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Transplantation  1990;49(6):1055-1059.
The capacity of specifically targeted erythrocytes to inhibit free radical–mediated injury to the endothelial cell after cold preservation, and improve liver function was studied in two experimental models: An isolated perfused rat liver (IPRL) system and syngeneic orthotopic rat liver transplantation. In the IPRL model, livers were preserved in University of Wisconsin solution for 24 h at 4°C. At the end of the preservation period, livers were flushed with lactated Ringer’s (control), immunoerythrocytes (IES), or blank intact erythrocytes prior to warm reperfusion for 2 h using an assanguinous Krebs-Henseleit buffer. Production of superoxide (O2−) anion during warm reperfusion in the IES-treated liver was reduced by 65% as compared with controls (P<0.001) and by 74% (P<0.001) when compared with blank erythrocyte–treated livers. Endothelial cell preservation, as assessed by levels of purine nucleoside phosphorylase (PNP), was much better in the IES-treated group (P<0.001) when compared with untreated livers. Hepatocellular preservation was markedly improved in the IES-treated livers.
In the syngeneic liver transplantation model, livers were preserved in UW solution for 24 h at 4°C. Prior to implantation, livers were flushed with 5 ml of cold lactated Ringer’s or immunoerythrocytes. Survival after three weeks was 60% in the IES-treated group and 30% in the untreated group. Survival in the IES-treated group was not significantly different from a control (no preservation) group.
IES-treated livers in both models demonstrated better endothelial cell integrity and ultimate liver function. IES treatment therefore appears to protect the hepatic microvascular endothelial cell from reperfusion injury and could prove to be an easy reproducible method of donor organ preparation after cold preservation.
PMCID: PMC3033047  PMID: 2163131
2.  A Modified Apparatus for Dual, Sterilized, Isolated Perfusion of the Rat Liver 
The isolated perfused rat liver (IPRL) has proven to be a useful model for the study of physiology and pathology of the liver. For research in nonparenchymal cell (NPC) function that includes measurement of cytokine production (eg, TNF), it is necessary to have a sterilized perfusion system. We have modified the IPRL apparatus so as to be able to perform sterile perfusions of two livers simultaneously. The perfusion apparatus is a recirculating closed system in which the oxygenator is a plastic container separated into two chambers by a fenestrated plastic wall. A disposable macropore filter functions as both a bubble trap and perfusate filter. The sterilization process is done by immersing the various components in Benz-All solution. The tubing is disinfected by irrigation with 10% Clorox followed by 0.9% sodium chloride solution. The perfusate used is filter-sterilized Krebs buffer solution containing 0.5 g Mandol/250 mL perfusate. Not only can two organs be conveniently perfused simultaneously, but the entire system can be reliably sterilized for up to 20 consecutive perfusions. Bile production is higher and more stable with less leakage of intracellular enzymes. Many of the components are disposable and can be altered to suit the needs of a particular experiment.
PMCID: PMC2950627  PMID: 2291894
Isolated perfused rat liver (IPRL); dual; sterilized
3.  Dynamic injection of the digital flexor tendon sheaths. 
Annals of the Rheumatic Diseases  1990;49(5):327-328.
PMCID: PMC1004081  PMID: 2344215
4.  Inhibitory role of endothelium-derived relaxing factor in rat and human pulmonary arteries. 
British Journal of Pharmacology  1990;101(1):166-170.
