ELECTRON MICROSCOPY OF THE HUMAN SYNOVIAL MEMBRANE

The structure of the lining cells at the surface of the synovial membrane facing the joint cavity has been studied by electron microscopy. The long cytoplasmic processes of these cells appear to be oriented toward the surface of the membrane, where they overlap and intertwine. The matrix of the lining cells contains dense material but no fibers with the periodicity of collagen. The lining cells are divided into two cell types or states of activity on the basis of their cytoplasmic contents. Type A is more numerous and contains a prominent Golgi apparatus, numerous vacuoles (0.4 to 1.5 microns in diameter) containing varying amounts of a dense granular material, many filopodia, mitochondria, intracellular fibrils, and micropinocytotic-like vesicles. Type B contains large amounts of ergastoplasm with fewer large vacuoles, micropinocytotic-like vesicles, and mitochondria. The probable functions of these cells are discussed in the light of current knowledge of the metabolism and function of the synovial membrane.     

Estrogen participation in induction of cervicovaginal adenosis-like lesions in immature mice exposed prenatally to diethylstilbestrol

The structure of human zygapophyseal joint synovial folds as seen by high-power light microscopy and transmission electron microscopy is described. Small myelinated nerves are demonstrated in association with some capillaries in the synovial folds. This may have clinical significance in the field of spinal pain.     

Effect of monochromatic red helium-neon laser on the morphology of zymosan arthritis in rats

Histological and electron microscopy findings suggesting the effect of monochromatic coherent polarized red light on zymosan arthritis in rats are presented. The data obtained have confirmed that treatment of inflamed joints with laser rays exerts an evident therapeutic effect which is determined by activated function of macrophages and fibroblasts of the synovial membrane, resulting in a more rapid change of inflammation phases. Analysis of the cases where the rats were treated with ordinary red light has revealed no essential differences in the joints of the control and irradiated groups.     

Ultrastructural characteristics of the human synoviocytes

The purpose of this study was to identify, by the scanning electron microscopy, the behaviour of the different cell types in the normal human synovial intima, in order to obtain information useful for interpreting pathological changes in the synovium. Our observations revealed that, in numerous areas of the synovial membrane (adipose or fibrous type), the synoviocytes were dispersed and the intercellular matrix, covered only by the cytoplasmic processes of cells deeply located, was in direct contact with the joint cavity. In the areolar type of the synovium the synoviocytes were more numerous; they tended to concentrate to give the appearance of a continuous tissue; but between the cells very large intercellular spaces were usually present. In this latter membrane type we identified the two main cellular types of the synoviocytes: A and B. B-synoviocytes were the predominant cell type of the synovium. These cells were characterized by long cytoplasmic processes, perpendicularly directed towards the joint cavity. Both the cellular body and the cytoplasmic processes were covered by small blebs and vesicles of various size. The A-synoviocytes were a small minority, rarely dispersed between the B-synoviocytes. They were characterized by numerous membrane infoldings which delimited intracellular canaliculi of various depth. Our ultrastructural observations demonstrated that, in normal conditions, the B-synoviocyte must be considered as a constitutive element which characterized the synovial intima, responsible for the specific structure of the interstitial tissue and for the regulation of the composition of the synovial fluid.(ABSTRACT TRUNCATED AT 250 WORDS)     

Differentiation of temporomandibular joint synovial mesenchymal stem cells into neuronal cells in vitro: an in vitro study

SMSCs (synovial mesenchymal stem cells) isolated from TMJs (temporomandibular joints) were induced to proliferate and differentiate in vitro by bFGF (basic fibroblast growth factor) and explore the potential of SMSC differentiation into neuronal cells. In this study, the cultured SMSCs were derived from the TMJ synovial membrane of condylar hyperplasia patients and were amplified with the indicated concentration of FCS (fetal calf serum) and DMEM (Dulbecco's modified Eagle's medium) in vitro. bFGF (25 ng/ml) was applied to induced synovial cells differentiated into neuronal cells. Inverted microscopy, scanning electron microscopy, immunocytochemical and RT‐PCR were used for checking the change of the induced cells. Morphology was mostly spindle; a small part was of a polygon. The undifferentiated SMSCs showed the fibroblast‐like morphology; however, most of the differentiated cells were in the shape of a spindle and the rest were polygonal. Furthermore, being induced by bFGF, SMSCs can be found to be a unique long extension from the cell body under the scanning electron microscope. RT‐PCR and immunocytochemical analysis was made to confirm nestin (neural stem cell marker) and NF‐L (neurofilament‐light or neurofilament 68‐kDa mature nerve cell marker) expression in SMSCs. SMSCs can differentiate into neuronal cells when induced by bFGF. The bFGF‐induced SMSCs not only changed into neural‐like cells but also expressed specific markers.     

