DP system can form only gypsum model and not for bioceramics. On the other hand, transformation of hardened gypsum into hydroxyapatite (HA) by treatment in ammonium phosphate solution was found lately. In the present study, transformation of the 3DP gypsum block to HA was attempted. However, the fabricated 3DP block was soluble in water. To insolubilize, it was heated at 300 degrees C for 10 min, and then, gypsum was transformed to calcium sulfate hemihydrate, CaSO(4) x 0.5H(2)O. The 3D block was immersed in 1M (NH(4))(3)PO(4) x 3H(2)O solution at 80 degrees C for 1-24 h, and the transformation into HA within 4 h was ascertained. A heat-treated plaster of Paris (POP) block was also investigated for comparison. The unheated POP block consisting of gypsum dihydrate took 24 h to complete the transformation, while the heat-treated POP consisting calcium sulfate hemihydrate promoted the transformation into HA; but the transformed thickness in the block was less than the 3DP block. This is probably due to higher solubility of the hemihydrate than gypsum dihydrate. Accelerated transformation of the 3DP block was also caused by its porous structure, which enabled an easy penetration of the phosphate solution. With the present method, it is possible to transform the fabricated gypsum by 3D printing that is adaptive to the osseous defect into HA prostheses or scaffold.

Bone Substitutes↗

The development of lysosomal apparatus. II. Incorporation, subcellular distribution, and intraparticulate hydrolysis of 131 I-albumin by liver of mice at perinatal stages.

Mouse fetuses at 15th and 18th day of gestation, and newborns aging 0, 5, 10, and 15 days were injected i.p. with 131 I-albumin (RISA). The degree of incorporation by liver and the intraparticulate hydrolysis of RISA in 27,000g x 10 min. particles increased after birth. By this time, changes in subcellular distribution of RISA and marker enzymes were also observed. Following an i.p. injection of India ink, numerous hepatic cells stained with carbon particles were observed at the light microscope from day 5 of life. The results suggest that the lysosomal apparatus acquires full capacity to incorporate colloidal particles and to degrade foreign macromolecules in the first week of life.

Acid Phosphatase↗

The effect of limiting light to the pineal on the rate of forelimb regeneration in the newt.

Newts with bilaterally amputated forelimbs were exposed to either continuous light (LL), total darkness (DD), or continuous light with the dorsal head epithelium painted with an India ink and Nile blue sulfate mixture (LL-II-NBS) that limited light penetration through the skull to the pineal. The LL-II-NBS animals regenerated their forelimbs more slowly then their counterparts in LL.

Animals↗

Ectodermal control of the avascular zone of the peripheral mesoderm in the chick embryo.

Prospective skin ectoderm is underlaid by a relatively thick (100 +/- 20 micrometer) avascular zone of mesoderm in most regions of the early embryo. To determine whether or not the ectoderm exercises a role in the establishment and maintenance of the avascular zone, trypsin-isolated pieces of backskin ectoderm from chick or quail embryos were implanted as a sheet into a slit cut deep into the capillary bed of the wing bud of host chick embryos of stages 19-23. In sham operations, slits were cut at various anteroposterior levels, and the wing was allowed to heal. At intervals of 3-48 hr after these operations, embryos were injected with India ink, fixed, and cleared. Implants formed flattened vesicles, usually in continuity with host ectoderm, but sometimes completely internalized. Periderm cells from each side of the vesicle faced each other, and the cells of the cuboidal layer faced an avascular mesodermal layer at least 100 micrometer thick at all points. The implantation of prospective skin ectoderm resulted in the formation of an avascular zone in normally vascularized mesoderm of the wing bud. In contrast, the vascular bed of the limb bud abutted directly on implants of Millipore filters or of Silastic silicone (Dow Corning). Likewise, the capillary bed came in direct contact with implants of retinal pigment epithelium, an ectodermal derivative normally in close contact with the vascular choroid coat of the eye. These results, taken in conjunction with earlier experiments that show the necessity of the apical ectodermal ridge for the formation of the marginal veins of the limb bud, suggest that epithelial-mesenchymal interactions are involved in important aspects of vasculogenesis in early embryos.

