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Histochemical Analysis (histochemical + analysis)
Selected AbstractsHistometric and Histochemical Analysis of the Effect of Trichloroacetic Acid Concentration in the Chemical Reconstruction of Skin Scars MethodDERMATOLOGIC SURGERY, Issue 10 2006SUNG BIN CHO MD BACKGROUND Atrophic scars can be induced by various causes, including severely inflamed acne, chicken pox, and trauma. Many treatment modalities are used for reconstructing and improving the appearance of scars with various treatment results. OBJECTIVE A recent report shows the clinical efficacy of the chemical reconstruction of skin scars (CROSS) method, which consists of the focal application of trichloroacetic acid (TCA) in a higher concentration. Histometric analysis of the CROSS method, however, has not yet been established. METHODS In this study, five hairless mice were used to evaluate the effect of the CROSS method and to analyze the difference between the CROSS method and simple TCA application. RESULTS Similar histologic changes were observed in the two methods, including epidermal and dermal rejuvenation with new collagen deposition. These changes, however, were more prominent in the CROSS method,treated areas, particularly when 100% TCA was used. CONCLUSION The results of this study suggest that treatment of atrophic scars using the CROSS method is more effective than simple application of TCA in activating fibroblasts in the dermis and increasing the amount of collagen. [source] Histochemical Analysis of Glycoconjugates in the Skin of a Catfish (Arius Tenuispinis, Day)ANATOMIA, HISTOLOGIA, EMBRYOLOGIA, Issue 1 2010A. Al-Banaw Summary A histochemical study using conventional carbohydrate histochemistry (periodic-acid staining including diastase controls, alcian blue staining at pH 1 and 2.5) as well as using a battery of 14 fluorescein isothiocyanate (FITC)-labelled lectins to identify glycoconjugates present in 10 different areas of the skin of a catfish (Arius tenuispinis) was carried out. The lectins used were: mannose-binding lectins (Con A, LCA and PSA), galactose-binding lectins (PNA, RCA), N -acetylgalactosamine-binding lectins (DBA, SBA, SJA and GSL I), N -acetylglucosamine-binding lectins (WGA and WGAs), fucose-binding lectins (UEA) and lectins which bind to complex carbohydrate configurations (PHA E, PHA L). Conventional glycoconjugate staining (PAS staining, alcian blue at pH 1 and 2.5) showed that the mucous goblet cells contain a considerable amount of glycoconjugates in all locations of the skin, whereas the other unicellular gland type, the club cells, lacked these glycoconjugates. The glycoproteins found in goblet cells are neutral and therefore stain magenta when subjected to PAS staining. Alcian blue staining indicating acid glycoproteins was distinctly positive at pH 1, but gave only a comparable staining at pH 2.5. The mucus of the goblet cells therefore also contains acid glycoproteins rich in sulphate groups. Using FITC-labelled lectins, the carbohydrate composition of the glycoproteins of goblet cells could be more fully characterized. A distinct staining of the mucus of goblet cells was found with the mannose-binding lectins LCA and PSA; the galactosamine-binding lectins DBA, SBA and GLS I; the glucosamine-binding lectin WGA; and PHA E which stains glycoproteins with complex carbohydrate configurations. No reaction occurred with the fucose-binding lectin UEA and the sialic acid-specific lectin SNA. In addition, the galactose-binding lectins PNA and RCA showed only a weak or completely negative staining of the mucus in the goblet cells. The specificity of the lectin staining could be proved by inhibiting binding of the lectins by competitive inhibition with the corresponding sugars. From these data, we can conclude that the mucus produced by the epidermal goblet cells of A. tenuispinis is rich in mannose, N -acetylgalactosamine and N -acetylglucosamine residues. [source] Lectin Histochemical Analysis of the Olfactory Bulbs in the Barfin Flounder (Verasper moseri)ANATOMIA, HISTOLOGIA, EMBRYOLOGIA, Issue 1 2010N. Nakamuta Summary Several lines of evidence have shown that the olfactory system of the fish contains the main and accessory olfactory systems. However, morphological data indicate that the accessory olfactory bulb, the primary centre for the accessory olfactory system, will not differentiate in the fish. Therefore, the fish olfactory bulb is supposed to engage in both main and accessory olfactory systems. To examine this possibility, we investigated the olfactory bulb of the barfin flounder (Verasper moseri) by histochemical examination using lectins. The olfactory bulb of the barfin flounder showed a laminar structure with four layers, and diffuse glomerular architecture was observed in the glomerular layer. Based on the expression patterns of sugar residues, the glomerular layer of the barfin olfactory bulb was largely divided into three portions. Heterogeneity in the lectin-binding pattern among olfactory glomeruli was clearly demonstrated by the fluorescent double-lectin staining. The results of this study suggest that the fish olfactory bulb contains both regions equivalent to the main and accessory olfactory bulbs, and they are subdivided into small subsets with different functions. [source] Expression of CP4 EPSPS in microspores and tapetum cells of cotton (Gossypium hirsutum) is critical for male reproductive development in response to late-stage glyphosate applicationsPLANT BIOTECHNOLOGY JOURNAL, Issue 5 2006Yun-Chia Sophia Chen Summary Plants expressing Agrobacterium sp. strain CP4 5-enolpyruvylshikimate-3-phosphate synthase (CP4 EPSPS) are known to be resistant to glyphosate, a potent herbicide that inhibits the activity of the endogenous plant EPSPS. The RR1445 transgenic cotton line (current commercial line for Roundup Ready® Cotton) was generated using the figwort mosaic virus (FMV) 35S promoter to drive the expression of the CP4 EPSPS gene, and has excellent vegetative tolerance