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Biomedical subjects

C Little

Publications and source records attributed to C Little.

At least 55 records · Page 3Linked to original sources
PubMed1989

The effect of the plasticizers TBEP (tris-(2-butoxyethyl)-phosphate) and DEHP (di-(2-ethylhexyl)-phthalate) on beta-adrenergic ligand binding to alpha 1-acid glycoprotein and mononuclear leukocytes.

The plasticizers tris-(2-butoxyethyl)-phosphate (TBEP) and di-(2-ethylhexyl)-phthalate (DEHP) and the beta-adrenergic receptor-blockers [3H]-(-)-dihydroalprenolol ([3H]-(-)-DHA) and [3H]-(-)-CGP 12177 were tested for their ability to interact with beta-adrenergic binding to alpha 1-acid glycoprotein (AAG) and mononuclear leukocytes (MNL). The IC50-values, obtained by displacement of [3H]-(-)-DHA bound to AAG, were 3.5 nM, 2 microM and 4 microM for TBEP, (-)-alprenolol and DEHP, respectively. (+/-)-CGP 12177 had virtually no effect on radioligand binding to AAG. The [3H]-(-)-CGP 12177 binding to MNL consisted of beta-adrenergic receptor binding (Kd = 210 pM) and non-saturable binding. [3H]-(-)-DHA was bound to two different classes of binding sites on MNL, the beta-adrenergic receptors (Kd = 440 pM) and a secondary class of binding sites (Kd = 64 nM). (+/-)-CGP 12177 displaced about 30% of [3H]-(-)-DHA from MNL with an IC50-value of 190 pm. (-)-ALP displaced about 85% of total bound radioligand and gave a biphasic displacement curve with IC50-values of 320 pM and 690 mM, respectively. TBEP displaced a considerable fraction of [3H]-(-)-CGP 12177 and [3H]-(-)-DHA bound to MNL beta-adrenergic receptors, whereas DEHP had no effect. In contrast, DEHP caused displacement of [3H]-(-)-DHA from the MNL low affinity sites, but was a markedly less potent displacer compared to TBEP. The present study shows that TBEP and DEHP interact with beta-adrenergic transport proteins, non-specific tissue binding sites and beta-adrenergic receptors coupled to adenylate cyclase. Plasticizers may thus affect the biology and pharmacology of the beta-adrenergic signal system.

Adult↗
PubMed1989

High-resolution (1.5 A) crystal structure of phospholipase C from Bacillus cereus.

Both the phosphatidylinositol-hydrolysing and the phosphatidylcholine-hydrolysing phospholipases C have been implicated in the generation of second messengers in mammalian cells. The phosphatidylcholine-hydrolysing phospholipase C (PLC) from Bacillus cereus, a monomeric protein containing 245 amino-acid residues, is similar to some of the corresponding mammalian proteins. This, together with the fact that the bacterial enzyme can mimic the action of mammalian PLC in causing, for example, enhanced prostaglandin biosynthesis, suggests that B. cereus PLC can be used as a model for the hitherto poorly characterized mammalian PLCs. We report here the three-dimensional structure of B. cereus PLC at 1.5 A resolution. The enzyme is an all-helix protein belonging to a novel structural class and contains, at least in the crystalline state, three Zn2+ in the active site. We also present preliminary results from a study at 1.9 A resolution of the complex between PLC and inorganic phosphate (Pi) which indicate that the substrate binds directly to the metal ions.

Bacillus cereus↗
PubMed1989

Effect of 2,3-butanedione on human myeloperoxidase.

1. Myeloperoxidase is inhibited by various diketones that are recognized arginine reagents. 2. Although arginine residues in the enzyme were modified in both the light and the dark, enzyme inactivation occurred only in the presence of light. 3. Under conditions where diketones caused inactivation of myeloperoxidase, spectral studies indicated marked damage to the haem residues of the enzyme. 4. It was concluded that diketones serve simply as photosensitizers of visible light-induced inactivation of myeloperoxidase. 5. Studies on other haemoproteins indicated the great ease with which the presence of diketones sensitized haem residues for photodestruction.

Amino Acids↗
PubMed1989

Phospholipase C.

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Bacillus cereus↗
PubMed1989

A phase I-II study of sequential infusion VP-16 and cisplatin therapy in advanced lung cancer.

Although the etoposide (VP-16) and cisplatin combination has shown therapeutic activity in lung cancer, human results to date have not matched the expectation of synergism raised by animal model studies. Laboratory studies suggest that therapeutic synergism of etoposide and cisplatin may be related to factors of drug concentration, time of exposure, and sequencing. To pursue this question, we developed regimens of etoposide given by 72 h infusion in conjunction with sequential bolus or infusion cisplatin. Thirty-two patients were entered. Fourteen of 15 small-cell lung cancer patients had a response (CR, PR, regression) with a median survival of 321 days. Nine of 17 patients with non-small-cell lung cancer achieved a response, including two CRs. The median survival is 201 days. The major toxicity was myelosuppression. At the highest etoposide dosage tested, 42% of patients had leukopenia less than 2000/mm3. There were no treatment-related deaths. This new approach of combined etoposide and cisplatin therapy shows promising therapeutic activity against both small cell and non-small-cell lung cancer.

