Degenerative changes in the temporomandibular joint after diagnostic arthroscopy. An experimental study in goats.

Arthroscopic procedures for diagnosis and treatment of temporomandibular joint (TMJ) diseases are widely used. Few adverse effects of the procedures have been reported. In this in vivo study, diagnostic arthroscopy was performed in the left TMJ of seven castrated male goats, the right TMJ serving as a control. The goats were killed 1, 2, and 3 months after the arthroscopy. Macroscopic and microscopic examination revealed no apparent pathologic changes in two animals, minor changes in two, and severe degenerative changes in three. This study indicates that arthroscopic intervention in the TMJ may cause irreversible changes in the articular tissues.

Inhibited morphological terminal differentiation and enhanced proliferation of cultured mouse epidermal cells at different concentrations of dimethyl sulphoxide.

Dimethyl sulphoxide (DMSO), at concentrations of 1-2%, induces terminal differentiation in several different cell types in vitro and enhances the growth of newborn mouse epidermal cells in primary culture under conditions that also permit terminal differentiation. We have found that DMSO concentrations approaching 4% reversibly inhibited (with little overt toxicity) terminal differentiation of normal epidermal cells from newborn SENCAR mice. Cells cultured in medium containing 4% DMSO and calcium in excess of 1 mM did not stratify extensively or slough large amounts of keratinized debris into the medium as occurred in control cultures, nor did they form large numbers of squamous cells or keratin bundles, as revealed by light and electron microscopy. The number of detergent-insoluble cornified envelopes was similarly reduced. Long-term growth of epidermal colonies in secondary culture was optimum in 1% DMSO, this concentration also permitting normal terminal differentiation of these cells. Since DMSO had these effects on epidermal cells in vitro, it may also affect epidermal cell proliferation and terminal differentiation in vivo, an important consideration should DMSO ever be approved for topical use in the US.

Antibodies to mouse lung capillary endothelium.

We are interested in developing monoclonal antibodies (MoAbs) that recognize specific cell types in the lung of BALB/c mice. Normal mouse lung homogenate was used to immunize F344 rats and hybridomas were produced by fusion of rat spleen cells with mouse myeloma SP 2/0. Two hybridomas were selected which produced MoAbs active in immunohistochemistry of lung cells. MoAb 273-34A and 411-201B both show extensive peroxidase staining of capillary endothelial cells within alveolar walls of lungs at the light microscopic level. To demonstrate cell specificity, immunoelectron microscopy with gold-labeled antibody was performed. Lightly fixed lungs were frozen and thin-sectioned before staining with MoAb and 5-nm gold particles coupled to secondary antibody. Quantitative analyses of these cryosections show that both antibodies, used at optimal concentrations, are specific for binding to capillary endothelial cells. More than 95% of the gold particles are associated with capillary endothelial cells on the thin side of the alveolar wall. When capillaries adjoined thick septa containing interstitial cells, about two thirds of the gold particles were associated with endothelial cells and about one quarter with interstitial cells. These MoAbs should be useful in studying the role of endothelial cells in toxic lung injury.

Rat monoclonal antibodies to mouse lung components for analysis of fibrosis.

Six rat monoclonal antibodies to mouse lung membrane fraction have been characterized. Each has unique binding properties and can be used to stain particular lung components in paraffin sections. One antibody, 133-13A, recognizes a 100-kDa glycoprotein on lung tumor cells, but stains only macrophage-like cells in normal or fibrotic lung sections as determined by immunogold electron microscopy. The monoclonal antibody 273-34A binds to a 112-kDa protein on the surface of normal mouse lung fibroblasts. Immunogold electron microscopy demonstrates antibody binding to capillary endothelial cells, but not to fibroblasts. Type I, or Type II cells. Staining of fibrotic sections with MoAb 273-34A is dramatically enhanced over staining of normal lung. A third antibody, 370-8A, gives a general staining pattern throughout normal lung that is intensified in fibrotic lung. Another MoAb 328-41A mediates intense nuclear fluorescence of lung tumor cells and cells in lung sections. It binds to 14- and 17-kDa proteins that may be high mobility group (HMG) nuclear proteins. Enzyme inhibition studies and immunofluorescence staining patterns on normal lung indicate that elastin may be the target of MoAb 328-35B. MoAb 327-5B binds to normal mouse lung fibroblasts and red blood cell membranes. These last three MoAbs stain macrophages in fibrotic lung, but give a general pattern of light epithelial cell stain in normal lung sections indicating macrophage engulfment of the normal cell antigen during fibrosis. These antibodies should be useful in identifying cell types and molecular mechanisms involved in early stages of fibrosis induced by different chemical insults.

