Eckart Stofft

Cellular ultrastructure of the ruptured anterior cruciate ligament. A transmission electron microscopic and immunohistochemical study in 55 cases.

To evaluate the cellular ultrastructure following injury, we examined the anterior cruciate ligaments in 55 patients with complete tears in different phases after the injury and compared them to a control group of 39 cadaver knees. Samples were analyzed by electron microscopy, immunofluorescence, and ultramorphometry. After an invasion of inflammatory cells into the stumps of the ruptured ligaments, a marked proliferation of fibroblasts was found at the end of Phase 1 (2-3 days after the ligament injury), that was even more pronounced at the beginning of Phase II (4-17 days). These cells were initially highly metabolically active and secreted Type III collagen precursors. In Phase III (4-45 days), fibroblast degeneration occurred with increasing frequency. Furthermore, some fibroblasts showed signs of cell death. Our findings suggest that the structural alterations of the intraligamentous fibroblasts diminish their function and, consecutively, disorganization of the developing repair tissue occurs. This mechanism might contribute to the poor healing potential of the ruptured anterior cruciate ligament.

Structure and Function of Matrix Components in the Cruciate Ligaments

In the present study, the matrix components of 100 cruciate ligaments were analyzed by conventional electron microscopy, immunohistology, morphometry, and immunoelectron microscopy. The anterior (ACL) and the posterior (PCL) cruciate ligaments contained collagen types III, IV, and VI. Several structural glycoproteins, like fibronectin, laminin, entactin, tenascin, and undulin were detected using monoclonal antibodies. Whereas laminin and entactin were higher concentrated in the PCL, type VI collagen was more frequently found in the ACL. The ACL had a critical nourishment in its distal and middle thirds. In all ligament parts the PCL revealed a better vascular supply with strong correlation to type IV collagen expression. The normal matrix of the cruciate ligaments represented a complicated regulatory network of proteins, glycoproteins, elastic systems, and glycosaminoglycans with multiple functional interactions.

Collagen ultrastructure in ruptured cruciate ligaments : an electron microscopic investigation

The ultrastructure of collagen fibrils was investigated in normal (n 39) and ruptured (n 23) human anterior cruciate ligaments. The normal ligament had a complex three-dimensional structure. Collagen fibrils predominantly had a undirectional course with parallel arrangement and a mean diameter of 75 (20-185) nm. Four days after anterior cruciate ligament rupture, the mean fibril diameter was increased; it later decreased, probably due to synthesis of young, thin 30-40 nm fibrils. Interfibrillar dysplastic collagen fibrils were detected in the extracellular matrix of ruptured ligaments. They were more frequently found later than 3 days after rupture and were seen also at a distance of 2-3 cm from the rupture zone. The presence of dysplastic fibrils may explain the functional insufficiency of the repair tissue in ruptured cruciate ligaments.

Einwachsverhalten von periostgedeckten Knochendübeln mit und ohne autologe Knorpelzellen