1. The inhibitory role of endothelium-derived relaxing factor was studied in both rat and human pulmonary arteries in vitro by inhibiting its synthesis with the L-arginine analogue NG-monomethyl-L-arginine (L-NMMA). 2. In rat pulmonary arteries, L-NMMA pretreatment (10-300 microM) dose-dependently inhibited acetylcholine-induced relaxation (which is endothelium-dependent). NG-monomethyl-D-arginine (D-NMMA, 100 microM) was without effect. L-Arginine, but not D-arginine, dose-dependently reversed this inhibition. L-NMMA had no effect on relaxation induced by sodium nitroprusside. 3. In human small pulmonary arteries L-NMMA (100 microM) pretreatment similarly inhibited the acetylcholine-induced relaxation but had no effect on the sodium nitroprusside-induced relaxation. 4. In both rat and human pulmonary arteries, L-NMMA, but not D-NMMA, always caused contraction of preconstricted tissues whereas it had no effect on baseline tone. In the rat this contraction was completely prevented by prior treatment with L-arginine. 5. L-NMMA (100 microM) pretreatment mimicked the effect of endothelium removal on phenylephrine-induced vasoconstriction, both resulting in an increase in tension development at each concentration of phenylephrine. This enhancement was greatest at low concentrations of phenylephrine but was still present even at the highest concentrations. Pretreatment with L-NMMA (100 microM) also significantly increased the responses to single doses of phenylephrine. 6. These results suggest that endothelium-derived relaxing factor from endothelial cells both mediates the relaxation response to acetylcholine and also acts as a physiological brake against vasoconstriction in pulmonary vessels.
PMCID: PMC1917660  PMID: 2282458
5.  The effects of (-)-daurisoline on Ca2+ influx in presynaptic nerve terminals. 
British Journal of Pharmacology  1990;101(1):45-48.
1. The effects of (-)-daurisoline on 45Ca2+ uptake and [3H]-gamma-aminobutyric acid ([3H]-GABA) release from synaptosomes of rat cerebral cortex and on contractile activity of rat aorta were examined. 2. Application of (-)-daurisoline (1-100 microM) produced concentration-related inhibition of high K(+)-stimulated 45Ca2+ uptake and [3H]-GABA release (IC50 = 7.7 +/- 0.9 microM and 10.0 +/- 1.5 microM, respectively) in synaptosomes but verapamil was only weakly active. 3. Neither (-)-daurisoline (100 microM) nor verapamil (100 microM) modified 45Ca2+ uptake in control medium (5 mM K+, resting uptake) and [3H]-GABA release in Ca-free medium (45 mM K+ basal release). 4. High K+ and noradrenaline-evoked contractions of rat aorta were inhibited by both (-)-daurisoline and verapamil. 5. In conclusion, (-)-daurisoline, which differed from verapamil in its mode of blocking Ca2+ influx may be a potent Ca2+ antagonist of Ca2+ channels in neurones.
PMCID: PMC1917626  PMID: 2282465
6.  Withdrawal phenomena after atenolol and bopindolol: haemodynamic responses in healthy volunteers. 
1. The effect of withdrawal of atenolol and bopindolol administration was studied in 12 normal volunteers; six on each drug. 2. Following sub-maximal cycle-ergometer exercise training six sets of base-line observations were made of heart-rate (HR) and blood pressure (BP) responses; supine, 60 degrees head-up tilt, during graduated isoprenaline infusion and sub-maximal cycle exercise. 3. The results show that withdrawal phenomena occur following both drug treatments. Atenolol produced a hypersensitivity to isoprenaline and a small overshoot of HR in response to physiological manoeuvres. In contrast bopindolol produced a prolonged state of reduced sensitivity to isoprenaline and some evidence of overshoot of HR with physiological manoeuvres. The differences between the responses may be explained by the different properties of the two beta-adrenoceptor blocking drugs. 4. Some subjects showed clear evidence of overshoot of HR and BP on exercise demonstrating that certain individuals may be more prone to have withdrawal effects than others. 5. The length of time during which withdrawal phenomena can occur is probably longer than has previously been realised. 6. Hormonal changes were found in the withdrawal period (Walden et al., 1990).
PMCID: PMC1368245  PMID: 1981318
7.  The viral envelope gene is involved in macrophage tropism of a human immunodeficiency virus type 1 strain isolated from brain tissue. 
Journal of Virology  1990;64(12):6148-6153.
Human immunodeficiency virus type 1 (HIV-1) strains isolated from the central nervous system (CNS) may represent a subgroup that displays a host cell tropism different from those isolated from peripheral blood and lymph nodes. One CNS-derived isolate, HIV-1SF128A, which can be propagated efficiently in primary macrophage culture but not in any T-cell lines, was molecularly cloned and characterized. Recombinant viruses between HIV-1SF128A and the peripheral blood isolate HIV-1SF2 were generated in order to map the viral gene(s) responsible for the macrophage tropism. The env gene sequences of the two isolates are about 91.1% homologous, with variations scattered mainly in the hypervariable regions of gp120. Recombinant viruses that have acquired the HIV-1SF128A env gene display HIV-1SF128A tropism for macrophages. Furthermore, the gp120 variable domains, V1, V2, V4, and V5, the CD4-binding domain, and the gp41 fusion domain are not directly involved in determining macrophage tropism.