to glyphosate. However, with high glyphosate application rates at developmental stages later than the four-leaf stage (late-stage applications: applications that are inconsistent with the Roundup® labels), RR1445 shows male sterility. Another transgenic cotton line, RR60, was generated using the FMV 35S promoter and the Arabidopsis elongation factor-1, promoter (AtEF1,) for the expression of CP4 EPSPS. RR60 has excellent vegetative and reproductive tolerance to applications of glyphosate at all developmental stages. Histochemical analyses were conducted to examine the male reproductive development at the cellular level of these cotton lines in response to glyphosate applications, and to investigate the correlation between glyphosate injury and the expression of CP4 EPSPS in male reproductive tissues. The expression of CP4 EPSPS in RR60 was found to be strong in all male reproductive cell types. Conversely, CP4 EPSPS expression in RR1445 was low in pollen mother cells, male gametophytes and tapetum, three crucial male reproductive cell types. Our results indicate that the FMV 35S promoter, although expressing strongly in most vegetative tissues in plants, has extremely low activity in these cell types. [source] Down-Regulation of Lignin Biosynthesis in Transgenic Leucaena leucocephala Harboring O -Methyltransferase GeneBIOTECHNOLOGY PROGRESS, Issue 3 2006Smita Rastogi In the present study, a 0.47 kb OMT gene construct from aspen, encoding for an enzyme O -methyltransferase (OMT, EC 2.1.1.6), in antisense orientation was used to down-regulate lignin biosynthesis in Leucaena leucocephala. The plants were transformed with Agrobacterium tumefaciens strain harboring the antisense gene, and the transformation was confirmed by PCR amplification of the npt II gene. The integration of a heterologous antisense OMT gene construct in transformed plants led to a maximum of 60% reduction in OMT activity relative to control. The evaluation of total lignin content by the Klason method revealed a maximum of 28% reduction. Histochemical analyses of stem sections depicted a reduction in lignin content and normal xylem development. The results also suggested a probable increase in aldehyde levels and a decrease in syringyl units. Lignin down-regulation was accompanied by an increase in methanol soluble phenolics to an extent that had no impact on wood discoloration, and the plants displayed a normal phenotype. Concomitantly, an increase of up to 9% in cellulose content was also observed. Upon alkali extraction, modified lignin was more extractable as evident from reduced Klason lignin in saponified residue and increased alkali soluble phenolics. The results together suggested that the extent of down-regulation of OMT activity achieved may lead to quality amelioration of Leucaena with respect to its applicability in pulp and paper manufacture as well as nutritive and easily digestible forage production. [source] Neuroprotective mechanisms of curcumin against cerebral ischemia-induced neuronal apoptosis and behavioral deficitsJOURNAL OF NEUROSCIENCE RESEARCH, Issue 1 2005Qun Wang Abstract Increased oxidative stress has been regarded as an important underlying cause for neuronal damage induced by cerebral ischemia/reperfusion (I/R) injury. In recent years, there has been increasing interest in investigating polyphenols from botanical source for possible neuroprotective effects against neurodegenerative diseases. In this study, we investigated the mechanisms underlying the neuroprotective effects of curcumin, a potent polyphenol antioxidant enriched in tumeric. Global cerebral ischemia was induced in Mongolian gerbils by transient occlusion of the common carotid arteries. Histochemical analysis indicated extensive neuronal death together with increased reactive astrocytes and microglial cells in the hippocampal CA1 area at 4 days after I/R. These ischemic changes were preceded by a rapid increase in lipid peroxidation and followed by decrease in mitochondrial membrane potential, increased cytochrome c release, and subsequently caspase-3 activation and apoptosis. Administration of curcumin by i.p. injections (30 mg/kg body wt) or by supplementation to the AIN76 diet (2.0 g/kg diet) for 2 months significantly attenuated ischemia-induced neuronal death as well as glial activation. Curcumin administration also decreased lipid peroxidation, mitochondrial dysfunction, and the apoptotic indices. The biochemical changes resulting from curcumin also correlated well with its ability to ameliorate the changes in locomotor activity induced by I/R. Bioavailability study indicated a rapid increase in curcumin in plasma and brain within 1 hr after treatment. Together, these findings attribute the neuroprotective effect of curcumin against I/R-induced neuronal damage to its antioxidant capacity in reducing oxidative stress and the signaling cascade leading to apoptotic cell death. © 2005 Wiley-Liss, Inc. [source] Abnormal accumulation of citrullinated proteins catalyzed by peptidylarginine deiminase in hippocampal extracts from patients with Alzheimer's diseaseJOURNAL OF NEUROSCIENCE RESEARCH, Issue 1 2005Akihito Ishigami Abstract Citrullinated proteins are the products of a posttranslational process in which arginine residues undergo modification into citrulline residues when catalyzed by peptidylarginine deiminases (PADs) in a calcium ion-dependent manner. In our previous report, PAD2 expressed mainly in the rat cerebrum became activated early in the neurodegenerative process. To elucidate the involvement of protein citrullination in human neuronal degeneration, we examined whether citrullinated proteins are produced during Alzheimer's disease (AD). By Western blot analysis with antimodified citrulline antibody, citrullinated proteins of varied molecular weights were detected in hippocampal tissues from patients with AD but not normal humans. Two of the citrullinated proteins were identified as vimentin and glial fibrillary acidic protein (GFAP) by using two-dimensional gel