Tissue response to suture materials implanted subcutaneously in a rabbit model

We compared the tissue response to a nonabsorbable monofilamented suture made of expanded polytetrafluoroethylene (ePTFE), which has recently been introduced for use in plastic surgery, with the response to 10 other commercially available absorbable sutures and nonabsorbable monofilamented and multifilamented sutures. The sutures were used to secure a patch of ePTFE implanted in the dorsum of adult New Zealand White rabbits. At 30, 60, and 120 days after implantation, the tissue response to the sutures was assessed with respect to the number of foreign-body giant cells present, the thickness of the fibrous capsule that developed, and the general inflammatory response (n = 4 for each suture for each time period). Analysis of variance revealed that specific suture type was significantly associated with foreign-body giant cell count and fibrous capsule thickness. Tevdek had a significantly higher value for mean number of foreign-body giant cells. Silk and Tevdek had significantly thicker fibrous capsules, and ePTFE suture had a significantly thinner capsule. Absorbable sutures and nonabsorbable multifilamented sutures evoked a more extensive tissue response than monofilamented sutures; the differences between nonabsorbable monofilamented and nonabsorbable multifilamented sutures were significant for capsule thickness. In general, suture made of ePTFE produced a minimal tissue response. It should be a good choice for use in facial plastic surgery, in which excellent functional and aesthetic results are critical.     

Chondrocyte-seeded collagen matrices implanted in a chondral defect in a canine model

The objective of our study was to evaluate reparative tissues formed in chondral defects in an adult canine model implanted with cultured autologous articular chondrocytes seeded in type I and II collagen GAG matrices. Two defects were produced in the trochlea grooves of the knees of 21 dogs, with cartilage removed down to the tidemark. This study includes the evaluation of 36 defects distributed among five treatment groups: Group A, type II collagen matrix seeded with autologous chondrocytes under a sutured type II collagen flap; Group B, type I collagen matrices seeded with chondrocytes under a sutured fascia flap; Group C, unseeded type I collagen matrix implanted under a sutured fascia flap; Group D, fascia lata flap alone; and Group E, untreated defects. All animals were killed 15 weeks after implantation. Six other defects were created at the time of death and evaluated immediately after production as 'acute defect controls'. In three additional defects, unseeded matrices were sutured to the defect and the knee closed and reopened after 30 min to determine if early displacement of the graft was occurring; these defects served as 'acute implant controls'. The areal percentages of four tissue types in the chondral zone of the original defect were determined histomorphometrically: fibrous tissue (FT); hyaline cartilage (HC); transitional tissue (TT, including fibrocartilage); and articular cartilage (AC). New tissue formed in the remodeling subchondral bone underlying certain defects was also assessed. Bonding of the repair tissue to the subchondral plate and adjacent cartilage, and degradation of the adjacent tissues were evaluated. There were no significant differences in the tissues filling the original defect area of the sites treated with chondrocyte-seeded type I and type II matrices. Most of the tissue in the area of the original defect in all of the groups was FT and TT. The areal percentage of HC plus AC was highest in group E, with little such tissue in the cell-seeded groups, and none in groups C and D. The greatest total amount of reparative tissue, however, was found in the cell-seeded type II matrix group. Moreover, examination of the reparative tissue formed in the subchondral region of defects treated with the chondrocyte-seeded collagen matrices (Groups A and B) demonstrated that the majority of the tissue was positive for type II collagen and stained with safranin O. These results indicate an influence of the exogenous chondrocytes on the process of chondrogenesis in this site. In all groups with implants (A-D), 30(50% of the FT and TT was bonded to the adjacent cartilage. Little of this tissue (6-22%) was attached to the subchondral plate, which was only about 50% intact. Remarkable suture damage was found in sections from each group in which sutures were used. Harvest sites showed no regeneration of normal articular cartilage, 18 weeks after the biopsy procedure. Future studies need to investigate other matrix characteristics, and the effects of cell density and incubation of the seeded sponges prior to implantation on the regenerative response.     