PMCID: PMC248789  PMID: 2243391
8.  Expression of type 8 capsular polysaccharide and production of toxic shock syndrome toxin 1 are associated among vaginal isolates of Staphylococcus aureus. 
Journal of Clinical Microbiology  1990;28(12):2612-2615.
A colony immunoblot method was developed for serotyping the capsular polysaccharides expressed by Staphylococcus aureus isolates. The method was rapid and specific and was performed with either polyclonal or monoclonal antibodies specific for each of the capsule types. S. aureus isolates were obtained from patients with toxic shock syndrome (TSS) or other staphylococcal infections and from asymptomatic women with vaginal colonization. Among the vaginal isolates of S. aureus, expression of the type 8 capsule was significantly (P less than 0.001) more frequent among strains that produced TSS toxin 1 (TSST-1) than it was among TSST-1-negative strains. In contrast, the frequency of type 8 capsule expression was similar among both TSST-1-positive and -negative strains of S. aureus from patients with nonvaginal TSS. When all vaginal and nonvaginal isolates were compared, TSST-1-negative S. aureus strains were equally distributed among the type 5 and 8 and nontypeable capsule groups, whereas TSST-1-positive strains were predominantly capsule type 8.
PMCID: PMC268243  PMID: 2279990
9.  The role of the CCAAT/enhancer-binding protein in the transcriptional regulation of the gene for phosphoenolpyruvate carboxykinase (GTP). 
Molecular and Cellular Biology  1990;10(12):6264-6272.
Previous studies have identified a region in the promoter of the gene for phosphoenolpyruvate carboxykinase (GTP) (PEPCK) (positions -460 to +73) containing the regulatory elements which respond to cyclic AMP, glucocorticoids, and insulin and confer the tissue- and developmental stage-specific properties to the gene. We report that CCAAT/enhancer-binding protein (C/EBP) binds to the cyclic AMP-responsive element CRE-1 as well as to two regions which have been previously shown to bind proteins enriched in liver nuclei. The DNase I footprint pattern provided by the recombinant C/EBP was identical to that produced by a 43-kDa protein purified from rat liver nuclear extracts, using a CRE oligonucleotide affinity column, which was originally thought to be the CRE-binding protein CREB. Transient contransfection experiments using a C/EBP expression vector demonstrated that C/EBP could trans activate the PEPCK promoter. The trans activation occurred through both the upstream, liver-specific protein-binding domains and the CRE. The CRE-binding protein bound only to CRE-1 and not to the upstream C/EBP-binding sites. The results of this study, along with physiological properties of C/EBP, indicate an important role for this transcription factor in providing the PEPCK gene with several of its regulatory characteristics.
PMCID: PMC362901  PMID: 2147222
10.  PC Programs for Adaptive Control of Drug Dosage Regimens and for Population Pharmacokinetic Modeling 
The USC*PACK PC clinical programs perform Bayesian adaptive control of drug dosage regimens. They improve prediction of serum levels of aminoglycosides and vancomycin, improve management of digoxin, reduce breakthrough arrhythmias in patients on lidocaine, and help optimize cancer chemotherapy. A new nonparametric EM (NPEM) program for population pharmacokinetic modeling computes the joint probability density function (PDF) for a 1-compartment pharmacokinetic model with IV dosing (see Figure 1). Output also includes marginal PDF plots, means, variances, modes, quartiles, skewness, kurtosis, and correlation coefficient. Results can be entered into population files for use with the PC clinical programs. The programs are available from the first author through a non-commercial license from the University of Southern California. (Supported by NIH Grant RR01629).
PMCID: PMC2245455
11.  Detection of Epstein-Barr virus DNA sequences in nasopharyngeal carcinoma cells by enzymatic DNA amplification. 
Journal of Clinical Microbiology  1990;28(11):2398-2402.