Animals↗

Vasculature of the parotoid glands of four species of toads (bufonidae: bufo).

The parotoid glands of toads (Bufonidae) consist of large aggregations of granular glands located between the otic region of the skull and the scapular region. To determine the circulatory pattern of these glands, we perfused the vascular systems of Bufo alvarius, B. marinus, B. terrestris, and B. valliceps with either India ink or Microfil, a fine latex. The perfused glands were studied by gross dissection, microscopic examination, and histology. The vascular patterns of the parotoid glands were compared to the arrangement of vessels in the dorsal skin of Rana sphenocephala (Ranidae), a frog that lacks parotoid glands. The parotoid glands of the four species of toads are supplied with blood by the lateral and dorsal cutaneous arteries and are drained by one or more branches of the internal jugular vein. The dorsal cutaneous artery supplies most of the blood to the parotoid glands in B. terrestris and B. valliceps. In B. alvarius and B. marinus, both the lateral and dorsal cutaneous arteries serve major roles in the blood supply of the glands. These patterns of blood flow have not been described previously for parotoid glands and conflict with earlier accounts for B. alvarius and B. marinus. The arteries and veins associated with the parotoid glands of toads are present in R. sphenocephala, but are arranged differently. In R. sphenocephala, the lateral cutaneous artery supplies the dorsal and lateral skin posterior to the shoulder region, whereas the dorsal cutaneous artery supplies the skin of the shoulder region. In toads, both the lateral and dorsal cutaneous arteries supply the skin of the shoulder region and ramify into subcutaneous capillaries that surround the secretory units of the parotoid glands. Extensive vasculature presumably is important for delivering cholesterol and other precursor molecules to the parotoid glands, where those compounds are converted into toxins.

Animals↗

Structure of long bones in mammals.

Techniques for staining (silver, osmium, metal sulfides, ink) and microphotography (epi-illumination) of polished bone surfaces have been developed to visualize the three-dimensional structure of the shafts of mammalian long bones. Bone is a two-compartment system with capillaries and some kinds of connective tissue in one compartment separated from fibers of bone collagen, often forming lamellae, in the other. Laminar bone consists of stacks of lamellae separated by vascular spaces containing capillary network sheets. It is deposited at the periosteal and endosteal surfaces. Osteonic bone, well described in the literature, consists of cylinders of lamellae with central vascular spaces. The primary structure of the shafts of mammalian long bones is laminar and laminae often remain as the main component. Secondary osteons are a replacement within laminae. As laminar bones mature, some of the irregular longitudinal capillary spaces in the network sheets enlarge and become less crooked to form secondary osteons. Parts of the random networks become ordered longitudinal ones, resulting in collapse of those network spaces not converted to osteons. The residual capillaries become bloodless, making the surviving network spaces difficult to resolve. This may account for them being overlooked in descriptions of bone structure. For example, laminar bone occurs with osteonic bone in the human femur, although it is rarely figured. Nearly mature bones switch the kind of primary bone deposited at the peripheral (periosteal) surface from laminar to primary osteonic.

Animals↗

Lymph drainage of the mammary glands in female cats.

The mammary gland is a common site of neoplasms in the female cat. All the malignant tumors metastasize to a lesser or a greater extent through the lymphatic system. However, the anatomical knowledge of this system is not sufficiently well known in cats to develop a reasoned model for the extirpation of these glands in case of malignant tumors. A study of the lymph drainage in 50 female cats was done by indirect injection in vivo of India ink inside the mammary parenchyma. After a waiting interval, mammary glands were extracted and the thoracic cavity opened. All the lymph nodes were examined after clearing. The success rate of the colorations of lymph nodes and lymph vessels was 91.8%. Out of the 100 observed mammary chains, the two intermediate mammary glands (T2, A1) may drain caudally to the superficial inguinal lymph center and/or cranially to the axillary lymph center. The T1 gland always drains exclusively cranially and A2 exclusively caudally. The two mammary glands (T1 and A1) often drain towards the sternal cranial lymph nodes, but 100% of the T2 drain towards it. This research assumes that the limit between the two directions of drainage can exist only between glands T2 and A1. The results obtained with the study of the 1st, 2nd, 3rd, and 4th mammary glands permit production of new and more complete data of functional significance that will eventually aid block dissection surgical technique in the removal of malignant tumors in cats.