Comparative assessment of articular cartilage and synovial membrane in experimental haemarthrosis

The changes in articular cartilage and synovial membrane of the knee joints were studied in two groups of rabbits and Wistar rats with experimental haemarthrosis, electron microscopically. Hamarthrosis was produced in group 1 by a single autologous blood injection, in group 2 by intraarticular fracture of the femoral condyles. Samples were taken from the intact articular cartilage, the menisci and the infrapatellar portion of the synovial membrane 12 h to 20 days after intervention. Blood resorption occurs only in the synovial membrane. Fragmentation of erythrocytes and erythrocytophagy by synovial macrophages is documented. The different stages of intracellular digestion of erythrocyte fragments are traced down. Synovial fibroblasts do not participate in erythrocytophagy, although they disclose morphological signs of enhanced functional activity. The findings show changes in the matrix and chondrocytes within the articular cartilage and menisci, and presence of free erythrocytes and lipoprotein complexes amidst the collagen fibres of the matrix. The chondrocytes are poor in cell organelles, while the intracytoplasmic filaments, lipid droplets and glycogen granules are augmented in number. There is no evidence of erythrocytophagy by cartilage cells. On single blood injection in the joint, the ensuing changes are reversible, and the normal synovial membrane structure is restored much quicker than the articular cartilage.     

Quantitative analysis of chondroitin sulphate retention by tannic acid during preparation of specimens for electron microscopy

Summary The ability of tannic acid to enhance binding of glycosaminoglycans to purified collagen was analysed in an in vitro system using amino sugar analysis on an amino acid analyser, transmission electron microscopy, and scanning electron microscopy. Collagen was purified by digestion with trypsin, papain, and hyaluronidase. Purified collagen was incubated with hyaluronic acid or with chondroitin sulphate glycosaminoglycan and then treated with tannic acid. Tannic acid was found to enhance retention during preparation for electron microscopy of either of the glycosaminoglycans onto collagen fibres. The ability of tannic acid to enhance binding of collagen and glycosaminoglycans might explain, at least in part, its structural reinforcement effect on resected synovial joint-apposing surfaces during preparation for scanning electron microscopy.     

Gap junctions in human synovial cells and tissue

Our objective was to establish the existence of intercellular communication through gap junctions in synovial lining cells and in primary and passaged cultures of human synovial cells. Communication between cells was assessed using the nystatin perforated-patch method, fluorescent dye transfer, immunochemistry, transmission electron microscopy, and immunoblotting. Functional gap junctions were observed in primary and passaged cultures and were based on measurements of the transient current response to a step voltage. The average resistance between cells in small aggregates was 300 +/- 150 MOmega. Gap junctions were also observed between synovial lining cells in tissue explants; the size of the cell network in synovial tissue was estimated to be greater than 40 cells. Intercellular communication between cultured cells and between synovial lining cells was confirmed by dye injection. Punctate fluorescent regions were seen along intercellular contacts between cultured cells and in synovial membranes in cells and tissue immunostained for connexin43. The presence of the protein was verified in immunoblots. Regular 2-nm intermembrane gap separations characteristic of gap junctions were seen in transmission electron micrographs of synovial biopsies. The results showed that formation of gap-junction channels capable of mediating ionic and molecular communication was a regular feature of synovial cells, both in tissue and in cultured cells. The gap junctions contained connexin43 protein and perhaps other proteins. The physiological purpose of gap junctions in synovial cells is unknown, but it is reasonable to anticipate that intercellular communication serves some presently unrecognized function.     