The presence of Epstein-Barr virus (EBV) DNA sequences was examined by the polymerase chain reaction in 50 nasopharyngeal carcinoma (NPC) biopsy specimens and in two primary epithelial tumor cell cultures derived from patients with NPC. The detection limit was a single EBV genome equivalent by agarose gel electrophoresis followed by Southern blot analysis of the amplified products. EBV DNA sequences were detected in all 41 undifferentiated NPC cell specimens, in 2 of 4 moderately differentiated NPC cell specimens, and in 3 of 5 keratinized NPC cell specimens. Undifferentiated NPC cells were also found to contain higher copy numbers of EBV than cells of the other two types of NPC. Our data suggest that EBV replication may be closely associated with the differentiation of NPC tumor cells. The results also demonstrated a sensitive and specific method for the detection of EBV DNA sequences in NPC tumor cells.
PMCID: PMC268195  PMID: 2174898
12.  Interactions of phosphatidylinositol kinase, GTPase-activating protein (GAP), and GAP-associated proteins with the colony-stimulating factor 1 receptor. 
Molecular and Cellular Biology  1990;10(11):5601-5608.
The interactions of the macrophage colony-stimulating factor 1 (CSF-1) receptor with potential targets were investigated after ligand stimulation either of mouse macrophages or of fibroblasts that ectopically express mouse CSF-1 receptors. In Rat-2 cells expressing the mouse CSF-1 receptor, full activation of the receptor and cellular transformation require exogenous CSF-1, whereas NIH 3T3 cells expressing mouse c-fms are transformed by autocrine stimulation. Activated CSF-1 receptors physically associate with a phosphatidylinositol (PI) 3'-kinase. A mutant CSF-1 receptor with a deletion of the kinase insert region was deficient in its ability to bind functional PI 3'-kinase and to induce PI 3'-kinase activity precipitable with antiphosphotyrosine antibodies. In fibroblasts, CSF-1 stimulation also induced the phosphorylation of the GTPase-activating protein (GAP)-associated protein p62 on tyrosine, although GAP itself was a relatively poor substrate. In contrast to PI 3'-kinase association, phosphorylation of p62 and GAP was not markedly affected by deletion of the kinase insert region. These results indicate that the kinase insert region selectively enhances the CSF-1-dependent association of the CSF-1 receptor with active PI 3'-kinase. The insert deletion mutant retains considerable transforming activity in NIH 3T3 cells (G. Taylor, M. Reedijk, V. Rothwell, L. Rohrschneider, and T. Pawson, EMBO J. 8:2029-2037, 1989). This mutant was more seriously impaired in Rat-2 cell transformation, although mutant-expressing Rat-2 cells still formed small colonies in soft agar in the presence of CSF-1. Therefore, phosphorylation of GAP and p62 through activation of the CSF-1 receptor does not result in full fibroblast transformation. The interaction between the CSF-1 receptor and PI 3'-kinase may contribute to c-fms fibroblast transformation and play a role in CSF-1-stimulated macrophages.
PMCID: PMC361316  PMID: 2172781
13.  Limited Multiplication of Symbiotic Cyanobacteria of Azolla spp. on Artificial Media 
Applied and Environmental Microbiology  1990;56(11):3623-3626.
We examined various media and conditions to isolate symbiotic cyanobacteria from the leaf cavities of Azolla spp. Cyanobacteria survived and multiplied to a limited extent on a medium with fructose, Casamino Acids, yeast extract, and NaNO3 under 1% O2. These cyanobacteria were antigenically identical to the endosymbionts.
PMCID: PMC185038  PMID: 16348366
14.  Fluorosilicone oil in the treatment of retinal detachment. 
We evaluated the use of a heavier-than-water fluorinated silicone oil in the treatment of 30 selected cases of complicated retinal detachment from January 1988 to July 1989. Proliferative vitreoretinopathy grade C-2 or greater accounted for 19 cases, proliferative diabetic retinopathy with traction detachment for two cases, giant retinal tears five, ruptured globe with retinal detachment two, massive choroidal effusion with retinal detachment one, and acute retinal necrosis with retinal detachment one. Initial retinal reattachment was achieved in all cases. Complications included redetachment seven (23%), cataract six (75% of phakic patients), raised intraocular pressure four (13%), hypotony four (13%), keratopathy three (10%), uveitis-synechia formation three (10%), phthisis two (3%), choroidal haemorrhage one (3%), and vitreous haemorrhage one (3%). Postoperative visual acuities with at least six months' follow-up range from no light perception to 20/50, with seven patients (23%) 20/400 or better.