Animals↗

Vasculogenesis of the bursa cloacalis (bursa of Fabricius) of the chick embryo.

Vasculogenesis of the bursa cloacalis (bursa of Fabricius) was examined i 10- to 21-day chick embryos and in chicks during the first 5 days post-hatching. The entire circulatory system was injected with India ink, and the bursae were then removed and either cleared for examination in toto or sectioned serially. The bursa was supplied by three pairs of extrinsic blood vessels. At 10 and 11 days of incubation, most intrinsic vessels were arranged in a superficial, hexagonal network. In regions of developing plicae, the hexagonal plexus extended into the core of each plica, forming middle plical vessels. The latter were interconnected across interplical areas by cross-connecting vessels. The middle plical vessels gave rise to small capillary offshoots, which soon increased in complexity, forming delicate loops. Branches extended from these loops through the subepithelial lamina propria to incipient epithelial buds by 12 days of incubation. All epithelial buds were supplied by at least one such branch, and similar branches extended to the basal aspect of the epithelium in areas where epithelial buds had not yet formed. This observation is consistent with the hypothesis that blood vessels induce formation of epithelial buds. At about 15 days of incubation, the cortex and medulla of each developing lymphatic follicle were defined clearly, and an intricate, web-like, capillary network coursed throughout the follicular cortex. The medulla appeared to be devoid of capillaries. The diameters of all intrinsic and extrinsic bursal blood vessels gradually increased throughout development. During post-hatching stages, the diameters of the extrinsic vessels continued to increase, whereas those of the intrinsic vessels were markedly decreased from late pre-hatching stages.

Animals↗

Adrenergic innervation of the heart of the bat, Miniopterus schreibersii.

The adrenergic innervation of the heart of the bat (Miniopterus schreibersii) was studied with a fluorescence histochemical technique. The appearance and distribution pattern of the terminal adrenergic nerve fibers demonstrated in the atria, sinoatrial and atrioventricular nodes, and ventricles is typically mammalian. Fine varicose adrenergic fibers run in and parallel to the ventricular muscle where they are common and uniformly distributed. Indian ink perfusion of the coronary vasculature demonstrates the high density of vessels in the ventricles, but obscures the terminal innervation of the ventricular muscle. This alone, or in combination with the apparent seasonal change in the terminal innervation of the ventricles, may explain the inability of previous workers to identify an adrenergic ventricular innervation in the bat Nyctalus noctula. The adrenergic ventricular innervation might be involved in the large increases in cardiac output associated with the commencement of flight and in the massive sympathetic activation that mediates arousal from torpor.

Animals↗

Diffusion tensor MRI of myocardial fibers and sheets: correspondence with visible cut-face texture.

PURPOSE: To test the hypothesis that the primary, secondary, and tertiary eigenvectors of the diffusion tensor (DT) measured with DT-MRI correspond to the fiber, sheet, and sheet normal directions, respectively, we compared DT-MRI data with the texture visible in the cut face of fresh bovine myocardium. MATERIALS AND METHODS: DT-MRI and optical images obtained under identical conditions were compared objectively. Ink prints were made of the cut tissue, and the local orientations within these images were defined by analysis of local autocorrelations for regions matching DT-MRI pixels. Deviation angles between the cleavage orientations and the diffusion eigenvectors were analyzed in eight specimens sliced in three orthogonal planes. RESULTS: Root-mean-square (RMS) angular disparity was 11 degrees between the first eigenvectors of the DT and the fiber direction, 14 degrees between the second eigenvector and the sheet direction, 14 degrees between the third eigenvector and the sheet normal direction, and 15 degrees between the tensor orientation in the imaging plane and the cleavage orientation of the cut face. CONCLUSION: The results support a parallel relationship between the eigenvectors of the DT and symmetry axes of the myocardial architecture. Specifically, the first, second and third eigenvectors correspond to the fiber, sheet, and sheet normal directions, respectively.

