Shiga-like toxin-converting phages from Escherichia coli strains that cause hemorrhagic colitis or infantile diarrhea.

Escherichia coli K-12 acquired the ability to produce a high titer of Shiga-like toxin after lysogenization by either of two different bacteriophages isolated from a highly toxinogenic Escherichia coli O157:H7 strain that causes hemorrhagic colitis. One of these phages and another Shiga-like toxin-converting phage from an Escherichia coli O26 isolate associated with infantile diarrhea were closely related in terms of morphology, virion polypeptides, DNA restriction fragments, lysogenic immunity, and heat stability, although a difference in host range was noted. These phages are currently the best-characterized representatives from a broader family of Shiga-like toxin-converting phages.

Protein crystal growth in microgravity.

The crystals of most proteins or other biological macromolecules are poorly ordered and diffract to lower resolutions than those observed for most crystals of simple organic and inorganic compounds. Crystallization in the microgravity environment of space may improve crystal quality by eliminating convection effects near growing crystal surfaces. A series of 11 different protein crystal growth experiments was performed on U.S. space shuttle flight STS-26 in September 1988. The microgravity-grown crystals of gamma-interferon D1, porcine elastase, and isocitrate lyase are larger, display more uniform morphologies, and yield diffraction data to significantly higher resolutions than the best crystals of these proteins grown on Earth.

Molecular structure of 1-(2-chloroethyl)-3-(trans-4-methylcyclohexyl)-1-nitrosourea.

The three-dimensional structure of 1-(2-chloroethyl)-3-(trans-4-methylcyclohexyl)-1-nitrosourea (MeCCNU, NSC-95441), an effective antitumor agent, has been determined by single-crystal x-ray diffraction. MeCCNU crystallizes in monoclinic space group P21/c, with cell dimensions a = 12.387, b = 10.810, and c = 10 .198 A , beta = 102.62 degrees , and Z = four molecules per unit cell. The structure was solved by direct phasing procedures and refinement by anisotropic least squares converged at a discrepancy index R = 0.065. The cyclohexyl ring is in the chair conformation with the plane of the nitrosourea moiety twisted approximately 90 degrees from the cyclohexyl ring. The carbon-nitrogen bonds of the urea group are significantly asymmetric.

Activated cyclophosphamide anticancer drugs: molecular structures of cis- and trans-4-hydroperoxyisophosphamides.

Molecular structures of two stereoisomers of 4-hydroperoxyisophosphamide (HPIPA) have been determined by single-crystal X-ray diffraction. These isomers are active cytostatic agents closely related to an active metabolite of the antitumor drug isophosphamide, an analogue of cyclophosphamide. Both isomers crystallize in monoclinic space group P21/c with cell dimensions for cis-HPIPA of a = 8.999 (2), b = 8.743 (2), c = 17.078 (4) A; beta = 107.91 (2) degrees, and Z = 4 molecules per unit cell, and cell dimensions for trans-HPIPA of a = 15.184 (3), b = 10.345 (3), c = 18.205 (3) A, beta = 114.15 (1) degrees, and Z = 8. The structures were solved by direct methods and refined by anisotropic least squares to a discrepancy index R = 0.048 for cis-HPIPA and R = 0.065 for trans-HPIPA. In both isomers the 4-hydroperoxy group is situated axial to the ring. The phosphoryl oxygen atom is situated axial to the ring and, thus, cis to the C(4) oxygen in the cis-HPIPA isomer. In the trans-HPIPA isomer, the phosphoryl oxygen is equatorial to the ring and trans to the C(4) oxygen.

Crystal structure of quinespar, a quinazoline analogue of methotrexate.

The molecular structure of quinespar, a quinazoline analogue of methotrexate and aminopterin, has been determined by X-ray crystallography. The molecule displays an extended conformation with the p-aminobenzoyl plane rotated 66 degrees from the plane of the quinazoline. The orientation of the quinazoline ring relative to the rest of the molecule is intermediate between the orientations of the comparable pteridine rings in folic acid and in DHFR-bound methotrexate. Evidence is presented to suggest that 2,4-diaminoquinazolines bind to DHFR in the same manner as do 2,4-diaminopteridines.

Discovery, synthesis, and bioactivity of bis(heteroaryl)piperazines. 1. A novel class of non-nucleoside HIV-1 reverse transcriptase inhibitors.