PMCID: PMC1042248  PMID: 2223698
15.  Radon in California homes. 
Western Journal of Medicine  1990;153(4):446-447.
PMCID: PMC1002591  PMID: 2244386
16.  groE genes affect SOS repair in Escherichia coli. 
Journal of Bacteriology  1990;172(10):6135-6138.
Repair of UV-irradiated bacteriophage in Escherichia coli by Weigle reactivation requires functional recA+ and umuD+C+ genes. When the cells were UV irradiated, the groE heat shock gene products, GroES and GroEL, were needed for at least 50% of the Weigle reactivation of the single-stranded DNA phage S13. Because of repression of the umuDC and recA genes, Weigle reactivation is normally blocked by the lexA3(Ind-) mutation (which creates a noncleavable LexA protein), but it was restored by a combination of a high-copy-number umuD+C+ plasmid and a UV dose that increases groE expression. Maximal reactivation was achieved by elevated amounts of the Umu proteins, which was accomplished in part by UV-induced expression of the groE genes. By increasing the number of copies of the umuD+C+ genes, up to 50% of the normal amount of reactivation of S13 was achieved in an unirradiated recA+ host.
PMCID: PMC526941  PMID: 2211529
17.  Expression of transforming growth factor-alpha in primary human colon and lung carcinomas. 
British Journal of Cancer  1990;62(3):425-429.
The expression of TGF-alpha in human colon and lung carcinoma cell lines has been reported previously, but its expression in primary tumours has not been described in detail. We have used the radio-immunoassay method to measure the specific content of immunoreactive TGF-alpha in the acid ethanol extracts of normal and cancerous tissues of human colon and lung. The average TGF-alpha content of colon carcinomas is 4 times that of the normal mucosa, and for non-small cell lung carcinomas it is twice that of the normal parenchyma. Because of variability in the TGF-alpha expression among individuals and in different segments of colon and lobes of lung, the ratio of TGF-alpha content of paired tumour and normal tissue was also calculated. On average, the tumour/normal ratio for colon carcinoma is higher than that for lung carcinoma. Although 55% of colon tumours show a ratio 4 times, or greater, only 33% of lung carcinomas demonstrate this ratio. The level of TGF-alpha in both colon and lung carcinomas does not correlate with histological type stage, grade nor degree of desmoplasia of these tumours. Northern blot analysis of total cellular RNA confirms the expression of an approximately 4.8 kb TGF-alpha mRNA in normal colonic mucosa and lung parenchyma. However, in contrast to the results of radio-immunoassay, significant over-expression of TGF-alpha mRNA is uncommon in primary human colon carcinomas.
PMCID: PMC1971446  PMID: 1698444
18.  Ion selectivity of the Vibrio alginolyticus flagellar motor. 
Journal of Bacteriology  1990;172(9):5236-5244.
The marine bacterium, Vibrio alginolyticus, normally requires sodium for motility. We found that lithium will substitute for sodium. In neutral pH buffers, the membrane potential and swimming speed of glycolyzing bacteria reached maximal values as sodium or lithium concentration was increased. While the maximal potentials obtained in the two cations were comparable, the maximal swimming speed was substantially lower in lithium. Over a wide range of sodium concentration, the bacteria maintained an invariant sodium electrochemical potential as determined by membrane potential and intracellular sodium measurements. Over this range the increase of swimming speed took Michaelis-Menten form. Artificial energization of swimming motility required imposition of a voltage difference in concert with a sodium pulse. The cation selectivity and concentration dependence exhibited by the motile apparatus depended on the viscosity of the medium. In high-viscosity media, swimming speeds were relatively independent of either ion type or concentration. These facts parallel and extend observations of the swimming behavior of bacteria propelled by proton-powered flagella. In particular, they show that ion transfers limit unloaded motor speed in this bacterium and imply that the coupling between ion transfers and force generation must be fairly tight.