electrophoresis and MALDI-TOF mass spectrometry. Interestingly, PAD2 was detected in hippocampal extracts from AD and normal brains, but the amount of PAD2 in the AD tissue was markedly greater. Histochemical analysis revealed citrullinated proteins throughout the hippocampus, especially in the dentate gyrus and stratum radiatum of CA1 and CA2 areas. However, no citrullinated proteins were detected in the normal hippocampus. PAD2 immunoreactivity was also ubiquitous throughout both the AD and the normal hippocampal areas. PAD2 enrichment coincided well with citrullinated protein positivity. Double immunofluorescence staining revealed that citrullinated protein- and PAD2-positive cells also coincided with GFAP-positive cells, but not all GFAP-positive cells were positive for PAD2. As with GFAP, which is an astrocyte-specific marker protein, PAD2 is distributed mainly in astrocytes. These collective results, the abnormal accumulation of citrullinated proteins and abnormal activation of PAD2 in hippocampi of patients with AD, strongly suggest that PAD has an important role in the onset and progression of AD and that citrullinated proteins may become a useful marker for human neurodegenerative diseases. © 2005 Wiley-Liss, Inc. [source] Histochemical analysis of lymphatic endothelial cells in lymphostasisMICROSCOPY RESEARCH AND TECHNIQUE, Issue 2 2001Rui-Cheng Ji Abstract The ultrastructure of endothelial cells of intestinal lymphatics and the thoracic duct (TD) and the relation to lymphostasis were examined in rats and monkeys. Localization of 5,-nucleotidase (5,-Nase) and endothelial nitric oxide synthase (eNOS) was studied. In normal lymphatic endothelial cells, 5,-Nase reaction product was evenly deposited on the cell surface in vivo and on cultured TD endothelial cells (TDECs), whereas eNOS was evenly distributed throughout the nucleus and cytoplasm. TDECs had a long filamentous process extending towards the subendothelial extracellular matrix but became flat and regular within 30,40 minutes after gastric perfusion with olive oil. According to their electron-density, two types of cells were found in the TD endothelial layer. The cells with low electron-density exhibited stronger 5,-Nase activity. Valves were bicuspid formations and the valvular endothelial surface of the convex side showed weaker 5,-Nase activity than the concave side. During TD blockage-induced lymphostasis in rats, the 5,-Nase product was almost not discernible in the TDECs within 2 weeks. Larger vesicles were found in the endothelial cytoplasm of the ligated TD. Their number decreased after 6,12 weeks. The small intestinal lymphatics in the mucosa and submucosa were dilated, with numerous open intercellular junctions. The endothelial lining appeared to have reduced activities for 5,-Nase and eNOS in 9 of 11 experimental animals. The results indicated that the inability of the open intercellular junctions, normally working as one-way endothelial flap valves, may be a key morphological feature after TD blockage. Reduced eNOS and 5,-Nase may functionally influence contractile activity and transport capability of the lymphatic vessels in the lymphostasis. Microsc. Res. Tech. 55:70,80, 2001. © 2001 Wiley-Liss, Inc. [source] The cell-cycle promoter cdc2aAt from Arabidopsis thaliana is induced in the lateral roots of the actinorhizal tree Allocasuarina verticillata during the early stages of the symbiotic interaction with FrankiaPHYSIOLOGIA PLANTARUM, Issue 3 2007Mame Ourèye Sy The symbiosis between the actinorhizal tree Allocasuarina verticillata and the actinomycete Frankia leads to the formation of root nodules inside which bacteria fix atmospheric nitrogen. Actinorhizal nodule organogenesis starts with the induction of cell divisions in the root cortex and in the pericycle cells opposite protoxylem poles near Frankia -infected root hairs. To study the ability of Frankia to induce progression through the cell cycle, we monitored the expression of the ,-glucuronidase (gus) gene driven by the promoter from cdc2aAt, an Arabidopsis cyclin-dependent kinase gene that displays competence for cell division, during plant growth and nodule ontogenesis. In non-symbiotic tissues, the gus gene was mainly expressed in primary and secondary meristems of roots and shoots. Auxins and cytokinins were found to induce reporter gene activity in the root system of whole plants, showing that the promoter cdc2aAt displayed the same regulation by hormones in Allocasuarina as that reported in Arabidopsis. In transgenic nodules, gus expression was found to be restricted to the phellogen. During the early stages of the interaction between Frankia and the plant root system, cdc2aAt was strongly induced in the lateral roots surrounded by hyphae of the actinomycete. Histochemical analysis of ,-glucuronidase activity revealed that cells from the pericycle opposite protoxylem poles were very deeply stained. These data indicate that upon Frankia infection, cells from the lateral roots, and notably pericycle cells that can give rise to a nodule or a root primordium, prepare to re-enter the cell cycle. [source] Changes in skeletal muscle size, fibre-type composition and capillary supply after chronic venous occlusion in ratsACTA PHYSIOLOGICA, Issue 4 2008S. Kawada Abstract Aim:, We have previously shown that surgical occlusion of some veins from skeletal muscle results in muscle hypertrophy without mechanical overloading in the rat. The present study investigated the changes in muscle-fibre composition and capillary supply in hypertrophied muscles after venous occlusion in the rat hindlimb. Methods:, Sixteen male Wistar rats were randomly assigned into two groups: (i) sham operated (sham-operated group; n = 7); (ii) venous occluded for 2 weeks (2-week-occluded group; n = 9). At the end of the experimental period, specimens of the plantaris muscle were dissected from the hindlimbs and subjected to biochemical and histochemical analyses. Results:, Two weeks after the occlusion, both the wet weight of plantaris muscle