Animals↗

Thin-layer chromatography/electrospray ionization triple-quadrupole linear ion trap mass spectrometry system: analysis of rhodamine dyes separated on reversed-phase C8 plates.

The direct analysis of separated rhodamine dyes on reversed-phase C(8) thin-layer chromatography plates using a surface sampling/electrospray emitter probe coupled with a triple-quadrupole linear ion trap mass spectrometer is presented. This report represents continuing work to advance the performance metrics and utility of this basic surface sampling electrospray mass spectrometry system for the analysis of thin-layer chromatography plates. Experimental results examining the role of sampling probe spray end configuration on liquid aspiration rate and gas-phase ion signal generated are discussed. The detection figures-of-merit afforded by full-scan, automated product ion and selected reaction monitoring modes of operation were examined. The effect of different eluting solvents on mass spectrum signal levels with the reversed-phase C(8) plate was investigated. The combined effect of eluting solvent flow-rate and development lane surface scan rate on preservation of chromatographic resolution was also studied. Analysis of chromatographically separated red pen ink extracts from eight different pens using selected reaction monitoring demonstrated the potential of this surface sampling electrospray mass spectrometry system for targeted compound analysis with real samples.

Journal Article↗

Characterization of ramified microglia in tissue culture: pinocytosis and motility.

Functional properties of ramified microglia were investigated in primary cultures of rat cerebral cortical cells. These microglia could be readily identified in both fixed and living cultures through previously established features. Based on their destruction by 5 mM L-leucine methyl ester, a high level of intrinsic endocytotic activity was established. When cultures were incubated with fluorescent latex beads to assess phagocytosis, little or no such activity was exhibited by ramified cells. However, when cultures were incubated with dyes or other soluble tracer compounds, these cells always exhibited labeling. This labeling was selective for ramified microglia in the cultures and was demonstrated using a variety of compounds, including trypan blue, lucifer yellow, horseradish peroxidase (HRP), and India ink. Intracellular label could be observed in vesicular structures; this localization corresponded to an active cellular process. Also, cellular labeling was inhibited by the presence of colchicine. These features supported the inference that the labeling was attributable to pinocytosis, and this process appeared to account for the vast majority of endocytotic activity in the ramified microglia. Possible physiological significance of this pinocytotic activity was indicated by the accumulation of various neurotransmitters/modulators: gamma-aminobutyric acid and vasoactive intestinal polypeptide (VIP). Ramified cells in these cultures have been previously noted to exhibit a constant and rapid pattern of motility, which was consistently observed here through time-lapse video recording; pinocytosis and rapid motility were shown to concur in individual cells. Based on their high intrinsic pinocytotic activity and pattern of cellular motility, the ramified microglia specifically are suggested to serve a constitutive function of fluid cleansing within the interstitial spaces of brain tissue.

Animals↗

Involvement of two genes of superinfecting phage kappa in curing and induction of prophage psi in Serratia marcescens HY.

Prophage psi carried along with prophage y by Serratia marcescens HY is subject of moderate curing at heteroimmune superinfection of cells from stationary phase with phage kappa. Curing becomes considerably more frequent when the bacteria are non-lysogenic for y. Both psi, y-double-lysogenic and psi-single-lysogenic cells with a mutation in the ink gene are very efficiently cured of psi if infected by kappa tay, although this mutant was characterized as being deficient in transactivation of certain genes in prophage y. On the other hand to get efficiently cured after kappa wild-type infection these cells too must be devoid of a y prophage. Thus a y function turned on by tay+ seems to counteract the elimination of psi. However, interestingly enough psi curing is boosted by a further y function under special circumstances. Efficient curing depends on an intact kappa tap gene, a gene reported to cause transactivation of certain psi genes. Curing at kappa tay infection is specifically accompanied by induction of the psi prophage in a part of the infected cells. However, there is no such induction at kappa wild-type infection, either in the absence or presence of a y prophage. An explanation of these findings is suggested which includes an antirepressive effect exerted on psi and a hypothetical interaction between the products of genes tap and tay.