Light and Electron Microscopic Investigation of Equine Synovial Membrane

Light and electron microscopic examination was made on equine synovial membrane from 23 healthy joints, nine joints with synovitis caused by intraarticular fracture and 10 joints with synovitis caused by osteochondrosis dissecans. Histologically as well as ultrastructurally the equine synovial membrane from healthy joints was of principally the same character as described in other species. Three types of synovial membrane — areolar, fibrous and adipose — and two types of lining cell were distinguished histologically. Ultrastructurally three types of lining cells were distinguished: A and Β type and an intermediate cell type. In healthy joints they were loosely arranged, parallel to the joint surface in an intercellular matrix, which was in direct continuity with the joint space. In joints with intraarticular fracture there was mild inflammation of the synovial membrane. There was elongation and hyperplasia of the lining cells with a relative increase in type A cells. The cell surface of lining cells was increased through filopodia. There was also an increase in cytoplasmic organelles i.e. hyperplasia of rough endoplasmic reticulum and Golgi complexes in Β type cells and an increase in lysosomes, and increased numbers of vesicles of varying types in A cells. In joints with osteochondrosis dissecans the lining cell hyperplasia and the inflammation in the synovial membrane were more prominent. Ultrastructurally the same alterations as in the previous group were seen including a relative increase in the number of A cells but degenerative changes were common in the lining cells. These changes were dilatation and vesiculation of rough endoplasmic reticulum, mitochondrial condensation, dilatation of the nuclear envelope and loss of plasma membranes, leading to disintegration of cells.     

Influence of synovial cells on cartilage in vitro: induction of breakdown and inhibition of synthesis.

Organoid or high density cultures of: (1) synovial cells from patients with rheumatoid arthritis, and (2) prechondrogenic mesenchymal cells from limb buds of 12-day-old mouse embryos, were co-cultured for 7-10 days using the Trowell culture system. Depending on the time of commencing co-cultivation, chondrogenesis was inhibited (co-cultivation from the start) or the cartilaginous matrix was partly degraded (co-cultivation after formation of embryonic cartilage, i.e. on day 4). These effects were obtained with cells from synovial fluid as well as from synovial tissue. Matrix degradation and the behaviour of the different cell types could be demonstrated well by electron microscopy under the in vitro conditions applied.     

运用偏振光观测聚乙烯磨屑颗粒诱导人工关节假体无菌性松动

目的:探讨运用偏振光显微镜来观测无菌性松动人工关节假体周围的聚乙烯颗粒分布,评估其在研究磨屑颗粒诱导假体无菌性松动机制及防治措施等实验研究中的可行性。方法:我们用雌性新西兰大白兔建立动物模型,在左侧胫骨髓腔内植入羟基磷灰石(hydroxyapatite,HA)涂层假体。并分别于假体表面和膝关节腔内植入0.5×107超高分子量聚乙烯(Ultra-high molecular weight polyethylene,UHMWPE)颗粒。术后行四环素荧光双标记。膝关节滑膜组织苏木精-伊红(hematoxylin-eosin,HE)染色、骨组织改良丽春红染色后分别用普通光镜和偏振光镜观察,未染色的骨组织行荧光显微镜和偏振光镜观察。结果:在聚乙烯颗粒刺激下,膝关节滑膜组织增生明显,骨-假体结合差,假体周围骨小梁稀疏,偏振光显微镜可清晰显示双折光性的聚乙烯颗粒在膝关节分布于滑膜及其深层结缔组织中,在骨-假体间隙间大量充填,阻碍骨-假体整合。结论:运用偏振光显微镜可以清晰而简便地观察滑膜和假体周围的聚乙烯颗粒分布,与传统实验方法相比,更加直观、简便和经济。     

A fine structural study of the development of the chick flexor digital tendon: a model for synovial sheathed tendon healing.

The development of the synovial sheathed flexor digital tendon in the chick was studied by light and electron microscopy in 12-day embryos to 22-day post-hatched chickens. Areas of specialized connective tissue differentiation were identified in this complex structure consisting of a lubricated synovial sheath, elastic vincula and fibrocartilaginous adaptations on the surface of the tendon. The presence of some of these specialized adaptations may be related to the specific types of mechanical forces and stresses applied to the developing connective tissue system. This model system appears to be appropriate for the experimental study of tendon injuries related to the human hand.     

