H. P. Dinges

Fresh autogeneic, frozen allogeneic, and decalcified allogeneic bone grafts in dogs

In fully-grown mongrel dogs, diaphyseal ulnar defects 25 mm long were stabilised by screws and plates, and were temporarily filled with silicone rubber blocks. After eight weeks the block was replaced either by fresh autogeneic cancellous bone, allogeneic deep-frozen cancellous bone, allogeneic decalcified bone matrix, or bone matrix gelatin. After 24 weeks the implants were evaluated by radiography, histology, and measurements of new bone volume, using computer-assisted density registration on microradiographs. Only the autogeneic bone grafts led to healing in all instances. Bone regeneration in the other groups was not significantly better than in the sham group in which no graft was employed. Decalcified bone matrix proved ineffective.

Immunohistochemical detection of tumor necrosis factor-alpha, other cytokines and adhesion molecules in human livers with alcoholic hepatitis

This immunohistochemical study was designed to investigate the possible contribution to and topographical distribution of some important cytokines, such as tumour necrosis factor α (TNFα) and interleukins, in acute alcoholic hepatitis. The well-known inductive capacity of these cytokines with respect to the expression and/or up-regulation of adhesion molecules, such as intercellular adhesion molecule-1 (ICAM-1) and endothelial leukocyte adhesion molecule-1 (ELAM-1), was a further point to be studied. Moreover, the proposed induction of adhesion molecules might also be associated with the activation and attraction of a special population of inflammatory cells characteristic for alcoholic hepatitis. Frozen liver samples from patients who died with signs of acute alcoholic hepatitis were evaluated using the alkaline phosphatase anti-alkaline phosphatase immunostaining technique and also single and double indirect immunofluorescence. In acute alcoholic hepatitis TNFα could be detected predominantly in ballooned hepatocytes, which often contained alcoholic hyalin (Mallory bodies). Moreover, TNFα showed a co-distribution with ICAM-1 expressed in the membranes of hepatocytes and with the occurrence of CD11b positive polymorphonuclear leukocytes (neutrophils) suggesting a possible major role of the β2-integrin Mac-1 as a ligand for ICAM-1. No induction of ELAM-1 could be found. In alcoholic hepatitis cytokines may be responsible for the induction of the adhesion molecule ICAM-1 on hepatocytic membranes and activate a defined population of inflammatory cells, thus contributing to the characteristic histological picture of acute alcoholic hepatitis with its concentration of neutrophils especially in areas with ballooned Mallory body-containing hepatocytes. Our results are in line with clinical findings showing high levels of TNFα and interleukin-1 in sera of patients with alcoholic hepatitis and with the already reported expression of ICAM-1 on hepatocytes.

Bacterial Translocation in a Baboon Model of Hypovolemic-Traumatic Shock

Bacterial translocation secondary to gut damage caused by traumatic shock is a source of posttraumatic sepsis. In several series, only one third of multiorgan failure patients with clinical manifestations of sepsis had an identifiable focus of infection (Goris 1985). The majority of ICU patients demonstrate the picture of clinical sepsis without a focus. In these patients, the gut frequently plays an important part as a shock organ involved in the subsequent development of sepsis.

Irradiation-sterilization of rat bone matrix gelatin

Bone matrix gelatin induces bone formation in muscle, and when implanted orthotopically it improves bone repair. Co-60 sterilization of bone gelatin impairs the protein-bound induction mechanisms. Gelatin samples nonirradiated or irradiated by 25 or 50 kGy were implanted into a pouch in the abdominal wall of Sprague-Dawley rats, as well as into a 7-mm calvarial defect. Evaluation was done by histologic studies, histomorphometry of orthotopic implants, and determination of alkaline phosphatase in ectopic implants. Gelatin irradiated with 50 kGy was absorbed in the muscle bed without evidence of any specific host reaction. Irradiation of 25 kGy led to histologically confirmed ectopic bone formation, but the wet weight of the explants was only half that of the nonirradiated control samples. Alkaline phosphatase activity was equal in both of these groups. With orthotopic implantation, neither a histologic nor a morphometric effect was seen with 25 kGy. Loss of osteoinduction with 25-kGy irradiation is apparently masked by osteoconductive mechanisms with orthotopic implantation.