Bacteriophages↗

Vascular reorganization and return of rigidity in fracture healing.

The present study addresses the relationship between vascular reorganization and the biomechanical changes occurring during canine long bone fracture healing. A middiaphyseal osteotomy of the radius was performed, and angiographic studies were carried out using India ink injections from 2 to 12 weeks postosteotomy. The radii were harvested, subjected to nondestructive four-point bend testing on an Instron, and then sectioned and cleared according to the Spaltholtz technique. The observed microangiographic patterns were correlated with the calculated rigidity values. Quantitative bone blood flow measurements around the osteotomy were performed on a different group of animals, using the radiotracer microsphere technique, at 3, 6, and 9 weeks postosteotomy to correlate with our qualitative microangiographic results. We demonstrated a direct and predictable relationship between the phase of vascular reorganization and the rigidity. This correlation was present even in our delayed union specimens.

Angiography↗

Role of interfragmentary strain in fracture healing: ovine model of a healing osteotomy.

It has been hypothesized that the histological pattern of fracture healing is controlled at least in part by the local mechanical strains in the interfragmentary region. To test this "interfragmentary strain hypothesis," we applied cyclic bending deformations to tibial osteotomies in 11 sheep. An instrumented flexible plate spanning a 1-mm osteotomy gap was deformed to create a gradient of tissue elongation from 10% under the plate to 100% at the opposite cortex. The cyclic deformations were applied three times per minute, 24 h per day, for 1-5 weeks. However, as a result of tissue differentiation, the bone-plate complex increased in stiffness with healing time, resulting in a marked reduction of the gap deformation at approximately 4 weeks. Fracture healing was evaluated using vascular injection of India ink and conventional histology. A nonlinear three-dimensional finite element model of the interfragmentary tissue at the initial stage of healing was used to predict the complex tissue strains. The ingrowth of vascularized soft tissue into the interfragmentary gap, as well as the subsequent differentiation of this tissue, occurred earlier and to a greater degree in regions of lower strain. In contrast, the proliferation of callus tissue was greatest at the periosteal and endosteal surfaces of the cortex opposite the plate. Direct comparison of the finite element predictions with the histology demonstrated that the spatial distribution of bone resorption at the fracture fragment ends directly corresponded to the locations of elevated tissue strain and stress. However, there was no consistent numerical relationship between the magnitude of these local peak strains and the corresponding volume of cortical bone resorption over the bone cross section.

Animals↗

Vascular assessment of the periarticular ligaments of the rabbit knee.

While the rabbit is being extensively utilized in animal models for orthopaedic research, the vascular anatomy of the knee ligaments has not been thoroughly described in this species. This study demonstrates the blood supply to the infrapatellar fat pad, the cruciate ligaments, the medial collateral ligament (MCL), and the menisci, such that the effects of manipulating these tissues may be properly interpreted. Vascular injection with India ink and iodinated i.v. contrast dye was performed in 11 New Zealand white rabbits, and routine histology done on six. The large vessel anatomy is similar to that described for humans and dogs, with a descending geniculate artery, medial and lateral superior and inferior geniculate arteries, a posterior geniculate artery, and a recurrent anterior tibial artery. The microvascular anatomy is also similar in that the infrapatellar fat pad and synovial membrane are highly vascular, the menisci are vascularized from their periphery (being avascular centrally), and the medial collateral ligament is relatively well vascularized. A difference from dogs and humans is present in the anterior cruciate ligament (ACL), which is poorly vascularized, with a single artery on its anterior aspect. High magnification histologic evaluation reveals numerous capillaries in the substance of the MCL, while the ACL is nearly devoid of such vessels. The interspecies variation in vascular anatomy is a variable that must be taken into consideration in any surgical or traumatic animal model investigation of knee pathology.