Nanoscale topography of the basement membrane underlying the corneal epithelium of the rhesus macaque

This paper quantitatively defines the nanoscale topography of the basement membrane underlying the anterior corneal epithelium of the macaque. Excised corneal buttons from macaques were placed in 2.5 mM ethylenediaminetetraacetate (EDTA) for 2.5 h, after which the epithelium was carefully removed to expose the underlying basement membrane. The integrity of the remaining basement membrane was verified using fluorescent microscopy in conjunction with antibody staining directed against laminin and collagen type IV as well as transmission electron microscopy. Characterization of the surface of the basement membrane was performed using transmission electron microscopy, high-resolution, low-voltage scanning electron microscopy, and atomic force microscopy. Quantitative data were obtained with all three imaging techniques and compared. The basement membrane has a complex topography consisting of tightly cross-linked fibers intermingled with pores. The mean elevation of features measured by transmission electron microscopy, scanning electron microscopy, and atomic force microscopy was 149 +/- 60 nm, 191 +/- 72 nm, and 147 +/- 73 nm, respectively. Mean fiber diameter as measured by SEM was 77 +/- 44 nm and pore diameter was 72 +/- 40 nm, with pores occupying approximately 15% of the total surface area. Similar feature types and dimensions were also found for Matrigel, a commercially available basement membrane-like complex, supporting that a minimum of artifact was introduced by corneal preparative procedures to remove the overlying epithelium. Topographic features amplified the surface area over which cell-substratum interactions occur by an estimated 400%. The three-dimensional structure of the basement membrane exhibits a rich complex topography of individual features, consisting of pores and fibers with dimensions ranging from 30 to 400 nm. These nanoscale substratum features may modulate fundamental cell behaviors such as adhesion, migration, proliferation, and differentiation.     

Activities of dipeptidyl peptidase II, dipeptidyl peptidase IV, prolyl endopeptidase, and collagenase-like peptidase in synovial membrane from patients with rheumatoid arthritis and osteoarthritis.

We examined the activities of peptidases in the synovial membrane from patients with rheumatoid arthritis (RA) and osteoarthritis (OA). Dipeptidyl peptidase II (DPP II), prolyl endopeptidase (PEP), and collagenase-like peptidase (CLP) activities were higher in knee joint synovial membrane from patients with RA than in that from patients with OA. DPP II and PEP activities in knee joint synovial membrane of patients with RA increased in parallel with the increase in joint fluid volume, whereas DPP IV activity decreased in parallel with the increase in joint fluid volume. These results suggest that these peptidases in the synovial membrane may play some role in immunological disturbances in the joints of patients with RA. Measurement of these peptidases in synovial membrane may be useful in the diagnosis of the severity of local joint inflammation.     

Combined immunofluorescence and field emission scanning electron microscope study of plasma membrane-associated organelles in highly vacuolated suspensor cells of white spruce somatic embryos

Highly vacuolated suspensor cells of spruce somatic embryos were examined by immunofluorescence light microscopy using butyl-methyl-methacrylate (BMM) and polyethylene glycol (PEG) embedded sections, transmission electron microscopy (TEM) and field emission scanning electron microscopy (FESEM). The use of PEG embedded embryos provided a rapid method for light microscope detection of antigens before committing to FESEM analysis. BMM embedded specimens provided well preserved suspensor cells for immunofluorescence. FESEM permitted high resolution observation of large areas of the inner surface of the plasma membrane and associated cell organelles. Suspensor cells contained mostly transversely oriented cortical microtubules linked to the plasma membrane and adjacent microtubules by cross- bridges. Light and electron microscopy revealed numerous clathrin coated structures on the plasma membrane. These included flat patches of clathrin, coated pits and coated vesicles. Many coated vesicles were associated with microtubules. Both tubular and lamellar endoplasmic reticulum were observed on the plasma membrane by FESEM.     

Loss of endoplasmic reticulum membrane integrity: an image analysis of the glucose-6-phosphatase system in human hepatocyte.