Microcarrier enhanced survival of human and rat fetal ventral mesencephalon cells implanted in the rat striatum

The transplantation of tissue containing dopamine-producing cells into the mammalian central nervous system is an emerging treatment for Parkinson's disease, despite relatively poor survival of implanted tissue. Recent evidence has suggested that Cytodex microcarriers enhance the survival of dopaminergic rat chromaffin cells transplanted into the rat striatum in the absence of immunosuppression. The current study was undertaken to evaluate the survival of rat and human fetal ventral mesencephalic neurons (VM) implanted alone or after attachment to microcarriers in the striatum of rats without immunosuppression. Rat fetal VM neurons demonstrated enhanced survival in the rat striatum when transplanted on microcarriers, compared to their transplantation alone during the 3-mo period examined in the present study. Transplants of human fetal VM neurons on microcarriers also survived remarkably well in the rat striatum without systemic immunosuppression. In contrast, human fetal VM cells transplanted alone into the rat striatum did not survive without systemic immunosuppression. There was no evidence of TH fiber sprouting in the vicinity of any transplant site. These data indicated that Cytodex microcarriers provide enhanced survival of both rat allograft and human xenograft fetal mesencephalic cells in the rat striatum without the necessity of systemic immunosuppression, perhaps by inducing a unique neuron-glia environment.     

Cellular distribution of exogenous aprotinin in the rat kidney

Aprotinin, an inhibitor of the enzymatic activity of kallikrein in vitro, has been used to study the possible contributions of the kallikrein-kinin systems to physiological and pathological conditions. Pharmacokinetic studies indicate that aprotinin is concentrated in the kidney; however, there is little information with regard to its cellular distribution. The purpose of the present work was to study the cellular distribution of aprotinin, which would be valuable for a better understanding of its intrarenal effects. Sprague-Dawley rats (200-250g, n = 36) received aprotinin (50000 KIU/rat) and were killed at different intervals after its administration. The kidneys were examined histologically and the cellular distribution of aprotinin was studied by immunohistochemistry. Aprotinin was localized at 30 min concentrated within vesicles in the apical border of the proximal tubule cells. Later (2 h) it was observed distributed over the cytoplasm, where it remained for the 24 h studied. Aprotinin was also detected in connecting tubule cells colocalized with kallikrein, and in the basal portion of collecting tubule cells. No evidence of endogenous aprotinin was observed. The binding of aprotinin to the connecting tubule cells and collecting ducts offers a partial explanation of its renal effects.     

Cellular morphology in the visual layer of the developing rat superior colliculus

Foldback sequences in nuclear DNA from cultured Hamster fibroblasts (BHK-21/C13 cells) have been characterized by electron microscopy. One half of the structures observed when denatured hamster DNA is allowed to anneal in the range O less than Cot1 less than 1 x 10(-4) M sec result from the annealing of inverted sequences forming foldback DNA. The remainder have a probable bimolecular origin. arising from rapidly-annealing sequences of satellite-like complexity. The average length of the inverted sequences in the foldback molecules is about 0.9 kilobases. There is estimated to be about 42,000 such sequences (21,000 pairs) in the hamster genome, approximately 45% of which form looped structures with a mean loop length of 1.74 kilobases. Contrary to previous reports, binding of the renatured duplex molecules to hydroxyapatite results in a poor recovery of structures containing identifiable foldback sequences, due to preferential enrichment of the bound fraction with duplexes formed by intermolecular annealing.     

