Initial clinical experience with a rationally designed, genetically engineered recombinant human hemoglobin.
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Acute-Phase Reaction↗
Biomedical subjects
S A Shoemaker
Publications and source records attributed to S A Shoemaker.
DNA molecular biology in the diagnosis of pulmonary disease.
DNA molecular biology is becoming increasingly important in clinical medicine. It has provided a new method to diagnose an inherited pulmonary disease in which the biochemical defect has been defined. Alpha-1-antitrypsin deficiency can now be diagnosed by direct analysis for the disease gene. In cystic fibrosis, another inherited pulmonary disease, the biochemical defect has not yet been defined. However, the search has been narrowed. The cystic fibrosis genetic defect has recently been localized to the long arm of chromosome 7. Because polymorphic DNA markers (RFLPs) are available for this region of chromosome 7, it is now often possible, using linkage analysis, to trace the inheritance of the cystic fibrosis gene(s) in families known to have cystic fibrosis. When the cystic fibrosis genetic defect is defined, it will then be possible to look directly for the disease gene. Finally, the greatest impact in clinical medicine may well be the development of rapid DNA hybridization techniques to diagnose infectious diseases that currently take days to weeks to diagnose.
Bacterial Infections↗
Restriction fragment analysis of chromosomal DNA defines different strains of Mycobacterium tuberculosis.
As an initial step in gaining a better understanding of the important clinical properties that vary between strains of mycobacteria, we attempted to find molecular markers that would define different strains of Mycobacterium tuberculosis. We used restriction fragment analysis with the endonuclease MboI and hybridization with total M. tuberculosis DNA to examine DNA differences between 15 strains of M. tuberculosis. We were able to identify different strains using this method. In order to assess the sensitivity of this method in identifying different strains, we compared it with phage typing. The 2 methods appear to be similar in sensitivity and also to be complementary. There were 2 examples where restriction fragment analysis did not separate strains with different phage types. In addition, there were 2 examples where phage typing did not separate strains with different restriction patterns. Finally, there were 2 epidemiologically unrelated strains with the same restriction pattern and the same phage type. This method of restriction fragment analysis of chromosomal DNA is potentially useful for epidemiologic studies of tuberculosis. Additionally, by analyzing the genome of M. tuberculosis, molecular markers may well be defined that will be useful in discovering the pathogenesis of the clinical properties of M. tuberculosis, which previously have been poorly understood.
DNA Restriction Enzymes↗
Mummies, mycobacteria, and molecular biology--the old and the new.
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Acquired Immunodeficiency Syndrome↗
Molecular biology and mycobacteria.
Mycobacteria and molecular biology provide an interesting combination of the old and the new. Using modern techniques, it will be possible to develop new ways to approach ancient diseases such as tuberculosis and leprosy and more modern diseases such as disseminated MAC infections in patients with AIDS. DNA probes will be developed to rapidly diagnose mycobacterial infections with greater accuracy. We will gain a better understanding of how infections spread if we can identify different strains of a given species and if we can monitor close contacts of infected patients with more specific immunologic tests. By understanding mechanisms of drug resistance and the host immune response to mycobacterial infection, it may be possible to develop more effective modes of treatment. Finally, it may be possible to develop effective vaccines to control the spread of infection.
DNA Restriction Enzymes↗
Techniques of DNA hybridization detect small numbers of mycobacteria with no cross-hybridization with non-mycobacterial respiratory organisms.
The traditional methods used in identifying mycobacteria, such as acid-fast bacillus stains and culture, are often time-consuming, insensitive, and nonspecific. As part of an ongoing program to improve diagnosis and characterization of mycobacteria, we have found that deoxyribonucleic acid (DNA) hybridization techniques using isotopically labeled, single-stranded, total DNA can be used to detect as little as 10(-4) micrograms of Mycobacterium tuberculosis (MTb) DNA. This amount of DNA represents approximately 2 X 10(4) genomes. We have also shown the MTb DNA is sufficiently different from the DNA of non-mycobacterial microorganisms such that cross-hybridization with MTb DNA does not occur under the hybridization conditions we employed. We speculate that DNA hybridization techniques may allow the rapid, sensitive, and specific identification of mycobacteria.
Autoradiography↗
The E7-associated cell-surface antigen: a marker for the 11p13 chromosomal deletion associated with aniridia-Wilms tumor.
Unbalanced interstitial deletions of the p13 region of human chromosome 11 have been associated with congenital hypoplasia or aplasia of the iris, mental retardation, ambiguous genitalia, and predisposition to Wilms tumor of the kidney. Utilizing somatic cell hybrids containing either the normal or abnormal chromosome 11 from a child with Wilms tumor and aniridia, we previously mapped the E7 cell-surface antigen to the 11p1300-to-11p15.1 region. To localize even further the site of this antigen on chromosome arm 11p, we have produced somatic cell hybrids from the fibroblasts of a second child with Wilms tumor and aniridia and a different deletion of 11p [46,XY, del (11)(pter----p14.1::p11.2----qter)]. Furthermore, the normal and deleted chromosome 11 could also be distinguished on the basis of a restriction fragment length polymorphism for the beta-globin gene. Hybrid cells containing the deleted chromosome were not killed in the presence of complement and the E7 monoclonal antibody (which recognizes E7 cell surface antigen), while hybrid cells containing the patient's normal chromosome 11 were killed. Thus, expression of the E7-associated cell-surface antigen can be mapped to the 11p13 region, and it appears to be a potential marker of the chromosome abnormality associated with aniridia-Wilms tumor.