ZusammenfassungDie Transplantation osteochondraler Dübel hat den Nachteil neuer Hebedefekte in gesunden Gelenkregionen.In diesem Tierexperiment sollte überprüft werden,ob die Transplantation von autologen periostgedeckten Knochendübeln zu einer adäquaten Knochen und Knorpelreparation führt und ob die Injektion autologer Knorpelzellen in das Periost eine zusätzliche Verbesserung darstellt.22 Minipigs wurden 2-mal bilateral operiert.Eine 1.Operation diente der Knorpelbiopsie im Kniegelenk zur Chondrozytenkultivierung. In einer 2.Operation wurde ein osteochondraler Defekt in der medialen Trochlea gesetzt und mit einem kortikospongiösen Knochenzylinder aus der Tibia unterschiedlich therapiert.Gruppe A: Spontanheilung (Kontrolle),Gruppe B: Knochenzylinder, Gruppe C: Zylinder mit anhaftendem Periost, Gruppe D: Zylinder mit Periost und darin injizierte autologe Chondrozyten. Die Tiere wurden nach 6, 12, 26 und 52Wochen getötet. Die Präparate wurden makroskopisch beurteilt, biomechanisch getestet (Langzeitpräparate; Indentationstest) und anhand eines histomorphologischen Scores verblindet begutachtet.Periostgedeckte Knochendübel führten zu einer guten Reparation des Knochen- und Knorpelgewebes.Die gebildeten Regenerate der Gruppe C und D bestanden überwiegend aus Faserknorpel, basal mit hyalinartigen Gewebeinseln und waren randständig in den Umgebungsknorpel integriert. Die zusätzliche Knorpelzellinjektion erbrachte keine verbesserte Regeneratbildung.Der bio-mechanisch Test zeigte eine deutliche Überlegenheit der Therapiegruppen C und D.Aufgrund unserer Resultate stellt die alleinige Transplantation periostgedeckter Knochendübel eine echte Alternative zur primären Therapie chondraler und osteochondraler Defekte dar und bietet eine gute Möglichkeit zur Hebedefektfüllung.Der zusätzliche Einsatz autologer Chondrozyten scheint nicht gerechtfertigt.AbstractAutologous osteochondral transplantation has the major disadvantage of significant damage to a healthy joint surface at the donor site.The purpose of this study was to examine the effect of autogenous chondrocytes injected into the periosteum of autologous bone grafts in order to provide an alternative method for cartilagerepair.A total of 22 Göttinger minipigs were operated twice on both knees.The first operation served for cartilage biopsy for the chondrocyte culture.During the second operation an osteochondral defect was created in the medial facet of the trochlear groove.The defect was treated differently with an autologous cortico-cancellous bone cylinder,harvested from the proximal tibia.Group A: untreated defect (control);B: bone-graft;C: bone-graft covered with periosteum; D: bone-graft with periosteum and injected autologous chondrocytes.The animals were killed after 6,12,26 and 52weeks.The regenerated areas were evaluated macroscopically, tested biomechanically (long-term specimens; indentation-test) and a histological, blind evaluation was carried out according to a semi-quantitative scoring system.The periosteum covered bone cylinders in Groups C and D showed good repair of the bone and cartilage defect.The repaired tissue consisted predominantly of fibrocartilage with the partial formation of hyalin like tissue.The regenerated areas were integrated with the adjacent cartilage and were biomechanically superior when compared with the other groups. The additional injection of chondrocytes did not produce significantly better results.Our findings suggest that the transplantation of periosteum-covered bone cylinders may provide an alternative method for treating chondral and osteochondral defects and can be recommended for filling large donor site defects in joint surgery.The additional transplantation of chondrocytes does not seem to be justified.

Vitamin A and ciliated cells I. Respiratory epithelia

SummaryTo estimate the role of vitamin A on ciliated cells we investigated whether ciliated cells undergo any alteration during vitamin A deficiency. The epithelia examined include the ciliated cells of the respiratory tract and the ciliated sensory cells of the inner ear, the tongue, and the olfactory cells. This part of the paper will describe the ciliated epithelium of the tracheobronchial tract and its relation to vitamin A status. During vitamin A deficiency a partial loss of ciliae can be observed before any squamous metaplasia (which usually occurs during longer lasting vitamin A deficiency) develops. The scanning electron microscopic data illustrate the altered surface of the epithelium during vitamin A deficiency better than transmission electron microscopy.ZusammenfassungUm zu prüfen, in welcher Weise Vitamin A zilientragende Epithelien beeinflußt, wurde untersucht, ob zilientragende Epithelien im Vitamin-A-Mangel besondere Veränderungen aufweisen. Die untersuchten Gewebe beinhalten das Respirationsepithel sowie die zilientragenden Sinnesepithelien des Innenohres, der Zunge und des Riechepithels. Dieser Teil beschreibt die Veränderungen des zilientragenden Epithels des Tracheobronchialtraktes in Relation zum Vitamin-A-Status. Mit rasterelektronenmikroskopischen Methoden konnte im Vitamin-A-Mangel ein fleckiger Verlust von Zilien des Respirationsepithels beobachtet werden, bevor es zu einer squamösen Metaplasie des Gewebes — wie sie im Vitamin-A-Mangel beschrieben ist — kam. Die rasterelektronenmikroskopischen Aufnahmen stellen die Veränderungen an den zilientragenden Zellen dieses Epithels anschaulicher dar, als dies mit transmissionselektronenmikroskopischen Verfahren möglich ist.

Characterization of glycoconjugate expression during development of Meckel's cartilage in the rat