Cellular mechanisms of hypoxia-induced contraction in human and rat pulmonary arteries

Two experiments examined the effects of age, genetic strain, and nicotine on acoustic startle response (ASR) amplitude and prepulse inhibition (PPI) in rats. ASR amplitude measures reactivity to external stimulation, and PPI is used as an index of sensory gating related to attention. Both ASR amplitude and PPI have been previously reported to be increased by nicotine in adult rats. Experiment 1 examined effects of chronically administered nicotine and saline on ASR and PPI in Wistar, Long-Evans, and Sprague-Dawley rats (40 days of age). Experiment 2 examined the effects of chronically administered nicotine and saline in Sprague-Dawley rats of two age groups: 40 and 70 days of age at the beginning of the study. ASR amplitude differed significantly across strains with the values for Wistar > Sprague-Dawley > Long-Evans, and there were no differences in percent of PPI among the three strains. In addition, results of Experiment 2 indicated that older rats had significantly greater ASR amplitudes and PPI than younger rats. Consistent with previous reports, nicotine increased ASR and PPI in the older rats; however, there were no significant differences in the younger rats. Therefore, age and genetic strain are important variables in the analysis of nicotine's effects on startle behaviors in rats.     

Cellular changes of rat embryonic fibroblasts by an actin-polymerization inhibitor, bistheonellide A, from a marine sponge

Bistheonellide A, an inhibitor of actin polymerization from the marine sponge Theonella sp., was introduced at a concentration of 100 nM into rat fibroblast of 2.4 x 10(4) cells/ml. Within 1 h, it disrupted stress fibers, accompanied by a marked change of the cell morphology, resulting in the formation of processes from the cell surface. Further incubation for 24 h in the presence of 100 nM bistheonellide A led to binucleation in most cells and subsequent inhibition of cell cycle progression. When bistheonellide A was withdrawn from the culture medium, binuclear cells began to grow again within 20 h and reverted to mononuclear morphology. Flow cytometric analysis fluorescence-activated cell sorting showed that 2C diploid DNA content in G1 phase was changed into 4C content of tetraploid for the bistheonellide A treated-cells in G1 phase and into 8C content during G2 and M phase. Therefore, we suggested that the bistheonellide A treatment inhibited cytokinesis, but not mitosis in M phase, and that treated cells were arrested at the early G1 phase. These effects of bistheonellide A on the cell cycle progression of 3Y1 fibroblast were also observed more prominently in cells synchronized in S phase with hydroxyurea. Cells in G0 phase were then activated by the addition of fetal calf serum in the presence of 100 nM bistheonellide A. Cell cycle progression of the bistheonellide A-treated cells was obviously slowed down or completely inhibited during G1 phase. These results reveal that actin filaments are not only essential to cytokinesis but also for promoting the progression of cell cycle G1 to S phase.     

Effect of subperiosteally implanted polytetrafluoroethylene (PTFE) material on alveolar bone in the rat

Dihydroergocryptine is a hydrogenated ergot derivative with pharmacological actions mainly related to its dopaminomimetic activity. Here we report that dihydroergocryptine can protect cultured rat cerebellar granule cells against glutamate-induced neurotoxicity, assessing cell viability with the fluorescein diacetate-propidium iodide technique. Dihydroergocryptine antagonized both the neuronal death produced by acute exposure to a toxic glutamate concentration as well as the normal age-dependent degeneration in culture. The effect of dihydroergocryptine might be mediated by a scavenger action as suggested by the fact that the compound in a concentration-dependent manner reduced the formation of intracellular peroxides produced in cerebellar granule cells by exposure to 100 microM glutamate. This action is apparently not mediated entirely by interactions with the dopamine D2 receptors. The neuroprotective action suggests that dihydroergocryptine might be a potential useful drug in the therapy and/or prophylaxis of acute and chronic neurodegenerative diseases related to excitotoxic damage.     

Age-dependent changes in the oxidative metabolism in rat brain

The effects of aging on the oxidation of labeled glucose and 3-hydroxybutyrate and on several mitochondrial enzymes in rat brain were investigated. The oxidation of labeled glucose and labeled 3-hydroxybutyrate was diminished by about 40 and 35%, respectively, in cerebral cortex slices from 2-year-old rats compared to those from 3-mo-old animals. A significant reduction in the activities of 3-hydroxybutyrate dehydrogenase, 3-oxo acid CoA transferase, acetoacetyl CoA thiolase, and NAD-isocitrate dehydrogenase was observed in brains of 1- and 2-year-old rats compared to 3-mo-old animals. However, aging had no effect on the activities of citrate synthase and pyruvate carboxylase. These findings show that specific alterations occur in the activities of several mitochondrial enzymes in aging brain.     