relative to body weight and absolute muscle weight showed significant increases in the 2-week-occluded group (,15%) when compared with those in the sham-operated group. The concentrations of muscle glycogen and lactate were higher in the 2-week-occluded group, whereas staining intensity of muscle lipid droplets was lower in the 2-week-occluded group than those in the sham-operated group. The percentage of type I muscle fibre decreased, whereas that of type IIb fibre increased in the 2-week-occluded group when compared with the sham-operated group. Although the expression of vascular endothelial growth factor-188 mRNA increased, the number of capillaries around the muscle fibres tended to decrease (P = 0.07). Conclusion:, Chronic venous occlusion causes skeletal muscle hypertrophy with fibre-type transition towards faster types and changes in contents of muscle metabolites. [source] Histochemical evidence of osteoclastic degradation of extracellular matrix in osteolytic metastasis originating from human lung small carcinoma (SBC-5) cellsMICROSCOPY RESEARCH AND TECHNIQUE, Issue 2 2006Minqi Li Abstract The aim of this study was to assess the dynamics of osteoclast migration and the degradation of unmineralized extracellular matrix in an osteolytic metastasis by examining a well-standardized lung cancer metastasis model of nude mice. SBC-5 human lung small carcinoma cells were injected into the left cardiac ventricle of 6-week-old BALB/c nu/nu mice under anesthesia. At 25,30 days after injection, the animals were sacrificed and their femora and/or tibiae were removed for histochemical analyses. Metastatic lesions were shown to occupy a considerable area extending from the metaphyses to the bone marrow region. Tartrate resistant acid phosphatase (TRAPase)-positive osteoclasts were found in association with an alkaline phosphatase (ALPase)-positive osteoblastic layer lining the bone surface, but could also be localized in the ALPase-negative stromal tissues that border the tumor nodules. These stromal tissues were markedly positive for osteopontin, and contained a significant number of TRAPase-positive osteoclasts expressing immunoreactivity for CD44. We thus speculated that, mediating its affinity for CD44, osteopontin may serve to facilitate osteoclastic migration after their formation associated with ALPase-positive osteoblasts. We next examined the localization of cathepsin K and matrix metallo-proteinase-9 (MMP-9) in osteoclasts. Osteoclasts adjacent to the bone surfaces were positive for both proteins, whereas those in the stromal tissues in the tumor nests showed only MMP-9 immunoreactivity. Immunoelectron microscopy disclosed the presence of MMP-9 in the Golgi apparatus and in vesicular structures at the baso-lateral cytoplasmic region of the osteoclasts found in the stromal tissue. MMP-9-positive vesicular structures also contained fragmented extracellular materials. Thus, osteoclasts appear to either select an optimized function, namely secreting proteolytic enzymes from ruffled borders during bone resorption, or recognize the surrounding extracellular matrix by mediating osteopontin/CD44 interaction, and internalize the extracellular matrices. Microsc. Res. Tech. 69:73,83, 2006. © 2006 Wiley-Liss, Inc. [source] Increased friction coefficient and superficial zone protein expression in patients with advanced osteoarthritisARTHRITIS & RHEUMATISM, Issue 9 2010C. P. Neu Objective To quantify the concentration of superficial zone protein (SZP) in the articular cartilage and synovial fluid of patients with advanced osteoarthritis (OA) and to further correlate the SZP content with the friction coefficient, OA severity, and levels of proinflammatory cytokines. Methods Samples of articular cartilage and synovial fluid were obtained from patients undergoing elective total knee replacement surgery. Additional normal samples were obtained from donated body program and tissue bank sources. Regional SZP expression in cartilage obtained from the femoral condyles was quantified by enzyme-linked immunosorbent assay (ELISA) and visualized by immunohistochemistry. Friction coefficient measurements of cartilage plugs slid in the boundary lubrication system were obtained. OA severity was graded using histochemical analyses. The concentrations of SZP and proinflammatory cytokines in synovial fluid were determined by ELISA. Results A pattern of SZP localization in knee cartilage was identified, with load-bearing regions exhibiting high SZP expression. SZP expression patterns were correlated with friction coefficient and OA severity; however, SZP expression was observed in all samples at the articular surface, regardless of OA severity. SZP expression and aspirate volume of synovial fluid were higher in OA patients than in normal controls. Expression of cytokines was elevated in the synovial fluid of some patients. Conclusion Our findings indicate a mechanochemical coupling in which physical forces regulate OA severity and joint lubrication. The findings of this study also suggest that SZP may be ineffective in reducing joint friction in the boundary lubrication mode at an advanced stage of OA, where other mechanisms may dominate the observed tribological behavior. [source] OCTN2 is associated with carnitine transport capacity of rat skeletal musclesACTA PHYSIOLOGICA, Issue 1 2010Y. Furuichi Abstract Aim:, Carnitine plays an essential role in fat oxidation in skeletal muscles; therefore carnitine influx could be crucial for muscle metabolism. OCTN2, a sodium-dependent solute carrier, is assumed to transport carnitine into various organs. However, OCTN2 protein expression and the functional importance of carnitine transport for muscle metabolism have not been studied. We tested the hypothesis that OCTN2 is expressed at higher levels in oxidative muscles than in other muscles, and that the carnitine uptake capacity of skeletal muscles depends on the amount of OCTN2. Methods:, Rat hindlimb muscles (soleus, plantaris, and the surface and deep portions of