The staining patterns of 24 biotinylated lectins were analyzed in serial sections of the mandible of 13- to 21-day-old rat embryos by means of the avidinbiotin-peroxidase method. A ubiquitous distribution of binding sites was demonstrated after incubation with Con A (Canavalia ensiformis), DSL (Datura stramonium; except bone matrix), and WGA (Triticum vulgare). ECL (Erythrina cristagalli), GSL I (Griffonia simplicifolia), SJA (Saphora japonica), VVL (Vicia villosa), DBA (Dolichus biflorus), UEA I (Ulex europeus), and LTA (Lotus tetragonobolus) were constantly negative. In early stages of development, GSL II (Griffonia simplicifolia II) was a selective marker of prechondral blastema. In contrast, PNA (Arachis hypogaea) did not stain condensing mesenchyme. During chondrogenesis of Meckelss cartilage a general decrease of lectin binding was observed. Mature cartilage matrix was constantly negative. Chondrocytes were marked by the lectins PSA (Pisum sativum), WGA, PHA-E, and PHA-L (Phaseolus vulgaris E and L). A strong GSL II binding was restricted to the mesial-superior region of the perichondrium. In later stages, several lectins revealed significant differences between preskeletal (“central”) areas and the remaining (“peripheral”) mesenchyme. A clear binding reaction was noted in central regions by applying LEA (Lycopersicon esculentum) and STL (Solanum tuberosum), while the peripheral tissue was only faintly stained. Developing bone was specifically marked by succinylated WGA (sWGA). The lectins LCA (Lens culinarus) and RCA (Ricinus communis) bound to fibers and extracellular matrix of the connective tissue. Jacalin (Artocarpus integrifolia) and SBA (Glycine max) binding sites were found in macrophages. Affinity of VAA (Viscum album) increased parallel with maturation of endothelial cells. Specific lectin-binding patterns revealed no correlation with the distribution of glycosaminoglycans. The results demonstrate a general reduction of oligosaccharide structures during development of Meckels cartilage. From our observations we conclude that intralaminar glucose and/or mannose sequences as well as terminal sialic acid molecules are ubiquitously distributed, while terminal α-fucose was constantly negative. Lectin-binding patterns of macrophages may reflect the presence of specifically linked terminal galactose. Our findings indicate that oligosaccharides terminating in N-acetylglucosamine are bone-specific. The significance of the restricted staining of the perichondrium by GSL II remains to be elucidated.

Rasterelektronenmikroskopische Untersuchung des hyalinen Gelenkknorpels

Guinea pigs (pirbright white) were subjected to fixed mechanical stress in an endless-belt experiment. Shoulder, elbow, hip, and knee joints were taken and the articular cartilage was prepared for exa

Development and morphology of rat synovial membrane.

The development of the knee joint and synovial membrane in Wistar rats was studied from the 13th fetal day until adulthood by means of light microscopy, transmission electron microscopy and scanning electron microscopy. Using acid phosphatase and peroxidase methods, the different cell types of synovial membrane could be identified.

Correlative histologic and arthroscopic evaluation in rheumatoid knee joints

SummaryThe correlation between arthroscopic observations and histologic changes in rheumatoid arthritis is still controversial. Synovial samples of 21 knee joints in rheumatoid arthritis patients were comparatively investigated by endoscopy and histology. Biopsies were scored by an endoscopist and subsequently dissected. Different histochemical and immunocytochemical staining techniques were used to define inflammatory activity. Arthroscopic and histological values were compared by rating scales and variance analysis. Our study indicates that synovial biopsy is of diagnostic value in rheumatoid arthritis. However, its usefulness depends on the histochemical methods used. The results revealed highly significant correlations of endoscopic features with the number of neutrophilic granulocytes, intravascular leukocytes, and peroxidase-positive macrophages. However, no relationship was found between the detection of lymphocytes or resident macrophages and inflammatory scores. The close correlation between endoscopic and histological findings suggests that arthroscopic evaluation allows a valuable classification of the inflammatory activity in rheumatoid synovitis.

Distribution patterns in glycoconjugate expression during the development of the rat palate.

SummaryThe distribution of complex carbohydrate structures during the embryonic development of the rat palate was analysed by examining lectin-binding patterns in serial paraffin and cryostat sections. With few exceptions, the binding patterns showed a general increase in lectin receptors in the more developed stages of palatogenesis. High mannose oligosaccharides were especially amplified during development. Terminal fucose molecules were not expressed. In contrast, terminal sialic acid molecules were ubiquitously distributed in epithelial and mesenchymal tissues. Non-sialylated terminal N-acetylglucosamine was specifically restricted to evolving bone matrix. Before palatal fusion, quantitative but not qualitative differences were detected between oral, nasal, and medial-edge epithelial surfaces. The only exception was LCA, which specifically marked epithelial cells at the tip of palatal shelves. A very selective affinity for Jacalin was demonstrated in the oral epithelium of the palate after day 16, suggesting the presence of sialylated terminal galactose-(β-1,3)-N-acetylgalactosamine. PNA specifically marked the basal lamina of the oral side of palatal processes. The binding patterns of DBA, GSL IA, SBA, and VVA indicated that the epithelium of the tongue is characterized by terminal α- and β-galactose residues, whereas palatine cells possess only molecules with β-anomery. During palatogenesis, glycosaminoglycans patterns were significantly modified. Our data suggest that alteration of complex carbohydrate structures may play a central role in modulating cell-cell and cell-matrix interactions. The significance of these findings, however, remains to be elucidated.