Co-expression of cytokeratin and vimentin filaments in rete testis and epididymis : an immunohistochemical study

In 11 testes of different developmental stages (from 10-week-old embryos to adult) the cytokeratin and vimentin expression patterns of rete testis and epididymis were investigated immunohistochemically in formaldehyde-fixed paraffin-embedded material. In addition, immunofluorescence microscopy including double immunofluorescence was performed on frozen sections of 3 of these 11 cases. Rete testis and epididymis cells displayed a heterogeneous co-expression of cytokeratin and vimentin. In double immunohistochemistry, differences in distribution of keratin and vimentin intermediate filaments with predominance of cytokeratins in the apical cytoplasmic regions and of vimentin filaments in the basal portions of the cells were found. Cytokeratin expression preceded the appearance of vimentin: cytokeratin was already detectable in 10-week-old embryos, while weak vimentin immunoreactivity was first seen in 12-week-old embryos and became conspicuous in testes around the perinatal period. In testes of children up to 2 years of age the cytoplasmic distribution of cytokeratin and vimentin was more homogeneous. Predominance of the basal cell portions for vimentin and the apical regions for cytokeratin staining were less pronounced than in adult testes. In the proximal and distal parts of the epididymis a different intermediate filament expression pattern was found with a clear predominance of cytokeratin near the rete.

Activation/Adherence Phenomena of Leukocytes and Endothelial Cells in Trauma and Sepsis

Leukocytes, in particular polymorphonuclear neutrophils (PMNs), release important mediators of microvascular injury seen after ischemia/reperfusion, hemorrhage, and sepsis. Adherence of PMNs to the endothelium is thought to be crucial in PMN-mediated injury and results in an increase in permeability and edema. Adhesion is also an in tegral part of the beneficial role of leukocytes during migration, phagocytosis and bacterial killing.

The influence of fibrin sealant on demineralized bone matrix-dependent osteoinduction. A quantitative and qualitative study in rats.

Allogeneic demineralized bone matrix (DBM) and bone matrix gelatin (BMG) were implanted with or without fibrin sealant (FS) ectopically (abdominal wall) and orthotopically (7-mm trepanation defect) in 38 male Sprague-Dawley rats. Evaluation was done by descriptive histology, histomorphometry of orthotopic implants, and determination of alkaline phosphatase in ectopic implants. The observation period was 21 days with ectopic implantation and 26 days with orthotopic implantation. In all ectopic specimens, new bone developed without any qualitative difference between specimens with and without FS. The alkaline phosphatase activity did not change significantly upon addition of FS. Morphometry revealed slight differences between the groups with and without FS. The peripheral bone deposits in the BMG + FS group, was significantly larger than in the BMG group. These investigations demonstrated neither a clearly positive nor negative effect of FS on ectopic osteoinduction or BMG-dependent osteoregeneration.

Dog bone less osteogenetic than rat bone Bone-matrix transplants in nude rats

Demineralized bone matrix and bone-matrix gelatin prepared from cortical rat bone, and from cortical and cancellous canine bone were implanted into muscle pouches of nude rats for 6 weeks. Evaluation was done by histology, histomorphometry, and determination of alkaline phosphatase. Rat matrix consistently induced new bone and high phosphatase levels. Canine matrix induced but small amounts of bone and lower phosphatase levels, with cortical matrix somewhat more inductive than cancellous matrix; demineralized cancellous bone matrix from the dog was the only material tested not showing any inductivity. Irrespective of bone type or species, gelatin had clearly higher induction capacity than demineralized bone matrix.