gastrocnemius) were used for Western blotting to detect OCTN2. Tissue carnitine uptake was examined by an integration plot analysis using l -[3H]carnitine as a tracer. Tissue carnitine content was determined by enzymatic cycling methods. The percentage of type I fibres was determined by histochemical analysis. Results:, OCTN2 was detected in all skeletal muscles although the amount was lower than that in the kidney. OCTN2 expression was significantly higher in soleus than in the other skeletal muscles. The amount of OCTN2 was positively correlated with the percentage of type I fibres in hindlimb muscles. The integration plot analysis revealed a positive correlation between the uptake clearance of l -[3H]carnitine and the amount of OCTN2 in skeletal muscles. However, the carnitine content in soleus was lower than that in other skeletal muscles. Conclusion:, OCTN2 is functionally expressed in skeletal muscles and is involved in the import of carnitine for fatty acid oxidation, especially in highly oxidative muscles. [source] Protective effect of curcumin, a Curcuma longa constituent, in early colonic inflammation in rats,DRUG DEVELOPMENT RESEARCH, Issue 6 2009Juan Manuel Sánchez-Calvo Abstract Curcumin, a polyphenol derived from the plant, Curcuma longa, has a variety of pharmacological effects, including chemotherapeutic, anti-inflammatory, antiangiogenic, and antioxidant activities. To gain a better understanding of the effects and mechanisms of action of curcumin on the acute injury caused by intra-colonic administration of acetic acid (AA) in rats, inflammation was assessed by histology and myeloperoxidase activity (MPO; an index of neutrophil infiltration in the mucosa); Th1 and Th2 cytokine production; histological and histochemical analysis of the lesions; nitrite production in colon mucosa; and the expression of iNOS, COX-1 and -2 using Western blotting and inmmunohistochemistry. We also studied the involvement of the p38 MAPK/JNK signalling pathway in the protective effect of curcumin in acute colonic inflammation. Curcumin (50,100,mg/kg/day) reduced the degree of colonic injury, the index of neutrophil infiltration and Th1 cytokine secretion, and increased IL-10 production, reduced colonic levels of nitrites, and reduced COX-2 and iNOS overexpression. A reduction in the activation of p38 and JNK MAPKs was also observed. Thus, we show that the widely used food additive, curcumin reduced the development of AA-induced colitis and alleviated the inflammatory response. Inhibition of MAPK signalling by curcumin could explain the changes on the cytokine Th1/Th2 profile, the reduction of COX-2 and iNOS signaling, as well as the decreased nitrite production in colonic mucosa, suggesting that curcumin may be useful in the treatment of ulcerative colitis. Drug Dev Res, 2009. © 2009 Wiley-Liss, Inc. [source] Anti-inflammatory effects of continuous passive motion on meniscal fibrocartilageJOURNAL OF ORTHOPAEDIC RESEARCH, Issue 5 2005Mario Ferretti Abstract Motion-based therapies have been applied to promote healing of arthritic joints. The goal of the current study was to determine the early molecular events that are responsible for the beneficial actions of motion-based therapies on meniscal fibrocartilage. Rabbit knees with Antigen-Induced-Arthritis (AIA) were exposed to continuous passive motion (CPM) for 24 or 48 h and compared to immobilized knees. The menisci were harvested and glycosaminoglycans (GAG), interleukin-1, (IL-1,), matrix metalloproteinase-1 (MMP-1), cyclooxygenase-2 (COX-2), and interleukin-10 (IL-10) were determined by histochemical analysis. Within 24 h, immobilized knees exhibited marked GAG degradation. The expression of proinflammatory mediators MMP-1, COX-2, and IL-1, was notably increased within 24 h and continued to increase during the next 24 h in immobilized knees. Knees subjected to CPM revealed a rapid and sustained decrease in GAG degradation and the expression of all proinflammatory mediators during the entire period of CPM treatment. More importantly, CPM induced synthesis of the anti-inflammatory cytokine IL-10. The results demonstrate that mechanical signals generated by CPM exert potent anti-inflammatory signals on meniscal fibrochondrocytes. Furthermore, these studies explain the molecular basis of the beneficial effects of CPM observed on articular cartilage and suggest that CPM suppresses the inflammatory process of arthritis more efficiently than immobilization. © 2005 Orthopaedic Research Society. Published by Elsevier Ltd. All rights reserved. [source] Co-localization of von Willebrand factor with platelet thrombi, tissue factor and platelets with fibrin, and consistent presence of inflammatory cells in coronary thrombi obtained by an aspiration device from patients with acute myocardial infarctionJOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 1 2006Y. HOSHIBA Summary.,Background:,Detailed histochemical analysis of coronary thrombi obtained freshly from acute phase of myocardial infarction patients may provide information necessary to understand the mechanism of coronary occlusive thrombus formation. Methods and Results:,Coronary thrombi causing myocardial infarction were obtained from 10 consecutive patients of myocardial infarction in the acute phase, using a newly developed aspiration catheter. All the fixed specimens of coronary thrombi, by hematoxylin and eosin staining, were found to contain three major constituents, namely, platelets, densely packed fibrin and inflammatory cells, including polymorphonuclear and mononuclear cells, although their distribution in each specimen is totally heterogeneous. Immunohistochemical staining revealed the prominent presence of von Willebrand factor (VWF) at the sites of platelet accumulation, presence of tissue factor and platelets at the sites of deposition of fibrin fibrils. It also revealed the presence of CD16-, CD45- and CD34-positive cells, yet the functional roles of these cells have still to be elucidated. There are weak positive correlation between the