Cellular and molecular changes during sex differentiation of embryonic mammalian gonads

Structural and molecular changes during sex differentiation and development of mammalian gonads from early embryonic phase until sexual maturity have been studied by morphologic and immunocytochemical methods in vivo and in experimental culture. The strategy has been to identify cellular macromolecules whose genes are differently expressed in the two sexes and to formulate a hypothetical regulatory chain of sex determination. This approach should provide new possibilities for finding the missing links between the final structural genes and the early regulatory genes, which are differentially expressed before and during gonadal differentiation. On the basis of accumulated structural and molecular evidence, the early epithelial differentiation from the precursor cells via cell aggregates to testicular cords or ovarian follicles is not sexually regulated. The biological consequences of sex determination in the differentiation of the genital organs include changes in the pattern formation of the gonadal epithelia and concomitant alterations in the synthesis and organization of the structural macromolecules.     

Cellular distribution and gene regulation of estrogen receptors alpha and beta in the rat pituitary gland

The pituitary gland is a heterogeneous tissue comprised of several hormone secreting and supporting cells, most of which are targeted by estrogens. Estrogen-induced changes in the pituitary are presumably mediated via the classical estrogen receptor, ER alpha. However, a novel receptor, ER beta, and pituitary-specific truncated estrogen receptor products (TERPs) were recently identified. The objectives of this study were to examine the distribution of these receptors in the rat pituitary and compare their regulation by estradiol in Sprague-Dawley and the estrogen-sensitive Fischer 344 rats. Pituitary cryosections were subjected to immunocytochemistry for specific cell types, followed by in situ hybridization for ER alpha or ER beta. ER alpha was expressed by approximately 45% of the lactotrophs and melanotrophs, 35% of the corticotrophs and folliculo-stellate cells, and 25% of the gonadotrophs. The expression of ER beta showed a similar pattern but was generally lower than ER alpha. In two cell types, melanotrophs and gonadotrophs, ER beta expression was significantly lower than ER alpha. In the second experiment, pituitary sections were immunostained for ER alpha, followed by in situ hybridization for ER beta. Only a minute population (6-10%) of either anterior or intermediate lobe cells coexpressed ER alpha and ER beta. In the next experiment, Fischer 344 and Sprague-Dawley rats were injected with oil or estradiol for 24 h. Total RNA from dissected anterior and posterior (neurointermediate) pituitaries was subjected to RT-PCR for ER alpha, ER beta, or TERPs. Interestingly, ER alpha and ER beta were unchanged by estradiol in either lobe of the pituitary. In contrast, estradiol increased pituitary TERP messenger RNA levels 4- to 7-fold. A 20-kDa TERP protein was detected by Western blots in the pituitary but not the uterus. There were no differences in the estradiol-induced expression of any of the receptors between the two strains of rats. We conclude that: 1) ER beta is expressed in all anterior and intermediate lobe cell types examined, albeit at a lower level than ER alpha; 2) no more than 10% of pituitary cells coexpress ER alpha and ER beta; and 3) estradiol markedly increases TERP messenger RNA levels but does not alter the expression of ER alpha or ER beta. We propose that estrogen receptor heterogeneity contributes to the diversity of pituitary cell responsiveness to estrogens.     

The chemical stability of enkephalin-like tetrapeptides in a cannula implanted in the rat neostriatum for multiple microinjections

A method for identification of Listeria in food samples was developed. It consisted of cultivating of suspected specimen on standard agar medium, direct absorption of grown colonies onto nitrocellulose membrane and processing of the latter with rabbit serum raised against purified cell wall protein Lm79/39 of L. monocytogenes. Analysis using anti-rabbit peroxidase conjugate and 4-chloro-1-naphthol and H2O2 solutions allowed direct detection of Listeria colonies which remained readily available for subsequent isolation.     