Histochemical and cytochemical methods induce a loss of endoplasmic reticulum (ER) membrane integrity in hepatocytes. In order to evaluate the degree of ER membrane integrity, glucose-6-phosphatase (G6P-A) was localized in light and electron microscopy using glucose-6-phosphate (G6P) and mannose-6-phosphate (M6P) as substrates. In case of ER membrane alteration, M6P diffuses inside the ER and is hydrolysed by a non-specific phosphohydrolase. G6P and M6P hydrolysis was quantified with image analysis methods. In light microscopy, the ratio of reaction of M6P hydrolysis/G6P hydrolysis gave 75% of non specific reaction. In electron microscopic study this ratio was about 30%. These results showed that enzyme localization methods in electron microscopy produced less ER membrane alteration than light microscopic methods.     

gamma-Aminobutyric acid in synovial membrane of rat knee joint

gamma-Aminobutyric acid (GABA) content was measured, and the release of GABA was studied in the synovial membrane of the rat knee joint. GABA content of the synovial membrane was 20.1 nmol/g tissue. Ten days after unilateral dissection of the sciatic nerve, femoral nerve or both nerves, the GABA contents of the ipsilateral membrane were 13.8, 14.6 and 7.8 nmol/g tissue, respectively. High K+ evoked the Ca2+-dependent release of [3H] GABA from the synovial membranes of intact rats preloaded with [3H] GABA, but did not evoke release from the membrane ipsilateral to the dissection of both sciatic and femoral nerves. Evoked release of [3H] GABA was obtained in the synovial membrane preloaded with [3H] GABA in the presence of beta-alanine, but not in the presence of 2,4-L-diaminobutyric acid. These results indicate that GABA is present in the neuronal elements of the synovial membrane of the rat knee joint.     

Silicone rubber membrane as a support for long-term cultivation and electron microscopic processing of nervous tissue

Silicone rubber membrane can be successfully used as a support for cultivation of nervous tissue; its processing for electron microscopy is described: The advantages of silicone rubber membrane as support, compared with glass, are. its biological inertness, the ability to withstand dry-heat sterilization at 160°C, transparency, easy processing for electron microscopy. Spatial configuration of cells and explants in specimens is preserved.     

Membrane tubules attach Salmonella Typhimurium to eukaryotic cells and bacteria

Using scanning electron microscopy techniques we measured the diameter of adhesive tubular appendages of Salmonella enterica serovar S. Typhimurium. The appendages interconnected bacteria in biofilms grown on gallstones or coverslips, or attached bacteria to host cells (human neutrophils). The tubular appendage diameter of bacteria of virulent flagellated C53 strain varied between 60 and 70 nm, thus considerably exceeding in size of flagella or pili. Nonflagellated bacteria of mutant SJW 880 strain in biofilms grown on gallstones or coverslips were also interconnected by 60-90-nm tubular appendages. Transmission electron microscopy studies of thin sections of S. Typhimurium biofilms grown on agar or coverslips revealed numerous fragments of membrane tubular and vesicular structures between bacteria of both flagellated and nonflagellated strains. The membrane structures had the same diameter as tubular appendages observed by scanning electron microscopy, indicating that tubular appendages might represent membrane tubules (tethers). Previously, we have shown that neutrophils can contact cells and bacteria over distance via membrane tubulovesicular extensions (TVE) (cytonemes). The present electron microscopy study revealed the similarities in size and behavior of bacterial tubular appendages and neutrophil TVE. Our data support the hypothesis that bacteria establish long-range adhesive interactions via membrane tubules.     

Visualization of the structures of the hepatitis C virus replication complex

Hepatitis C viral RNA synthesis has been demonstrated to occur on a lipid raft membrane structure. Lipid raft membrane fraction purified by membrane flotation analysis was observed using transmission electron microscopy and atomic force microscopy. Particles around 0.7 um in size were found in lipid raft membrane fraction purified from hepatitis C virus (HCV) replicon but not their parental HuH7 cells. HCV NS5A protein was associated with these specialized particles. After several cycles of freezing-thawing, these particles would fuse into larger sizes up to 10 um. Knockdown of seven proteins associated with lipid raft (VAPA, COPG, RAB18, COMT, CDC42, DPP4, and KDELR2) of HCV replicon cells reduced the observed number of these particles and suppressed the HCV replication. Results in this study indicated that HCV replication complexes with associated lipid raft membrane form distinct particle structures of around 0.7 um as observed from transmission electron microscopy and atomic force microscopy.