number of inflammatory cells involved in the unit area of coronary thrombi specimen and the time of collection of the specimens after the onset of chest pain. Conclusions:,In spite of various limitations, our results contain information suggesting the possible role of VWF in platelet-thrombus formation, possible important role played by tissue factor and activated platelets in the formation of fibrin fibrils, and the positive relationship between inflammatory cells migration and the formation of occlusive thrombi in human coronary arteries. [source] Ex vivo and in vivo evaluation of laser-induced thermotherapy for nodular thyroid diseaseLASERS IN SURGERY AND MEDICINE, Issue 7 2009Jörg-P. Abstract Background and Objective The prevalence of thyroid nodules ranges between 2% and 60% depending on the population studied. However, minimally invasive procedures like laser-induced thermotherapy (LITT) are increasingly used to treat tumors of parenchymatous organs and seem to be suitable for singular thyroid nodules as well. Their successful clinical application depends on the induction of sufficiently large lesions and a knowledge of the energy parameters required for complete thermal ablation. The aim of this study was to establish a dose,response relationship for LITT of thyroid nodules. Materials and Methods Thermal lesions were induced in healthy porcine thyroid glands ex vivo (n,=,110) and in vivo (n,=,10) using an Nd:YAG laser (1,064,nm). Laser energy was applied for 300,seconds in a power range of 10,20,W. During the ablation, continuous temperature measurement at a distance of 5 and 10,mm from the applicator was performed. The lesions were longitudinally and transversally measured, and the volume was calculated. Furthermore, enzyme histochemical analysis of the thyroid tissue was performed. Results The maximum inducible lesion volumes were between 0.74,±,0.18,cm3 at a laser power of 10,W and 3.80,±,0.41,cm3 at 20,W. The maximum temperatures after ablation were between 72.9,±,2.9°C (10,W) and 112.9,±,9.2°C (20,W) at a distance of 5,mm and between 49.5,±,2.2°C (10,W) and 73.2,±,6.7°C (20,W) at a distance of 10,mm from the applicator. The histochemical analysis demonstrates a complete loss of NADPH dehydrogenase activity in thermal lesions as a sign of irreversible cell damage. Conclusions This study is the first to demonstrate a dose,response relationship for LITT of thyroid tissue. LITT is suitable for singular thyroid nodules and induces reproducible clinically relevant lesions with irreversible cell damage in an appropriate application time. Lasers Surg. Med. 41:479,486, 2009. © 2009 Wiley-Liss, Inc. [source] A histopathological and lectin-histochemical study of the lining epithelium in postoperative maxillary cystsORAL DISEASES, Issue 5 2002M Maruyama OBJECTIVE:,Histopathological and lectin-histochemical characteristics were studied in the lining epithelium of postoperative maxillary cysts (POMC). MATERIALS AND METHODS:,Histological (HE, PAS, AB), immunohistochemical (CD3 and L26) and lectin (wheat germ agglutinin, WGA; Ulex europaeus agglutinin I, UEA-I; concanavalin A, ConA) stainings were performed in the 360 POMC specimens. The number of goblet cells and inflammatory cells was counted and statistically analyzed. RESULTS:,The lining epithelium was classified into three types based on histopathological characteristics; pseudostratified ciliated epithelium (pSCE), transitional epithelium (TE) and stratified squamous epithelium (SSE). Local infiltration of inflammatory cells into the cyst wall was associated with an increased number of goblet cells in the lining epithelium. The observed association between the infiltration of inflammatory cells and an increase in the number of goblet cells was statistically significant in groups with lining pSCE and TE. Glycoconjugate histochemical analysis revealed that the surfaces of the lining epithelium with squamous metaplasia showed an increased degree of staining reactivity with UEA-I, whereas the staining reactivity with ConA was reduced. Goblet cells were able to be stained with WGA and UEA-I, but showed extremely low reactivity with ConA. CONCLUSION:,Changes in the glycoconjugate expression of the metaplastic lining epithelium and goblet cell development play an important role in the local defense mechanisms against inflammatory factors in POMC. [source] Versican expression during skeletal/joint morphogenesis and patterning of muscle and nerve in the embryonic mouse limbTHE ANATOMICAL RECORD : ADVANCES IN INTEGRATIVE ANATOMY AND EVOLUTIONARY BIOLOGY, Issue 2 2005Holly E. Snow Abstract Versican, an extracellular matrix proteoglycan, has been implicated in limb development and is expressed in precartilage mesenchymal condensations. However, studies have lacked precise spatial and temporal investigation of versican localization during skeletogenesis and its relationship to patterning of muscle and nerve during mammalian limb development. The transgenic mouse line hdf (heart defect), which bears a lacZ reporter construct disrupting Cspg2 encoding versican, allowed ready detection of hdf transgene expression through histochemical analysis. Hdf transgene expression in whole mount heterozygous embryos and localization of versican relative to cartilage, muscle, and nerve tissues in paraffin-embedded limb sections of wild-type embryos from 10.5,14 days postcoitum were evaluated by lacZ histochemistry, immunohistochemistry, and in situ hybridization. Versican was localized within precartilage condensations and nascent cartilages with expression diminishing during maturation of the cartilage model at later time points. Interestingly, versican remained highly expressed in developing synovial joint interzones, suggesting potential function for versican in joint morphogenesis. Isolated myoblasts, incipient skeletal muscle masses, and neurites were not present in areas of strong versican expression within the developing limb. Versican-expressing tissues may reserve space for the future limb skeleton and