A variety of analogues of 1-[4-methoxy-3,5-dimethylbenzyl]-4-[3-(ethylamino)-2-pyridyl]piperazine hydrochloride (U-80493E) were synthesized and evaluated for their inhibition of human immunodeficiency virus type 1 (HIV-1) reverse transcriptase (RT). Replacement of the substituted aryl moiety with various substituted indoles provided bis(heteroaryl)piperazines (BHAPs) that were 10-100-fold more potent than U-80493E. The pyridyl portion of the lead molecule was found to be very sensitive to modifications. Extensive preclinical evaluations of several of these compounds led to the selection of 1-[(5-methoxyindol-2-yl)carbonyl]-4-[3-(ethylamino)-2- pyridyl]piperazine methanesulfonate (U-87201E, atevirdine mesylate) for clinical evaluation.

Contribution of leukotriene B4 to airway inflammation and the effect of antagonists.

Inhalation of aerosols of ovalbumin in sensitized guinea pigs produced a marked, bronchoalveolar eosinophilia 24 hr after challenge. The lung eosinophilia was not prevented by the cyclooxygenase inhibitors, indomethacin or PAF antagonists (WEB-2086 and L-652731) but was inhibited by methylprednisolone, the 5-LO inhibitor, U-66858 and a series of structural analogs of LTB4, U-75302, U-77692, U-75485 and U-78489. The effectiveness of LTB4 antagonists but not PAF antagonists in vivo was consistent with in vitro studies in which LTB4 was shown to be far more chemotactic than PAF for guinea pig eosinophils. LTB4 elicited maximal directional migration of guinea pig eosinophils at concentrations from 10(-7) M to 10(-9) M while PAF showed no effect over the same concentration range. The structural analogs of LTB4 were shown to inhibit LTB4 induced chemotaxis of guinea pig eosinophils and produced a dose-related inhibition of binding of LTB4 to guinea pig eosinophil membranes. To add further proof to the hypothesis that LTB4 contributed to the antigen-induced lung eosinophilia we attempted to measure LTB4 release into BAL fluid immediately after and at various time points up to 24 hr after antigen inhalation. However, using a sensitive radioimmunoassay (detection limit 10 pg/ml) very low levels of LTB4 (24.9-67.9 pg/ml) or its metabolite, 20-OH LTB4 (24.9-98.2 pg/ml) were detected in BAL fluid and these levels did not increase significantly following antigen provocation. Inhalation of LTB4 aerosols in unsensitized Brown-Norway rats or inhalation of aerosols of ovalbumin in sensitized Brown-Norway rats also produced a marked "late-phase" eosinophil-rich influx of inflammatory cells into the lungs. The lung eosinophilia in the rat was prevented by two structurally unrelated leukotriene B4 (LTB4) antagonists, U-75302 and Ly255283. These data implicate LTB4 as a mediator of allergen-induced bronchopulmonary eosinophilia. Leukotriene B4 antagonists may provide leads for the development of compounds which inhibit the chronic airway inflammation associated with asthma in man.

Experimental infection of calves, piglets and lambs with mixtures of invasive and enteropathogenic strains of Escherichia coli.

Mixtures of mutant organisms of an invasive (IN) strain, a strain enteropathogenic (EP) for calves, lambs and piglets, and a non-pathogenic (NP) strain of Escherichia coli, that could be differentiated from each other by their antibiotic resistances, were given orally to an immunoglobulin-negative (IG-) and an immunoglobulin-positive (IG+) calf, lamb and piglet. By the use of appropriate antibiotic-containing culture media, the concentrations of organisms of each of the three strains in the contents of the alimentary tracts and tissues (liver, spleen, kidney, blood and mesenteric lymph glands) of the animals were estimated when they were killed. In the three IG- animals, the IN strain proliferated in the tissues and the EP strain proliferated in the small intestines; in general, the concentrations of the IN strain in the small intestines and the EP strain in the tissues, apart from the mesenteric lymph glands, were as low as those of the NP strain in these sites. The only strain that proliferated in the small intestine of the IG+ animals was the EP strain, which proliferated in the small intestine of the calf and piglet; no organisms of either strain were isolated from the tissues of these three animals, except from their mesenteric lymph glands.

A transmissible plasmid determining lactose fermentation and multiple antibiotic resistance in a strain of Klebsiella pneumoniae.

