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Biomedical subjects

R E Morris

Publications and source records attributed to R E Morris.

At least 163 records · Page 9Linked to original sources
PubMed1993

Rapamycin inhibits arterial intimal thickening caused by both alloimmune and mechanical injury. Its effect on cellular, growth factor, and cytokine response in injured vessels.

The effect of rapamycin (RPM) on the extent of arterial intimal thickening was determined in rat recipients of orthotopic femoral artery allografts or in rats that had undergone balloon catheter injury to carotid arteries. In untreated rats, neointima comprised approximately 50% of the arterial wall area in both models. Although treatment of allograft recipients for 40 days with 1.5 mg/kg/day RPM was ineffective, a dose of 6 mg/kg/day (days 0-7) followed by 3 mg/kg/day (days 8-39) reduced intimal thickening by 98% (P < 0.0001). The higher RPM dose reduced T cell and macrophage infiltration significantly and decreased the expression of IL-2 receptor, class II Ag, and mRNAs for growth factors and cytokines. Treatment with 1.5 mg/kg/day RPM (days 0-13) after balloon-catheter injury reduced intimal thickening by 45% (P = 0.0254) and substantially decreased macrophage infiltration and expression of class II Ag in the adventitia. Within the neointima, however, mRNAs for platelet-derived growth factor-alpha, basic fibroblast growth factor, and transforming growth factor-beta were still expressed. In summary, we have shown that RPM inhibits not only the vascular response to injury caused by allograft rejection, but also the response to balloon catheter injury. This new information is important to our understanding of: (1) the fundamental processes responsible for intimal thickening regardless of the cause of vascular injury, (2) mechanisms of action of RPM that explain its effects on the response to very different types of vascular injury, and (3) the potentially diverse therapeutic applications of drugs, like RPM, that inhibit the actions of both immune and nonimmune cytokines and growth factors.

Animals↗
PubMed1993

Hepatic lobular patterns of phosphoenolpyruvate carboxykinase, glycogen synthase, and glycogen phosphorylase in fasted and fed rats.

The goal of this study was to localize phosphoenolpyruvate carboxykinase (PEPCK), glycogen synthase (GS), and glycogen phosphorylase (GP) in the liver lobule by immunocytochemical techniques and to describe the effects of feeding and fasting on the distribution and quantity of these enzymes. Livers from ad lib fed and overnight fasted normal adult male rats were frozen in liquid nitrogen after transcardial perfusion with 30% sucrose. Serial cryostat sections of tissue were collected on slides, fixed by immersion in 4% paraformaldehyde, and incubated with antibodies against PEPCK, GS, and GP. Antibodies to these enzymes were visualized with a gold-conjugated secondary antibody and a silver enhancement technique. Fed animals demonstrated a periportal to pericentral gradient of PEPCK. Fasting increased the periportal content of PEPCK, induced the midlobular and centrilobular cells to express the enzyme, and steepened the periportal to pericentral gradient. The increase of PEPCK was confirmed by Western blot analysis. GS and GP were distributed throughout the lobule in the fed animal but often showed a centrilobular pattern, and fasting did not alter the lobular distribution of either enzyme. Western blot analysis revealed no changes in the amount of these enzymes in the fed or fasted state. The cellular distribution of the three enzymes is similar to that of hepatic glycogen, in that the immunoreactive material has a clumped appearance in the periportal hepatocytes and is more dispersed in the pericentral cells. On fasting the periportal hepatocytes lose the dense compact localization of the enzymes and the protein becomes more homogeneously distributed throughout the cytosol. Further studies are needed to elucidate the functional significance of the regional heterogeneity of the glycogen-metabolizing enzymes and the molecular mechanisms regulating their gene expression.

Animals↗
PubMed1993

Enhancement of antigenic site detection with gold labeled secondary and tertiary antibodies using the immunogold-silver staining method.

We report a modification of the immunogold-silver staining method (IGSS) for localizing hepatic phosphoenolpyruvate carboxykinase (PEPCK) in tissue sections, and we compare the efficacy of localizing the primary antibody with either a 5 nm gold labeled secondary antibody or 5 nm gold labeled secondary and tertiary antibodies. Light microscope examination of 10 microns frozen sections demonstrated that the use of combined secondary and tertiary gold labeled antibodies was superior to using a secondary gold labeled antibody alone. The increased labeling density (number of colloidal gold particles/antigenic site/cell) achieved by combined gold labeled antibodies was confirmed by electron microscopy. The increased labeling density resulted in a two-thirds reduction in the time needed for the IGSS physical development of the silver shells and less background. We achieved intense specific staining of hepatocytes expressing PEPCK while minimizing background staining. The use of combined secondary and tertiary gold labeled antibodies enhances the signal-to-noise ratio, achieves high resolution and is a suitable method for use in both light and electron microscopy.