Animals↗

Physical properties of rabbit articular cartilage after transection of the anterior cruciate ligament.

The effect of unilateral transection of the anterior cruciate ligament on the confined compression and swelling properties of the distal femoral articular cartilage of skeletally mature rabbits at 9 weeks after surgery was determined. Gross morphological grading of the transected and contralateral control distal femora stained with India ink confirmed that cartilage degeneration had been induced by ligament transection. Osteochondral cores, 1.8 mm in diameter, were harvested from the medial femoral condyles. The modulus, permeability, and electrokinetic (streaming potential) coefficient of the articular cartilage of the osteochondral cores were assessed by confined compression creep experiments. The properties (mean +/- SD) of control cartilage were: confined compression modulus, 0.75 +/- 0.28 MPa; hydraulic permeability, 0.63 +/- 0.28 x 10(-15) m2/Pa*sec; and electrokinetic coefficient, 0.16 +/- 0.31 x 10(-9) V/Pa. In transected knees, the modulus was reduced by 18% (p = 0.04), while the permeability and electrokinetic coefficient were not detectably altered. The change in modulus was accompanied by a trend (p = 0.07) toward a decrease (-11%) in the glycosaminoglycan density within the tissue, a significant increase (p < 0.001) in the water content of the cartilage after equilibration in 1 x phosphate buffered saline from 70.3 +/- 4.1% in control knees to 75.2 +/- 4.0% in transected knees, and little further swelling after tissue equilibration in hypotonic saline. The compressive modulus of the cartilage from both control and transected knees was positively correlated with the density of tissue glycosaminoglycan. The alterations in the physical properties of the articular cartilage after transection of the anterior cruciate ligament in the rabbit show trends similar to those observed in human and other animal models of osteoarthritis and provide further support for the use of this model in the study of cartilage degeneration.

Animals↗

Role of skin cooling in improving patient tolerability of Q-switched Alexandrite (QS Alex) laser in nevus of Ota treatment.

BACKGROUND: The development of a high-energy laser device with very short pulse duration has revolutionized the treatment of nevus of Ota. Nevertheless, patients still suffer from complications that range from the pain and swelling that occur immediately after laser surgery to post-operative pigmentary changes and scarring. However, the simultaneous use of skin surface cooling and laser surgery may limit the damage of non-target tissue. OBJECTIVES: To assess whether epidermal cooling reduces the pain and swelling that commonly occur after laser treatment for nevus of Ota. STUDY DESIGN/PATIENTS AND METHODS: Thirty-seven patients with nevus of Ota were recruited from a dermatology outpatient clinic. Before treatment, the research nurse used an ink pen to divide the lesions into two halves. Half of each lesion was treated with a Q-switched Alex laser system that had a cool sapphire plate in contact as a mean of skin cooling. The other half was treated with the same laser, but with the cooling device switched off. Immediately after treatment, and again 1 week later, the patients answered a questionnaire, which assessed the symptoms that are associated with laser surgery. Dose assessment was performed in each half of the nevus to obtain the optimal fluence that could be used for the treatment of nevus of Ota before the entire half was treated. RESULTS: There was no difference in terms of the optimal fluence that was used, but in terms of immediate pain the patients associated the side that was treated with the cooling plate with a significantly lesser degree of pain than the non-cooled side (P = 0.001). Eighty two percent of the patients preferred the cooled side to the non-cooled side. CONCLUSIONS: Pre- and post-operative skin cooling is effective in improving the tolerability of nevus of Ota patients to Q-switched laser treatment. However, the use of cooling during laser treatment of nevus of Ota did not allow the use of higher fluence.

Adult↗