In a wild strain of Klebsiella pneumoniae the plasmid that determined lactose fermentation also determined resistance to chloramphenicol, ampicillin, tetracyclines, streptomycin, spectinomycin, and sulphonamides. The plasmid transferred at a very low rate to Escherichia coli K12 and Salmonella typhi. By implanting other transfer factors in the strain the rate of transfer and the recipient range were increased. Plasmid transfer from the modified strain to Salm. typhimurium and Salm. gallinarum was detected in the alimentary tract of experimentally infected chicks fed diets containing ampicillin.

Treatment of experimental Escherichia coli infection in mice with colicine V.

Concentrated non-toxic preparations of colicine V were obtained by filtering centrifugates of soft-agar cultures of a Col V+ K12 strain of E. coli through Millipore filters in which the colicine V was retained. These preparations, given subcutaneously, favourably influenced the course of disease in mice infected intraperitoneally with a pathogenic strain of E. coli that was very sensitive to colicine V in vitro. A beneficial effect was noted even when treatment was delayed until the mice were visibly ill.

Immunogenicity of biopharmaceuticals in laboratory animals.

The potential immunogenicity of new protein therapeutics raises concerns about the possibility of inducing untoward immune reactions in humans. It is generally assumed that all animals will make antibody to human proteins and therefore, there is sentiment among some scientists that this makes the issue of immunogenicity as a safety concern irrelevant. However, recent clinical trials with some proteins have detected the presence of autoantibodies that have resulted in clinical sequelae. These reactions were also observed in preclinical animal studies. In fact, non-human primate and transgenic mouse models can be useful for predicting the relative immunogenicity of human proteins. In addition, the characterization of the immunogenicity of biotechnology molecules provides a practical basis for determining the significance of antibody formation in preclinical safety studies.

Calibrated otoplasty.

The number of patients seeking correction of abnormally protruding ears has decreased in the past ten years because of changing hairstyles and concepts of body image. During the same time there has been an increase in the number of surgeon and surgical disciplines seeking to perform cosmetic otoplasty. To compensate for this deficit in total volume experience of the individual operator and to insure patients of a consistently acceptable cosmetic result, the concept of calibrated otoplasty is presented. A study was made of a large number of ears which appeared to be esthetically in harmony with each other and with the side of the head. The distance relationships obtained from this study were applied not only to the creation of a new antihelix in the first ear but the degree to which the ear projects from the side of the head. At the conclusion of the first ear operation, careful measurements were taken for use in the second ear operation to insure its being as near normal as the first in form and degree of projection of the ear from the head. This concept of calibrated otoplasty is explained in detail with illustrations of all required measurements plus a series of surgical illustrations showing application of the technique.

Palato-pharyngeal cine photography: a technique for developing continuing education films.

A method of Cine photography has been developed to record a variety of plastic and reconstructive surgical procedures in the oral cavity and pharynx. Effective classroom teaching of surgical closure of the cleft palate and the various palatal lengthening procedures including pharyngeal flaps, has been hampered by the lack of quality teaching films demonstrating surgical techniques in color. The small size of the structures involved and the delicate nature of the surgery requires that the surgeon work at close range and directly in front of the mouth. To avoid the shadows cast by the sides of the oral cavity, the surgeon's hands and frequent change of position by the professional photographic team, a head-mounted Cine camera was employed. A "through the surgeon's eye" view for the camera was accomplished by using a parallelogram of beam-splitting mirrors. The resulting photographic film provides the classroom audience with a sense of being in the surgeon's operating position thus having a view superior to that obtained by the surgeon's first assistant. An elaborate system of counter-balancing provides the surgeon with a weightless camera, mirror, and light complex mounted on a football helmet. The football helmet merely served as an interlocking unit enabling the surgeon to compose his own movie while operating.

Directional handle for Zeiss operating microscope.

A new directional handle for the Zeiss operating microscope is presented. This handle offers the advantage of single-handed (right or left) control by natural movement of the wrist. The handle may be used by grasping it through the see-through sterile drape or sterilized separately and placed outside the sterile microscope drape.

Steroid use in idiopathic sudden sensorineural hearing loss.

Few studies have investigated the effect of steroids on idiopathic sudden hearing loss (ISHL). Recently, Wilson, et al., reported a double blind study demonstrating a statistically significant improvement of hearing using steroids is ISHL. This report, fashioned after the Wilson study, confirms his findings and analyzes the relationship of several factors to the probability of hearing recovery.

Ethical use of Yellow Pages listings by physicians.