Animals↗
PubMed1993

HIV and adolescents.

Human immunodeficiency virus has entered the adolescent population, and pediatricians will be caring for youngsters who are HIV positive or at risk of being infected. We should remember that it is risky sexual behavior and injection drug use that places a teen at risk for HIV infection, not their sexual orientation, ethnicity, or gender. Parents and their teenagers desire and expect their pediatricians to provide care and guidance for the diseases and problems that adolescents face. A screening psychosocial assessment of teenagers can identify those who need more in-depth counseling. Although the subjects of sex and drug use initially may be uncomfortable for a physician, practice using the HEADSS assessment will rapidly lead to comfort in discussing these important subjects. An understanding of HIV testing and pre- and post-test counseling will prepare the physician for the inevitable patient who wishes testing or who is HIV positive. Much of the treatment of HIV-positive adolescents involves patience and support while the adolescent grapples with the serious implications of being HIV positive. The initial history and physical exam establishes baselines regarding previous infections and illnesses that may bear on HIV infection, as well as determining which symptoms and signs of HIV infection are present. The initial laboratory tests further define the patient's current clinical state and will determine what therapies are immediately needed. Human immunodeficiency virus continues to evolve toward a manageable chronic illness that responds most favorably to early intervention.

Adolescent↗
PubMed1992

The alpha 2-macroglobulin receptor/low density lipoprotein receptor-related protein binds and internalizes Pseudomonas exotoxin A.

The alpha 2-macroglobulin receptor/low density lipoprotein receptor-related protein (alpha 2 MR/LRP) is a large cell-surface glycoprotein consisting of a 515-kDa and an 85-kDa polypeptide; this receptor is thought to be responsible for the binding and endocytosis of activated alpha 2-macroglobulin and apoE-enriched beta-very low density lipoprotein. A similar high molecular weight glycoprotein has been identified as a potential receptor for Pseudomonas exotoxin A (PE). We demonstrate that the alpha 2 MR/LRP and the PE-binding glycoprotein have a similar mobility upon sodium dodecyl sulfate-polyacrylamide gel electrophoresis and are immunologically indistinguishable. Furthermore, affinity-purified alpha 2 MR/LRP binds specifically to PE but not to a mutant toxin defective in its ability to bind cells. The 39-kDa receptor-associated protein, which blocks binding of ligands to alpha 2 MR/LRP, also prevents binding and subsequent toxicity of PE for mouse fibroblasts. The concentration of receptor-associated protein that was required to reduce binding and toxicity to 50% was approximately 14 nM, a value virtually identical to the KD measured for the interaction of receptor-associated protein with the purified receptor. Overall, the studies strongly suggest that the alpha 2 MR/LRP is responsible for internalizing PE.

ADP Ribose Transferases↗
PubMed1992

Brief treatment with rapamycin in vivo increases responsiveness to alloantigens measured by the mixed lymphocyte response.

Rapamycin (RPM) is a macrolide fermentation product that prolongs rodent allograft survival more potently and effectively than cyclosporin A (CsA) and FK506. Experiments in vitro have shown that RPM inhibits lymphoproliferation by mechanisms of action that are different from other immunosuppressants. Much less is known, however, about the effects of RPM on immune cells in vivo compared to other immunosuppressive drugs. Others have shown that in vivo treatment with CsA suppresses the responsiveness of cells in the mixed lymphocyte response (MLR). Therefore, to investigate the effects of RPM in vivo, rats were treated with RPM and their lymphoid cells used as responder cells in the MLR. We confirmed that the proliferation of cells in the MLR was decreased after treatment with CsA in vivo. In contrast, treatment with RPM in vivo greatly increased the proliferative response to alloantigen in the MLR. These findings show that the effects of RPM and CsA on immune cells in vivo differ. Perhaps the cells proliferating in the MLR after in vivo RPM treatment play a role in the regulation of the immune system that enables this immunosuppressant to prolong allograft survival so effectively in rodents.

Animals↗
PubMed1992

Lectin activity of the major surfactant protein (SP-A) may participate in, but is not required for, binding to rat type II cells.