Experimental osteoinduction in rats: Collagen-apatite versus osteogenin-containing gelatine

SummaryAn experimental study in rats was done to investigate the bone-regenerating properties of collagen apatite (Collapat) and to compare it with osteoinduction dependent on osteogenin-containing gelatine (OCG). The test substances were implanted orthotopically (calvarial defect — 7 mm in diameter) and heterotopically (paravertebral muscles, abdominal muscles). The results were evaluated histologically and enzymatically (alkaline phosphatase). Collapat caused neither osteoinduction in the heterotopic site nor healing of the bone defects. Foreign body reaction without new bone formation was encountered. OCG implantation leads to new bone formation in the muscles within 3 weeks, associated with a significant increase in alkaline phosphatase activity, and to extensive new bone formation in the calvarial defect within 4 weeks. The defects did not heal if left empty. The value of clinical application of Collapat appears to be doubtful. Osteoinduction with OCG requires further experimental investigation.ZusammenfassungIn einer experimentellen Untersuchung an der Ratte wurden die osteoregenerativen Eigenschaften von Collagen-Apatit (Collapat) geprüft und mit der Osteogenin-Containing-Gelatine abhängigen Osteoinduktion verglichen. Die Prüfsubstanzen wurden orthotop (7 mm ∅ Trepanationsdefekt) und heterotop (paravertebrale Muskulatur, Bauchwand) implantiert. Die Auswertung erfolgte deskriptiv histologisch und enzymatisch quantitativ (alkalische Phosphatase). Collapat bewirkte keine Osteoinduktion im Weichteillager und führte nicht zur Knochendefektheilung, sondern zeigte in allen Fällen ein Fremdköorpergranulationsgewebe ohne Osteogenese. OCG führt zur Knochenneubildung in der Muskulatur innerhalb von drei Wochen, verbunden mit einem signifikanten Anstieg der alkalischen Phosphatase, und zur ausgedehnten Knochenbildung im Trepanationsdefekt innerhalb vier Wochen. Diese Defekte heilten in keinem Fall, wenn keine Substanz implantiert wurde. Der Wert der klinischen Applikation von Collapat erscheint deshalb zweifelhaft; die Osteoinduktion mittels OCG bedarf weiterer experimenteller Untersuchungen.

Cryopreservation of cytological specimens for immunocytochemistry.

A method has been established for storage and preservation of cytological specimens in liquid nitrogen and further processing for immunocytochemistry as smears prepared from thawed cells or cryo-sections of frozen cell pellets. For the experiments cultured cells of a T-lymphoblastic leukemia cell line (ATCC CCL 119) and blood cells of the buffy coat of healthy humans were treated with a cryo-solution (fetal calf serum +5% dimethylsulfoxid) and after freezing stored in liquid nitrogen. Alternatively, cells preincubated with cryo-solution followed by suspension in fetal calf serum without cryo-additive were frozen and stored in liquid nitrogen for the production of cryo-sections. Indirect immunofluorescence and alkaline phosphatase--antialkaline phosphatase based immunoreactions were performed for the decoration of various surface antigens with a panel of monoclonal antibodies. All immunoreactions were repeated at least three times and the stored cell preparations were investigated after different periods of storage (up to four months). The immunoreactions of fresh cells in suspension (which were used as controls) were comparable with those of cryopreserved cells, e.g. cells on smears after thawing and on cryo-sections of cell pellets. The strongest immunoreactions were achieved on fixed cryo-sections. The maintenance of cell morphology of smears from cryopreserved cells was slightly better than of cells from cryo-sections. In our hands the preparation of cell pellets, which are suitable for the storage in liquid nitrogen and the production of cryosections, is a very useful method for immunocytochemical investigations of cytological specimens especially in situations where immunoreactions cannot be performed on fresh material.(ABSTRACT TRUNCATED AT 250 WORDS)