A recent article in the New England Journal of Medicine examined the percentage of physicians listed under specialty headings in a Yellow Pages publication who were board certified. The authors presented findings that many of the physicians listed in the Yellow Pages, particularly under the heading of "Plastic Surgery," were not board certified. The source data employed in that article were obtained and reexamined, and it was found that the assumptions used were inaccurate and the methodology flawed. Almost every physician listed under "Plastic Surgery" in the Yellow Pages is board certified. We discuss the conclusions of the original article and its recommendations for control of Yellow Pages listings. We believe that these recommendations are not in the public's best interests and may be illegal as well.

Cleft lip and palate surgery in La Ceiba, Honduras.

The volume of experience in the surgical repair of cleft lip and palate deformities is limited in many areas of the United States. This deficit in experience exists not only for those in resident training programs but also for those who practice at universities or are in private practice. Physicians are performing too few operations to maintain their surgical skills in cleft lip and palate surgery. This deficit is also encountered by physicians in the major specialties of plastic surgery, oral surgery, and otolaryngology who perform cleft lip and palate surgery. Physicians from the United States can gain surgical experience in other countries through groups like the Medical Group Mission Christian Medical and Dental Society. Surgical experience is available in Central and South America, India, China, and Africa through similar organizations. The complexity of providing service to under-privileged people through organizations such as the Medical Group Mission Christian Medical and Dental Society is described.

Concanavalin A and wheat germ agglutinin receptors in the ascitic fluid of rat hepatomas.

The presence of glycopeptide lectin receptors in the ascitic fluid of rats bearing Novikoff or AS-30D hepatoma was investigated. Macrosialoglycopeptides, resistant to pronase digestion, were partially purified from the ascitic fluid of hepatoma-bearing rats by gel filtration on Sephadex G-50. A macrosialoglycopeptide fraction, isolated from the ascitic fluid of rats bearing the Novikoff hepatoma, possessed potent concanavalin A (Con A) receptor activity. This fraction possessed higher Con A receptor activity than did the comparable macrosialoglycopeptide fraction from the ascitic fluid of rats bearing the AS-30D hepatoma; this obeservation is in agreement with the Con A-induced agglutination properties of these 2 hepatoma cell lines and with the Con A receptor activities of the glycopeptides released from the surface of the hepatoma cells by papain digestion. Rat blood serum contained a comparable macrosialoglycopeptide fraction, which possessed weak Con A receptor activity. The macrosialoglycopeptide fractions from the ascitic fluid of hepatoma-bearing rats possessed wheat germ agglutinin receptor activity. However, this activity was also present in normal rat serum. These results suggest that glycopeptides present on the surface of Novikoff hepatoma cells are shed into the ascitic fluid and may be distinguished from components in normal serum by their Con A receptor activity.

Radiology of the normal maxillary sinus and related structures.

The radiographic anatomy of the maxillary sinus and related structures as shown in the Waters, lateral, submentovertical, and Caldwell views has been presented. A number of structures, which are of clinical importance but infrequently emphasized, are discussed. Careful study of these four standard sinus projections should provide an excellent knowledge of the radiographic anatomy of the maxillary sinus and its normal variants.

Effect of Piriprost, a 5-lipoxygenase inhibitor, on leukocyte accumulation during thioglycollate-induced acute inflammation.

We evaluated the effect of a prototype 5-lipoxygenase enzyme inhibitor, Piriprost [6,9-deepoxy-6,9-(phenylimino-)delta 6,8-prostaglandin I1], on the leukocyte accumulation induced by intraperitoneal injection of thioglycollate in rats. Piriprost, at 40 mg/kg, i.p., inhibited leukocyte accumulation by 34 +/- 12% (mean +/- S.D., n = 9). With experimental groups containing small numbers of rats (6 to 9) inhibition was statistically significant in 7 of 9 cases. Piriprost shared this property with dexamethasone; however, dexamethasone at 5 mg/kg, p.o., inhibited leukocyte accumulation to a greater extent, 61 +/- 6% (mean +/- S.D., n = 5) and the inhibition was statistically significant in 5 of 5 cases. Conventional non-steroidal antiinflammatory agents including aspirin and phenylbutazone were ineffective in our model. Our results suggest that certain compounds that inhibit the 5-lipoxygenase enzyme, without inhibiting the cyclooxygenase enzyme, can reduce the leukocyte accumulation associated with acute inflammation.