PMCID: PMC213185  PMID: 2394685
19.  Comparison of ofloxacin, gentamicin, and tobramycin concentrations in tears and in vitro MICs for 90% of test organisms. 
Concentrations of three anti-infective agents in tear film were monitored after one topical application in rabbits. Ofloxacin concentrations exceeded the MIC for 90% of the organisms tested (MIC90) (gram-negative and gram-positive organisms) for 240 min. Tobramycin concentrations exceeded the MIC90 for 10 min. Gentamicin concentrations exceeded the MIC90 for 20 min for gram-positive organisms and 120 min for gram-negative organisms.
PMCID: PMC171882  PMID: 2221871
20.  Some chemical and physical properties of nisin, a small-protein antibiotic produced by Lactococcus lactis. 
Nisin is a small gene-encoded antimicrobial protein produced by Lactococcus lactis that contains unusual dehydroalanine and dehydrobutyrine residues. The reactivity of these residues toward nucleophiles was explored by reacting nisin with a variety of mercaptans. The kinetics of reaction with 2-mercaptoethane-sulfonate and thioglycolate indicated that the reaction pathway includes a binding step. Reaction of nisin at high pH resulted in the formation of multimeric products, apparently as a result of intramolecular and intermolecular reactions between nucleophilic groups and the dehydro residues. One of the nucleophiles had a pKa of about 9.8. The unique vinyl protons of the dehydro residues that give readily identifiable proton nuclear magnetic resonances were used to observe the addition of nucleophiles to the dehydro moiety. After reaction with nucleophiles, nisin lost its antibiotic activity and no longer showed the dehydro resonances, indicating that the dehydro groups had been modified. The effect of pH on the solubility of nisin was determined; the solubility was quite high at low pH (57 mg/ml at pH 2) and was much lower at high pH (0.25 mg/ml at pH 8 to 12), as measured before significant pH-induced chemical modification had occurred. High-performance liquid chromatography on a C18 column was an effective technique for separating unmodified nisin from its reaction products. The cyanogen bromide cleavage products of nisin were about 90% less active toward inhibition of bacterial spore outgrowth than was native nisin. These results are consistent with earlier observations, which suggested that the dehydro residues of nisin have a role in the mechanism of antibiotic action, in which they act as electrophilic Michael acceptors toward nucleophiles in the cellular target.
PMCID: PMC184764  PMID: 2119570
21.  An abnormality of the gene that encodes neutrophil Fc receptor III in a patient with systemic lupus erythematosus. 
Journal of Clinical Investigation  1990;86(1):341-346.
In the course of examining the structure and function of Fc receptors on peripheral blood cells of patients with systemic lupus erythematosus, we identified a patient whose neutrophils did not react with either monoclonal or polyclonal antibodies to Fc receptor III. However, neutrophils from the patient were comparable to neutrophils from healthy controls with respect to their expression of Fc receptor II, complement receptor 1, complement receptor 3, and the phosphatidylinositol-linked, complement regulatory protein, decay-accelerating factor. The abnormality of expression of Fc receptor III was limited to the patient's neutrophils (her natural killer cells reacted normally with anti-Fc receptor III antibodies), and was associated with abnormal recognition and binding of IgG-coated erythrocytes. Analysis of genomic DNA revealed evidence that failure of the patient's neutrophils to express Fc receptor III was most likely due to an abnormality of the gene that encodes this receptor.
PMCID: PMC296727  PMID: 1694867
22.  Functional analysis of elements affecting expression of the beta-actin gene of carp. 
Molecular and Cellular Biology  1990;10(7):3432-3440.