developing joints and may aid in patterning of muscle and nerve by deterring muscle migration and innervation into these regions. © 2005 Wiley-Liss, Inc. [source] Delivery of an Adenoviral Vector to the Crushed Recurrent Laryngeal Nerve,THE LARYNGOSCOPE, Issue 6 2003Adam Rubin MD Abstract Objectives Objectives were to create a model of recurrent laryngeal nerve injury for testing the efficacy of potential therapeutic viral gene therapy vectors and to demonstrate that remote injection of a viral vector does not cause significant additional neuronal injury. Study Design Animal model. Methods Rats were randomly assigned to three groups of 10 animals each. In group I, the recurrent laryngeal nerve was crushed. In group II, the nerve was crushed and then injected with an adenoviral vector containing no transgene. In group III, the nerve was identified but was not crushed. Rats were killed at 1 week, and their larynges and brainstems were cryosectioned in 15-,m sections. Laryngeal cryosections were processed for acetylcholine histochemical analysis (motor endplates) followed by neurofilament immunoperoxidase (nerve fibers). Percentage of nerve,endplate contact was determined and compared between groups. Fluorescent in situ hybridization was performed on brainstem sections from rats in group II to confirm the presence of virus. Results No significant difference in percentage of nerve,endplate contact exists between the two crushed-nerve groups (groups I and II) (P = .88). The difference between both crushed-nerve groups and the group with noncrushed nerves (group III) was highly significant (P <.0001). The presence of virus was confirmed in group II rats. Conclusions Crush provides a significant measurable injury to the recurrent laryngeal nerve and may be used as a model to explore therapeutic interventions for nerve injury. The remote injection of viral vector did not cause significant additional neuronal injury. Remote delivery of viral vectors to the central nervous system holds promise in the treatment of recurrent laryngeal nerve injury and central nervous system diseases. [source] All-trans retinoic acid affects subcellular localization of a novel BmNIF3l protein: functional deduce and tissue distribution of NIF3l gene from silkworm (Bombyx mori),ARCHIVES OF INSECT BIOCHEMISTRY AND PHYSIOLOGY (ELECTRONIC), Issue 4 2010Jianqing Chen Abstract A novel cDNA sequence encoding a predicted protein of 271 amino acids containing a conserved NIF3 domain was found from a pupal cDNA library of silkworm. The corresponding gene was named BmNIF3l (Bombyx mori NGG1p interacting factor 3-like). It was found by bioinformatics that BmNIF3l gene consisted of five exons and four introns and BmNIF3l had a high degree of homology to other NIF3-like proteins, especially in the N-terminal and C-terminal regions. A His-tagged BmNIF3l fusion protein with a molecular weight of approximately 33.6,kDa was expressed and purified to homogeneity. We have used the purified fusion protein to produce polyclonal antibodies against BmNIF3l for histochemical analysis. Subcellular localization revealed that BmNIF3l is a cytoplasmic protein that responds to all-trans retinoic acid (ATRA). Western blotting and real-time reverse transcription polymerase chain reaction showed that the expression level of BmNIF3l is higher in tissues undergoing differentiation. Taken together, the results suggest that BmNIF3l functions in transcription. © 2010 Wiley Periodicals, Inc. [source] Inflammation and angiogenesis in osteoarthritisARTHRITIS & RHEUMATISM, Issue 8 2003L. Haywood Objective To quantify the relationship between inflammation and angiogenesis in synovial tissue from patients with osteoarthritis (OA). Methods Hematoxylin and eosin staining and histologic grading for inflammation were performed for 104 patients who met the American College of Rheumatology criteria for OA and had undergone total joint replacement or arthroscopy. A purposive sample of synovial specimens obtained from 70 patients was used for further analysis. Vascular endothelium, endothelial cell (EC) proliferating nuclei, macrophages, and vascular endothelial growth factor (VEGF) were detected by immunohistochemical analysis. Angiogenesis (EC proliferation, EC fractional area), macrophage fractional area, and VEGF immunoreactivity were measured using computer-assisted image analysis. Double immunofluorescence histochemical analysis was used to determine the cellular localization of VEGF. Radiographic scores for joint space narrowing and osteophyte formation in the knee were also assessed. Results Synovial tissue samples from 32 (31%) of 104 patients with OA showed severe inflammation; thickened intimal lining and associated lymphoid aggregates were often observed. The EC fractional area, EC proliferation, and VEGF immunoreactivity all increased with increasing histologic inflammation grade and increasing macrophage fractional area. In the synovial intimal lining, VEGF immunoreactivity was localized to macrophages and increased with increasing EC fractional area and angiogenesis. No inflammation or angiogenic indices were significantly correlated with radiographic scores. Conclusion Inflammation and angiogenesis in the synovium are associated with OA. The angiogenic growth factor VEGF generated by the inflamed synovium may promote angiogenesis, thereby contributing to inflammation in OA. [source] Tissue Engineering of Urethra Using Human Vascular Endothelial Growth Factor Gene-Modified Bladder Urothelial CellsARTIFICIAL ORGANS, Issue 2 2008Yong Guan Abstract:, Acquired or congenital abnormalities may lead to urethral damage or loss, often requiring surgical reconstruction. Urethrocutaneous fistula and strictures are common complications, due to inadequate blood supply. Thus, adequate blood supply is a key factor for successful urethral tissue reconstruction. In this study, urethral grafts were prepared by seeding rabbit bladder urothelial cells (UCs) modified with human vascular endothelial growth factor (VEGF165) gene in the