Pathological and immunocytochemical changes in chronic calcium oxalate nephrolithiasis in the rat

In the present study, we exposed rats to a crystal-inducing diet (CID) consisting of vitamin D3 and 0.5% ethylene glycol (EG), and we investigated histologically the kidney damage induced by the deposition of calcium oxalate (CaOx) crystals. After 28 days, 50% of the animals had renal CaOx crystals, of which 60% also had small papillary stones. Most crystals were present in the cortex. The occurrence of these crystals coincided with morphological and cytochemical changes: glomerular damage, tubular dilatation and necrosis, and an enlargement of the interstitium. The number of epithelial and interstitial cells positive for the proliferating cell nuclear antigen (PCNA) was increased. Tamm-Horsfall protein (THP) was not only demonstrable in the thick ascending limb of the loop of Henle (TAL), but also frequently in glomeruli, in the proximal tubular epithelium, and in the papilla. In the lumen of the tubular system, it was associated with urinary casts. Reflection contrast microscopy (RCM) showed that the crystals were coated with a thin layer of THP. In spite of the high urinary oxalate concentrations, the above described cellular changes were not observed in CID-fed rats without renal crystals. We conclude, therefore, that in the kidney, the retained CaOx crystals rather than the urinary oxalate ions are responsible for the observed morphological and immunocytochemical changes.     

Proliferation in the rat olfactory epithelium: age-dependent changes

Vertebrate olfactory sensory neurons are replaced continuously throughout life. We studied the effect of age on proliferation in olfactory epithelium in postnatal rats ranging in age from birth (P1) until P333. Using BrdU to label dividing cells, we determined the proliferation density of basal cells, i.e., the number of labeled nuclei/unit length (240 microm) of olfactory epithelium in coronal sections from six different anterior-posterior levels from each animal. A total length of >1 m of olfactory epithelium was counted in each age group. We observed a dramatic decrease of proliferation density from P1 through P333. At P1, proliferation density is 151 cells/mm; it decreases to approximately half at P21 (70 cells/mm), and half again at P40 (37 cells/mm). At P333 the proliferation density was only 8/mm, approximately 5% of that seen at P1. The changes were clearly related to age and not to body weight, because the values were essentially identical for males and females of the same age but of different body weight. Proliferating cells appear in patches that, after P40, become more separated from one another and contain fewer cells. In 6- and 11-month-old rats, 30 and 45% of all units contained no labeled cells. We confirmed the data of others that the olfactory surface area continuously increases with age; we showed that there is a reciprocal relationship between proliferation density and surface area. The proliferating cells provide neurons to sustain growth as well as to replace dying cells.     

Comparison of primate and canine models for bone ingrowth experimentation, with reference to the effect of ovarian function on bone ingrowth potential

Bone growth into porous composite (mesh-bead) titanium plugs was compared in elderly (postmenopausal) female monkeys and female dogs as a means of validating the cross-species interpretations so often made between data from research on dogs and human applications. The effect of oophorectomy on bone ingrowth in the canine model was defined by the comparison of data on fractional ingrowth in animals that had had oophorectomy and in control animals that had had a sham operation. No significant difference in bone growth into the experimental plugs was identified between the two animal models, which lends credence to cross-species interpretation of existing data from dogs. The presence or absence of active ovaries did not affect the ingrowth fraction in the canine model; this suggests that existing data are not confounded by the lack of control of ovarian function. Estrogen depletion does not appear to influence bone ingrowth adversely.     