Pulmonary surfactant is a complex mixture of lipids and proteins, of which surfactant protein A (SP-A) is the most abundant glycoprotein. The SP-A molecule has several distinct structural features that include a lectin-like domain, sharing structural features with other mammalian lectins. We have tested the hypothesis that lectin activity of the SP-A molecule is required for the binding to its receptor on the surface of alveolar Type II cells. By using colloidal gold immunocytochemistry in conjunction with electron microscopy, we evaluated the ability of mannosylated proteins to inhibit canine SP-A binding to rat Type II cells in vitro. After preincubation of SP-A with the mannosylated protein horse-radish peroxidase (HRP), SP-A was incubated with isolated filter-grown Type II cells. HRP did not alter the binding of SP-A to the Type II cell surface. Evidence that SP-A did bind to HRP was shown by coincident observation of gold-labeled SP-A and HRP precipitates. These results provide visual evidence that the lectin activity associated with SP-A is not required for its binding to receptor on rat alveolar Type II epithelial cells.

Animals↗
PubMed1992

Validation of the biotinyl ligand-avidin-gold technique.

We demonstrate here that the intracellular routing of biotinylated ligands was not affected by the attachment of streptavidin gold colloids so long as the electron-dense marker was added after the biotinyl ligand-receptor interaction had occurred. The binding, internalization, and intracellular routing of three different biotinyl ligands were followed in mouse LM fibroblasts. The biotinyl (B) ligands included B-choleragenoid (B-CTd), B-wheat germ agglutinin (B-WGA), and B-Pseudomonas exotoxin A (B-PE). All three ligands showed distinct intracellular trafficking patterns. B-WGA and B-PE entered via clathrin-coated pits, whereas B-CTd did not. After entry, B-CTd was routed to the lysosomal compartment without involvement of the Golgi. Although B-PE and B-WGA were also routed to the lysosomal compartment, a significant portion of these two ligands was observed in association with the Golgi. B-WGA, however, remained in the endosomal and Golgi compartments longer than did B-PE. We also monitored the internalization and routing of native PE by an indirect immunoperoxidase technique done in conjunction with saponin solubilization. The results corroborated the observations with the biotinyl-PE-streptavidin-gold method. In contrast, biotinyl-PE added to streptavidin-gold before addition to LM cells was poorly internalized and routed aberrantly. From these observations we conclude that the biotinyl ligand-avidin-gold technique is a valid method for following the binding, internalization, and intracellular routing of ligands.

ADP Ribose Transferases↗
PubMed1991

Effects of cyclosporin, FK506, and rapamycin on graft-vessel disease.

Graft-vessel disease (GVD) limits the long-term survival of heart-transplant patients, and this effect has not been altered by use of cyclosporin for immunosuppression. We compared the effects of the immunosuppressants cyclosporin, FK506, and rapamycin on GVD in a rat-heart transplantation model. Allografted hearts from rats treated with 1 mg/kg FK506 for 50 days showed the same degree of myocardial rejection but a significantly worse (p less than 0.05) grade of GVD compared with grafted hearts from rats treated with 1.5 mg/kg cyclosporin for the same time. 2 mg/kg FK506 for 50 days prevented cellular rejection but GVD was as severe as that found with 1 mg/kg FK506. Moderate GVD was present in two of five allografted hearts after treatment with 4 mg/kg FK506. 1.5 mg/kg rapamycin for 50 days was an effective inhibitor of rejection and GVD. Based on our results in rats, the possibility that GVD may occur in human heart-transplant recipients treated with FK506 cannot be excluded.

Animals↗
PubMed1991

Isolation and characterization of Pseudomonas aeruginosa exotoxin A binding glycoprotein from mouse LM cells.

A Pseudomonas aeruginosa exotoxin A (PE) binding glycoprotein was affinity purified from toxin sensitive mouse LM cells. The binding protein was solubilized with Triton X-100 or Nonidet P-40 and purified on a PE-Sepharose affinity column. Polyacrylamide gel electrophoresis yielded a single band with an estimated molecular mass of greater than 300,000 Da. N-Linked carbohydrate was present, accounting for approximately 10% of the total mass of the molecule. The purified protein specifically bound PE. Incubation of purified protein specifically bound PE. Incubation of purified PE binding protein with toxin reduced toxicity to LM cells. We speculate on the role of this toxin binding glycoprotein in the intoxication process.

ADP Ribose Transferases↗