decellularized artery matrix. A retroviral pMSCV-VEGF165 -GFP vector was cloned by insertion of VEGF open reading frame into the vector pMSCV-GFP (murine stem cell virus [MSCV]; green fluorescent protein [GFP]). Retrovirus was generated using package cell line 293T. Rabbit UCs were expanded ex vivo and modified with either MSCV-VEGF165 -GFP or control MSCV-GFP retrovirus. Transduction efficiency was analyzed by fluorescence-activated cell sorting. The expression of VEGF165 was examined by immunofluorescence, reverse transcript-polymerase chain reaction, Western blot, and enzyme-linked immunosorbent assay (ELISA). Decellularized rabbit artery matrix was seeded with genetically modified UCs and was subsequently cultured for 1 week prior to subcutaneous implantation into nude mice. Four weeks after implantation, the implants were harvested and analyzed by fluorescence microscopy, and by histologic and immunohistochemical staining. Ex vivo transduction efficiency of UCs was greater than 50% when concentrated retrovirus was used. The modified cells expressed both VEGF and GFP protein. Furthermore, the VEGF-modified UCs secreted VEGF in a time-dependent manner. Scanning electron microscopy and histochemical analysis of cross sections of the cultured urethral grafts showed that the seeded cells were attached and proliferated on the luminal surface of the decellularized artery matrix. In the subcutaneously implanted vessels, VEGF-modified cells significantly enhanced neovascularization and the formation of a urethral layer compared to GFP-modified cells. These results indicate that VEGF gene therapy may be a suitable approach to increase the blood supply in tissue engineering for treatment of urethral damage or loss. [source] Alterations in the epidermal,dermal melanin axis and factor XIIIa melanophages in senile lentigo and ageing skinBRITISH JOURNAL OF DERMATOLOGY, Issue 1 2006N. Ünver Summary Background, Senile lentigo (SL) is a pigmentation disorder that occurs predominantly on the dorsa of the hands, the forearms and the face; its incidence increases with age. Histological hallmarks of SL lesions are hyperpigmentation of the epidermis and elongation of the epidermal rete ridges. Various factors such as , -melanocyte-stimulating hormone, endothelin-1 or stem cell factor are involved in the onset and maintenance of the increased pigmentation. Alterations of the dermal compartment have not yet been analysed in detail in SL. Objectives, To study the occurrence and distribution of melanin in the dermis from SL and aged skin, biopsies from 12 subjects were morphologically analysed by light and electron microscopy in comparison with unaffected skin. Methods, Punch biopsies of SL and adjacent skin from 12 male or female volunteers aged 52,81 years were prepared for light and electron microscopy and samples were analysed by morphological, morphometric, histochemical and immunohistochemical methods. Results, The epidermis from SL revealed morphological features such as hyperpigmentation of basal keratinocytes and the formation of elongated rete ridges. S100+ melanocytes in the stratum basale were not markedly increased, indicating that the hyperpigmentation is predominantly due to changes in melanin synthesis, distribution or turnover. Quantification of epidermal cells expressing the proliferation marker Ki67 did not show an increase of this parameter in SL, indicating that at least in the established lesion cell proliferation is not enhanced. We further focused on the dermal compartment and observed granulated cells which were more abundant in SL. Electron microscopic and histochemical analysis revealed that the granulation of these cells is based on melanosomes, mostly present in large melanosomal complexes. Immunohistochemistry using antibodies to CD68 and factor XIIIa (FXIIIa) showed these melanophages to be predominantly FXIIIa+ dermal dendrocytes, which were about six times more abundant than CD68+ macrophages. Conclusions, In SL an increased number of melanophages was found compared with unaffected skin from the same subject. These melanophages were identified as FXIIIa+ dermal dendrocytes. Possible functional consequences of the massive melanin uptake by dermal dendrocytes are discussed. [source] Microplasmin improves surgical outcome in a rabbit model for trabeculectomyACTA OPHTHALMOLOGICA, Issue 2009E VANDEWALLE Purpose This study was designed to study the efficacy and safety of Microplasmin as an anti-scarring agent after trabeculectomy in a rabbit model. Methods The effect of Microplasmin was investigated in vivo in a rabbit model for glaucoma surgery. Clinical outcome measures were intra-ocular pressure, bleb area and survival, side effects on slit lamp examination. Moreover, (immuno-) histochemical analysis of the eyes was performed, with quantification of inflammation (CD 45) and collagen deposition (Trichrome and Sirius Red). In the first experiment (n=10), Microplasmin anterior chamber injection was compared to placebo injection. In the second experiment (n=3), topical Microplasmin drops were compared to placebo drops. In the third experiment (n=5) the combination of Microplasmin anterior chamber injection and topical drops was compared to placebo injection and drops. All experiments were conducted in a masked observator way. Results Microplasmin significantly augmented the bleb area and survival in a rabbit model of trabeculectomy after a single anterior chamber injection or combination therapy (injection combined with drops) compared to control. Collagen deposition was borderline reduced after Microplasmin administration compared to control. No significant changes in inflammation were noticed in the anterior chamber or in the conjunctiva. Conclusion Microplasmin single injection or combination with postoperative drops improved the outcome after trabeculectomy. In a rabbit model, larger blebs were produced for a longer period compared to control, and collagen deposition tended to decrease in this small series. [source] | |||||||||||||