Polarity of epithelial cells of the liver. Cellular and molecular mechanisms, and pathologic changes

Smooth muscle contraction is primarily regulated not only by changes in cytosolic Ca2+ concentrations ([Ca2+]i) but also by changes in the force/[Ca2+]i ratio. The use of membrane-permeabilization technique facilitated demonstration of an increase in the level of force at constant [Ca2+]i (Ca2+ sensitization). It was clarified that Rho-associated kinase (Rho-kinase) is a novel mediator of Ca2+ sensitization of the smooth muscle contraction, by introducing the recombinant catalytic domain of Rho-kinase into the cytosol of vascular smooth muscle permeabilized with beta-escin. This review article focuses on novel mechanisms, by which activation of receptor-coupled G-protein(s) increases Ca2+ sensitivity of the contractile apparatus in smooth muscle: Rho-kinase and protein kinase C.     

Cellular localization of dopamine D2 receptor messenger RNA in the rat trigeminal ganglion

The actions of dopamine are mediated by specific, high-affinity, G protein-coupled receptors. Multiple subtypes of dopamine receptors have been characterized, including the D2 subtype (D2R). Cells within the dorsal root and petrosal ganglia of the rat express D2R messenger RNA (mRNA) consistent with D2R expression by primary sensory neurons. We hypothesized that neurons of the trigeminal ganglion express D2R mRNA. Total cellular RNA from rat trigeminal ganglia was analyzed on Northern blots under high stringency conditions. Hybridization of trigeminal ganglion RNA resulted in a signal which comigrated with striatal, pituitary, and hypothalamic D2R mRNA. To determine the distribution of D2R expressing cells in the trigeminal ganglion, cryostat sections were analyzed by in situ hybridization followed by emulsion autoradiography. We identified a population of clustered cells labeled with dense grain concentrations over their cytoplasms. These findings demonstrate the expression of D2 dopamine receptor mRNA in discrete subpopulations of neurons in the rat trigeminal ganglion. Our observations suggest that drugs active at dopamine receptors of the D2 subtype are potential modulators of sensory activity of neurons whose cell bodies reside in the trigeminal ganglion. D2 dopamine receptors may thus have a role in clinical pain syndromes involving the head and neck.     

Percutaneous computed tomography lymphography in the rabbit by subcutaneously injected nanoparticulates

RATIONALE AND OBJECTIVES: Surgical lymphangiography is infrequently used in staging cancer because of its inherent limitations. Radiopaque nanoparticulates target lymph nodes draining interstitial tissues and could make percutaneous lymphography feasible. METHODS: Experimental nanoparticulate contrast agent formulations were injected subcutaneously in the forepaw or hindpaw of normal rabbits or rabbits with induced reactive nodal hyperplasia. Axillary and popliteal nodes were imaged with thin-section computed tomography (CT) using quantitative methods to measure node enhancement. Dose-response (0.1-2.0 ml) and time course (4 hr to 10 weeks) of enhancement were assessed. RESULTS: Nodal enhancement above 100 Hounsfield units was consistently obtained. Enhancement was significantly related to dose and peaked at 10 hr with slow washout over the observation period. Nodes with reactive hyperplasia were larger and had heterogeneous enhancement patterns distinctly different from normal nodes. CONCLUSION: Percutaneous CT lymphography effectively depicts the macroscopic intranodal architecture in rabbits.     

Developmental changes of tetrahydrobiopterin in rat and human erythrocytes

In the present study, we investigated the developmental changes of (1) plasma and erythrocyte tetrahydrobiopterin (BH4); (2) erythrocyte GTP cyclohydrolase (the rate-limiting enzyme of BH4 biosynthesis); (3) the permeability of erythrocyte membrane to BH4; and (4) plasma phenylalanine, both in healthy human subjects and Wistar rats. In vitro experiments demonstrated passive transport of BH4 into erythrocytes. In humans, BH4 levels as well as the other parameters were fairly consistent across all age groups. In contrast, Wistar rats showed significant developmental changes in erythrocyte BH4, which were not simply correlated to either GTP cyclohydrolase, permeability to BH4 or plasma phenylalanine levels. This may suggest the existence of other factors regulating the homeostasis of BH4, such as BH4-binding capacity in plasma and/or erythrocytes. These species/age differences in erythrocyte characteristics may influence the pharmacological behavior and clinical efficacy of BH4 in humans and experimental animals.