Regulatory regions of the beta-actin gene of the common carp (Cyprinus carpio) have been examined by linking upstream, 5'-flanking sequences and regions of the first intron to a bacterial chloramphenicol acetyltransferase (CAT) reporter gene. By analysis of the mRNA products and encoded CAT activity, we have identified four putative regions that influence expression: (i) a negative regulatory region 2,300 to 1,100 base pairs (bp) ahead of the gene; (ii) a proximal promoter element, containing the highly conserved CCAAT, CC(A/T)6GG, and TATA boxes, that is within the first 204 bp upstream of the initiation site; (iii) a negative element of 426 bp in the 5' region of the first intron; and (iv) a positive 304-bp element near the end of the first intron that contains highly conserved sequences found in all characterized beta-actin genes. The positive intron element is not a classical enhancer; it is position and orientation dependent, as has been observed in other housekeeping genes in vertebrates. Depending on the elements joined together, CAT gene expression can be modulated more than 500-fold in transfected mouse cells.
PMCID: PMC360779  PMID: 2355913
23.  Current approaches to vaccine preparation 
The Canadian Veterinary Journal  1990;31(3):181-189.
Numerous conventional vaccines for animal use are currently available, and many of these vaccines have been instrumental in the control of infectious diseases of major economic importance. A vaccine has even been instrumental in global eradication of smallpox, an important human disease. However, many of the current vaccines are deficient in efficiency, potency, or safety. It has been recognized that the conventional methodologies are a limitation to further vaccine development. Introduction of monoclonal antibodies, recombinant DNA, and protein engineering techniques has facilitated a rather rapid increase in the knowledge of pathogenetic mechanisms, as well as of protective antigens at the molecular level. This knowledge provides the basis for development of a new generation of vaccines. As a rule, these vaccines contain purified immunogens, or even isolated epitopes, identified and prepared by molecular biological techniques. The efforts to find better delivery systems and better adjuvants accompany the research on vaccines.
PMCID: PMC1480769  PMID: 17423533
24.  Serological evaluation of Escherichia coli-expressed human T-cell leukemia virus type I env, gag p24, and tax proteins. 
Journal of Clinical Microbiology  1990;28(6):1139-1142.
Three proteins (env, gag, and tax) encoded by the human T-cell leukemia virus type I (HTLV-I) genome were cloned and expressed in Escherichia coli. The env protein contained a substantial part of the gp46 domain and a majority of the p21e domain. The gag protein contained all of p24 and portions of p19 and p15. In addition to these two structural proteins, a full-length tax (p40X) construct was obtained. All three recombinant proteins were purified to near homogeneity. When used in an immunoblot assay, the three recombinant proteins detected antibodies in more HTLV-I antibody-positive patient sera than did the corresponding native proteins. Antibodies to at least two of these three different gene products were detected in 98.4% of adult T-cell leukemia patients, 100% of HTLV-I-associated myelopathy patients, 97.4% of asymptomatic carriers, and 94% of uncharacterized HTLV-I-positive patients. Antibody to recombinant tax was found in 4.9% of adult T-cell leukemia patients, whereas antibody to recombinant env could not be detected. These recombinant proteins from three different gene products may be useful in detecting or confirming the presence of antibodies to HTLV-I.
PMCID: PMC267892  PMID: 2199486
25.  Characterization of pp85, a target of oncogenes and growth factor receptors. 
Molecular and Cellular Biology  1990;10(6):2909-2915.
An 85,000-molecular-weight polypeptide (85K polypeptide) has previously been identified as a common substrate for tyrosine phosphorylation upon polyomavirus middle T transformation or upon platelet-derived growth factor stimulation of 3T3 cells. In each case, pp85 has an associated phosphatidylinositol kinase activity. The tissue distribution of pp85 was determined by middle T blotting experiments; the highest levels were found in brain, lung, and spleen tissues. High-resolution examination of 85K by isoelectric focusing demonstrated that there are at least 10 different forms. These were resolved into two families, 85K and 86K; the ratio of the two families changed in different cells. Similar forms were found for pp85 associated with pp60v-src. Individual species within each family differed by phosphorylation. Analysis of pp85 and pp86 by immunoprecipitation with anti-phosphotyrosine antibody showed increasing phosphorylation in response to middle T or pp60v-src transformation. The association of middle T with pp85 and pp60c-src was examined in pulse-chase experiments. Association of middle T with pp60c-src was slow and was accompanied by progressive modification of middle T. pp85 formed a dissociable complex with middle T within 2.5 min.
PMCID: PMC360653  PMID: 2160590

Results 1-25 (41)