Adult↗
PubMed1989

Binding of prazosin and propranolol at variable alpha 1-acid glycoprotein and albumin concentrations.

1. The effect of variable alpha 1-acid glycoprotein (AAG) and albumin (HSA) concentrations on the binding of prazosin and propranolol was assessed in plasma after surgery and in mixtures of AAG/HSA with concentrations mimicking those found in vivo. 2. On the pre-operative day the binding of prazosin and propranolol was 94.8% and 89.0%, respectively and 97.3% and 93.2%, respectively, 5 days after surgery. 3. In solutions containing mixtures of highly purified AAG and HSA representing the pre-operative state, 88.6% and 83.9% binding of prazosin and propranolol was observed, whereas for solutions mimicking post-operative plasma, the equivalent values were 94.6% and 91.4%, respectively. 4. The ratios between bound and unbound concentrations of both drugs were closely correlated to the concentrations of AAG, but not to the concentrations of HSA. 5. The present study demonstrates that AAG is responsible for the binding variability of prazosin and propranolol in plasma from post-operative patients.

Aged↗
PubMed1989

Centrifugation of very freshly donated blood may yield platelets unstable to storage in the new generation of containers.

The stability of platelets stored in the second generation of container has been examined further. Platelet concentrates prepared in CPD anticoagulant were found to deteriorate fairly rapidly during storage if prepared from very freshly donated blood, whereas a more than 4-hour delay between donation and platelet isolation yielded platelet concentrates much more stable to storage. Delays of more than 2 h (relative to the time of donation) in the isolation of platelets led to decreases in the aggregation response to low or moderate doses of collagen. Platelets from blood collected into CPD adenine showed decreased aggregation responses relative to equivalent cells from blood collected into adenine-free anticoagulant. It seems that a decreased aggregation response is associated with improved storage properties.

Adenine↗
PubMed1989

Binding of prazosin to alpha 1-acid glycoprotein and albumin: effect of protein purity and concentrations.

Prazosin is extensively bound in human serum/plasma. In the present study a bound fraction of 93-95% was observed at 37 degrees for therapeutic drug concentrations. Both alpha 1-acid glycoprotein (AAG) and albumin (HSA) are established as transport proteins for prazosin, but their individual contribution to the extent and variability of protein binding in serum/plasma is unclear. The present study showed that AAG possesses one binding site per molecule with high affinity (Kd approximately 0.8 microM) for prazosin. HSA, essentially globulin-free, bound prazosin with lower affinity (Kd approximately 30 microM) with an average of 0.3 binding sites per molecule. However, less purified HSA, containing globulins, exhibited apparently higher affinity (Kd approximately 8 microM), but lower binding capacity (0.07 sites per molecule) for prazosin. In mixtures of highly purified proteins, the concentrations of AAG, and not HSA, determined the extent and variability of prazosin binding.

Dialysis↗
PubMed1989

Apparent phosphate retrieval system in Bacillus cereus.

Bacillus cereus secretes three different phospholipases C. We studied the effect of Pi levels in the growth medium on the production of these exoenzymes. Production of both phosphatidylcholine-preferring phospholipase C and sphingomyelinase C was repressed by Pi in the growth medium, whereas production of phosphatidylinositol phospholipase C was unaffected. We also found that B. cereus secretes a phosphate-repressed alkaline phosphatase activity. Together with a previously reported highly efficient, active uptake system for Pi, these three phosphate-repressed exoenzyme activities seem to be part of a phosphate retrieval mechanism that operates under growth-limiting concentrations of Pi. In natural soil systems, which are the natural habitats of B. cereus, the scarcity of Pi is the major growth-limiting factor. A phosphate-repressed metalloprotease activity was also detected in culture supernatants of B. cereus. It is unclear whether this exoenzyme activity also participates in the proposed phosphate-scavenging system.

Alkaline Phosphatase↗
PubMed1988

Nucleotide sequence and expression in Escherichia coli of the gene coding for sphingomyelinase of Bacillus cereus.

Bacillus cereus secretes phospholipases C, which hydrolyze phosphatidylcholine, sphingomyelin and phosphatidylinositol. A 7.5-kb HindIII fragment of B. cereus DNA cloned into Escherichia coli, with pUC18 as a vector, directed the synthesis of the sphingomyelin-hydrolyzing phospholipase C, sphingomyelinase. Nucleotide sequence analysis of the subfragment revealed that it contained two open reading frames in tandem. The upstream truncated open reading frame corresponds to the carboxy-terminal portion of the phosphatidylcholine-hydrolyzing phospholipase C, and the downstream open reading frame to the entire translational portion of the sphingomyelinase. The two phospholipase C genes form a gene cluster. As inferred from the DNA sequence, the B. cereus sphingomyelinase has a signal peptide of 27 amino acid residues and the mature enzyme comprises 306 amino acid residues, with a molecular mass of 34233 Da. The signal peptide of the enzyme was found to be functional in protein transport across the membrane of E. coli. The enzymatic properties of the sphingomyelinase synthesized in E. coli resemble those of the donor strain sphingomyelinase. The enzymatic activity toward sphingomyelin was enhanced 20-30-fold in the presence of MgCl2, and the adsorption of the enzyme onto erythrocyte membranes was accelerated in the presence of CaCl2.