The construction and operation of a simple inexpensive slam freezing device for electron microscopy.

A simple and inexpensive slam freezing device has been constructed that allows consistently good rapid freezing of uncryoprotected tissue. The device employs a dry liquid-nitrogen-cooled cooper surface and a slamming plunger designed to provide rapid and uniform tissue to metal contact.

The intracellular fate of [3H]benzo[a]pyrene diol epoxide diastereomers localized in vivo in SENCAR mouse epidermis: a quantitative electron microscopic study.

The intracellular localization of topically applied anti and syn diastereomers of racemic benzo[a]pyrene-7,8-diol-9,10-epoxide (BPDE) was investigated in SENCAR mouse epidermis using conventional electron microscopic (EM) autoradiography. The shaved backs of adult mice were treated in vivo with the active skin tumor initiator anti [3H]BPDE or with its stereoisomer syn [3H]BPDE, which is inactive as an initiator. After 3, 12 and 24 h of exposure, thin sections of the treated skin were prepared for high resolution autoradiography to determine the intracellular distributions of radioactivity bound within the epidermis. For each treatment and exposure time, the interfollicular area of epidermis examined by EM encompassed 350-700 keratinocytes, and extended from the basal lamina to the innermost keratinizing layer at a magnification that permitted basal and suprabasal cell populations to be distinguished in situ following autoradiography. Statistical analysis of the distributions of silver grains localized over keratinocytes at 3-, 12- and 24-h timepoints revealed that 27.2% of the carcinogenic anti [3H]BPDE grains occurred over epidermal nuclei, which was significantly more (P less than 0.001) than 20.0% of the non-carcinogenic syn [3H]BPDE grains associated with nuclei at all timepoints examined. In the basal cell population at the 3-h time point 33% of both racemic diastereomers were found within nuclei; at 12 and 24 h of exposure the nuclear association of anti [3H]BPDE grains increased (P less than 0.01) to 39% and 47% respectively, while the nuclear-associated syn [3H]BPDE grains remained at 29% and 33%. The localization of [3H]BPDE observed in our experiments suggests that the epidermal cell nucleus may be an important intracellular target for anti BPDE.

Replication of the scrapie agent in ocular neural tissues.

Optic nerves and retinas removed from hamsters experimentally inoculated with the scrapie agent contain a high titer of infectivity. Ophthalmoscopic examination of these animals revealed gross lesions of retinopathy as early as 3 weeks before the onset of clinical signs of brain degeneration. These results suggest that the scrapie agent may spread centrifugally in nerve fibers after intracerebral inoculation and that the scrapie-associated retinopathy seen in hamsters is directly induced by the agent rather than the result of retrograde degeneration from central neural damage.

Hepatic and metabolic effects of ethanol on rhesus monkeys.

Rhesus monkeys were tube-fed 100 calories per kg of a liquid diet based on casein in which 41% of the calories were derived from grain alcohol. The alcohol intake was 5.8 g per kg per day. Control diets contained isocaloric amounts of glucose. The protein content of the diet was 15% and fat supplied 21% of the calories. After 28 days the animals which had been fed ethanol developed hepatic fatty change and serum L.D.H. levels were elevated. The most striking electron microscopic changes in the alcohol animals were mitochondrial swelling, focal cytoplasmic degradation, and dilatation of the rough endoplasmic reticulum. In the monkeys which had received ethanol the metabolism of alcohol increased from 17.4 mg per 100 ml per hour to 26.6 mg and antipyrene half-life decreased from 61.0 minutes to 49.9 minutes. The carbohydrate animals showed no significant change in alcohol metabolism or antipyrene half life. The ethanol animals lost weight significantly while the carbohydrate animals gained significantly. The metabolic effects of alcohol thus were not reproduced by glucose. Administration of phenobarbital at 30 mg per kg for 5 days increased alcohol metabolism from 16.5 mg per hour to 22.5 mg per hour and shortened antipyrene half life from 76.5 minutes to 33.6 minutes. Alcohol and phenobarbital both induced enhanced drug metabolism but alcohol was a more powerful inducer of its own metabolism than phenobarbital. Phenobarbital on the other hand was a better inducer of antipyrene metabolism than alcohol.