Abnormalities, Multiple↗
Wilms' tumor-aniridia association: segregation of affected chromosome in somatic cell hybrids, identification of cell surface antigen associated with deleted area, and regional mapping of c-Ha-ras-1 oncogene, insulin gene, and beta-globin gene.
Fusion of an auxotrophic mutant hamster cell with the skin fibroblasts of a child with the Wilms' tumor-aniridia association produced clones which, on the one hand, contained the child's normal chromosome 11 and, on the other, the chromosome 11 with the 11p13 deletion associated with the syndrome. Both hybrids were positive for human LDH-A by enzymatic assay. Clones containing the normal human chromosome 11 were killed by a cytotoxic monoclonal antibody to a cell surface antigen previously mapped to the 11p13----11pter region of chromosome 11. Clones with the abnormal 11 were not killed. Thus, we have produced hybrids from the same patient distinct from each other on the basis of their chromosome 11. These hybrids have been used to map the locus for a cell surface antigen to the deleted region on chromosome 11 of a patient with the Wilms tumor-aniridia association. The linkage between this antigen and the syndrome should be helpful in further study of the genetics of this disease. In addition, we have found that the c-Ha-ras-1 oncogene is distal to the p13 region of chromosome 11 and the position of insulin and beta-globin on the chromosome. Finally, by producing segregants of the hybrids containing the abnormal chromosome 11, we have provided evidence that chromosome 11-associated c-Ha-ras-1 is syntenic with chromosome 11 and not moved to a different portion of the genome.
Animals↗
Staphylococcus aureus and human platelets cause pulmonary hypertension and thromboxane generation in isolated saline-perfused rabbit lungs.
The potential contributions of bacterial-platelet interactions to the development of acute edematous lung injury, such as that seen in the adult respiratory distress syndrome (ARDS), remains unknown. We found that the addition of Staphylococcus aureus, 502A to isolated rabbit lungs perfused with saline, and human platelets rapidly decreased the number of circulating platelets, increased pulmonary artery perfusion pressures, and generated thromboxane B2, the stable derivative of thromboxane A2. In contrast, increases in perfusion pressures or thromboxane levels did not occur when platelets treated with acetylsalicylic acid (ASA) were used, even though ASA-treated platelets disappeared from the perfusates. The results suggest that activation of platelets by bacteria may account for thrombocytopenia, platelet microemboli, and/or contribute to increases in pulmonary artery pressures seen in some patients with ARDS.
Animals↗
Mepacrine but not methylprednisolone decreases acute edematous lung injury after injection of phorbol myristate acetate in rabbits.
A good model of the adult respiratory distress syndrome (ARDS) is intravenously injected phorbol myristate acetate (PMA), which causes pulmonary sequestration of neutrophils and a neutrophil-dependent acute edematous lung injury in rabbits. In the present study, pretreatment of rabbits with the antimalarial agent, mepacrine, prevented lung edema after injection of PMA without altering initial accumulations of neutrophils in the lung. Mepacrine also decreased oxygen radical production and degranulation by neutrophils stimulated by PMA in vitro. In contrast, pretreatment with methylprednisolone did not decrease edematous lung injury in rabbits given PMA nor did it inhibit O2 radical production or degranulation by neutrophils treated with PMA in vitro. Our results suggest that agents that modify neutrophil function may be useful in decreasing lung injury after PMA treatment and, perhaps, in treating patients with ARDS.
Animals↗
Acetyl glyceryl ether phosphorylcholine-stimulated human platelets cause pulmonary hypertension and edema in isolated rabbit lungs. Role of thromboxane A2.
Macrophages, neutrophils, and platelets may play a role in acute edematous lung injury, such as that seen in the adult respiratory distress syndrome (ARDS), but their potential actions and interactions are unclear. Because stimulated human macrophages and neutrophils can release acetyl glyceryl ether phosphorylcholine (AGEPC), a potent platelet activator, we hypothesized that in ARDS, leukocyte release of AGEPC might stimulate platelets to release thromboxane A2 (TXA2), which then produces pulmonary hypertension and lung edema. In support of this premise, we found that pulmonary hypertension and edema occurred in isolated rabbit lungs perfused with human platelets and AGEPC, but not with platelets or AGEPC alone. Infusion of a vasodilator (nitroglycerin) to maintain base-line pulmonary artery pressures in lungs perfused with platelets and AGEPC prevented the development of lung edema suggesting that platelet and AGEPC-induced edema was hydrostatic in nature. Additional experiments suggested that the increased pressure was a result of TXA2 release from platelets stimulated by AGEPC. Specifically, preincubation of platelets with imidazole, a thromboxane synthetase blocker, prior to infusion with AGEPC significantly diminished pulmonary hypertension and prevented lung edema. Furthermore, pretreating lung preparations with 13-azaprostanoic acid, a TXA2 antagonist, before infusion of AGEPC and untreated platelets also reduced the pulmonary hypertension and blocked the lung edema. The role of TXA2 was further suggested when perfusates from lungs infused with platelets and AGEPC developed high levels of TXA2, whereas perfusates from controls did not. These results suggest that platelet aggregation induced by AGEPC may contribute to ARDS by releasing TXA2, which raises microvascular pressure and increases edema formation, especially when an underlying permeability defect is present.
Animals↗
Platelet-induced pulmonary hypertension and edema. A mechanism involving acetyl glyceryl ether phosphorylcholine and thromboxane A2.
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Albumins↗