Amino Acid Sequence↗
PubMed1988

Cloning and sequencing of the gene encoding the phosphatidylcholine-preferring phospholipase C of Bacillus cereus.

A synthetic oligodeoxynucleotide probe was used to clone the gene encoding the phosphatidylcholine-preferring phospholipase C of Bacillus cereus. The sequence of a 2050-bp restriction fragment containing the gene was determined. Analysis of the gene-derived amino acid (aa) sequence showed that this exoenzyme is probably synthesized as a 283-aa precursor with a 24-aa signal peptide and a 14-aa propeptide. The mature, secreted enzyme comprises 245 aa residues. Sonicates of Escherichia coli HB101 carrying the gene on a multicopy plasmid showed phospholipase C activity. This activity was inhibited by Tris, a known inhibitor of the B. cereus enzyme and also by antiserum raised against pure B. cereus phospholipase C. We conclude therefore that the gene is expressed in E. coli. The cloning and sequencing described here complete the first step toward using in vitro mutagenesis for investigations of the structure-function relationships of B. cereus phospholipase C.

Amino Acid Sequence↗
PubMed1988

Effect of centrifugation on the storage properties of platelets.

Some of the recommended centrifugation methods for the preparation of platelet concentrates may cause accelerated deterioration of platelets stored in second-generation containers. The deterioration is characterized by increasing pH, pO2 and decreasing pCO2, a high discharge of lactate dehydrogenase (LDH) and increasing amounts of small particles which have recently been shown to have platelet factor 3 activity [Solberg, C.; Osterud, B.; Little, C.: Thrombosis Res. 48: 559-565, 1987]. A short first centrifugation (3,270 g, 2 min 15 s) yielded platelets with better storage properties than platelet-rich plasma prepared with longer centrifugation times (2,200 g, 4 min 30 s and 1,100 g, 6 min). By using multivariate data analysis the effect of different platelet concentrations and metabolic parameters can be used to predict the discharge of LDH or the change in morphology.

Anticoagulants↗
PubMed1988

A cause-specific hazard rate analysis of prognostic factors among 199 adults with acute lymphoblastic leukemia: the Memorial Hospital experience since 1969.

Results of a multivariable analysis of prognostic factors are reported for 199 previously untreated adults with acute lymphoblastic leukemia (ALL). These patients have long-term follow-up, and the probability of cure is estimated at approximately 35%. The cause-specific hazard rate analysis found lower rates of achieving complete remission (CR) in patients with WBC greater than 10,000/microL, AUL (undifferentiated) morphology, and older age. Since these patients required additional time to respond, fewer of them actually achieved CR. Characteristics directly associated with a higher rate of death during induction therapy due to severe bone marrow suppression were low serum albumin concentration (less than or equal to 3.5 g/dL), age greater than 50 years, acute undifferentiated leukemia (AUL) morphology, low Karnofsky performance status, and weight loss greater than 5%. Factors associated with a higher rate of relapse were WBC greater than 20,000/microL, non-T cell ALL, age greater than 60 years, Ph' + ALL, and time to achieve CR greater than 5 weeks. These criteria were used to identify patients at high risk of relapse. In addition, the predictive value of high WBC was found to disappear by 18 months of continuous CR. Finally, the rate of death following first relapse was higher in patients with a short first remission duration, high percentage weight loss at initial diagnosis, and older age. In summary, factors associated with a higher rate of death during attempted induction (ie, low albumin, high percent weight loss, and poor performance status) had no association with the patient's ability to remain relapse-free. Conversely, factors correlating with more extensive or resistant disease (ie, high WBC, null or B cell ALL, or Ph' + ALL) showed no association with the ability to tolerate therapy. Thus, a less toxic but more effective induction regimen is needed for patients with a poor clinical status, whereas a more intensive form of therapy appears warranted for patients presenting with more extensive or resistant disease.

Adult↗
PubMed1987

Platelet storage lesion: formation of platelet fragments with platelet factor 3 activity.

Certain samples of stored platelet concentrates exhibited marked platelet factor 3 activity. This activity was associated almost completely with platelet fragments formed during storage. Platelet concentrates containing high levels of these fragments and hence high platelet factor 3 activity were characterized after storage by alkaline plasma pH values, high levels of extracellular lactate dehydrogenase activity and relatively low cell counts.

Blood Platelets↗