Improved detection of cytomegalovirus viremia in AIDS patients using shell vial and indirect immunoperoxidase methodologies.

One hundred twelve peripheral blood specimens were tested for the presence of cytomegalovirus (CMV) by the tube culture indirect immunoperoxidase (TC-IPA) procedure, the shell vial assay [shell vials were pre- and postinoculation treated with medium containing 2 of 10% fetal bovine serum (FBS) or 100 micrograms% cortisol] (SV-IFA), and conventional (MRC-5) tube cultures (TC-CPE). CMV was detected in 25 (22%) of the 112 specimens tested by at least one of these methods. The detection/isolation of CMV among the 25 positive specimens in shell vials maintained with 2% FBS, 100 micrograms% cortisol + 2% FBS, and 10% FBS was 36, 44, and 52%, respectively. Detection/isolation of the virus from blood by TC-IPA and TC-CPE was 52% and 76%, respectively. A significantly greater CMV detection rate occurred using TC-CPE compared to SV-IFA treated with medium supplemented with an FBS concentration of 2% (P = .0132), but not medium containing the higher serum supplement or the glucocorticoid (P greater than .05). Differences in the identification of a CMV viremia were observed by IPA, SV-IFA, and TC-CPE methodologies on a patient-to-patient basis, denoting the necessity of incorporating each methodology into the CMV screening panel. Demographic analysis of 82 AIDS patients showed a CMV viremia prevalence of 9% (2/28) in intravenous drug users, 57% (27/47) in homosexual patients, and 22% (2/9) in heterosexual and transfusion patients. Overnight (24 hr) storage of whole blood at 4 or 24 degrees C, respectively, reduced CMV recovery by 40% and 65%, when tested by TC-CPE.(ABSTRACT TRUNCATED AT 250 WORDS)     

Liver disease and the PI ElembergM phenotype of alpha 1-antitrypsin

The purpose of this article is to document clinical and pathologic observations concerning liver disease associated with the PI ElembergM phenotype of alpha 1-antitrypsin. Deposits of alpha 1-antitrypsin that were periodic acid-Schiff positive and stained with an antiserum to alpha 1-antitrypsin were found in the liver of a markedly jaundiced, terminally ill patient with Stage IV primary biliary cirrhosis. A biopsy performed three years earlier failed to reveal alpha 1-antitrypsin deposits. The phenotype PI ElembergM was verified by both acid starch gel electrophoresis and isoelectric focusing in agarose. The deposits of alpha 1-antitrypsin in the liver appear to be a consequence of the patient's disease and age and not due to an association with the PI*Elemberg allele. Accumulation of alpha 1-antitrypsin in the liver of this patient may be due to an accelerated synthesis of this protease inhibitor exceeding the liver's capacity for glycosylation or other steps in its secretion.     

Human polymorphonuclear leukocyte interaction with cyclosporine A.

The effects of cyclosporin A (cyA) on human polymorphonuclear leukocyte function, including phagocytosis, its associated metabolic burst, bacterial killing, and chemotaxis, were evaluated. Both Pseudomonas aeruginosa and Staphylococcus aureus were used as test particles. Polymorphonuclear leukocytes incubated in 10 and 50 micrograms of cyA per ml behaved normally with respect to phagocytosis and hexose monophosphate shunt activity at both high (10:1) and low (2:1) S. aureus/leukocyte ratios. With a small bacterial inoculum, killing of S. aureus was slightly impaired at early times only in the presence of 50 micrograms of cyA per ml. Phagocytosis and killing of P. aeruginosa with both large and small bacterial inocula were unaffected by cyA. Chemotaxis was within normal limits under all conditions. In addition, polymorphonuclear leukocytes from four renal transplant recipients receiving both cyA and prednisone demonstrated normal metabolic bursts and bacterial killing with both small and large inocula of S. aureus.     

Comparison of two rapid culture methods for detection of cytomegalovirus in clinical specimens.

The conventional virus isolation technique was compared with a 24-h shell vial centrifugation culture technique and with a 48-h tube culture method for the detection of cytomegalovirus (CMV) in MRC-5 cells. Of 200 clinical specimens tested, 41 were positive for CMV by at least one procedure. Indirect immunoperoxidase staining was positive for 32 (78.0%) of 41 specimens in the tube culture method and for 30 (73.2%) of 41 specimens in the shell vial centrifugation method. CMV was detected in 23 (56.1%) of 41 specimens by the development of cytopathic effect within 14 days.     

Pharmacological evidence for stimulation of growth hormone secretion by a central noradrenergic system in dogs

The role of brain catecholamines in the regulation of growth hormone secretion was investigated in pentobarbital-anesthetized dogs by using drugs which modify the function of adrenergic neurons and receptors. Intravenous administration of L-dopa produced a prompt, statistically significant increase in plasma growth hormone concentration. This response was not significantly reduced by blockade of peripheral dopa decarboxylase activity with carbidopa. Clonidine, an alpha-agonist which penetrates the brain, increased plasma growth hormone secretion. Norepinephrine, epinephrine, dopamine and isoproterenol, catecholamines which do not penetrate the blood-brain barrier, failed to affect plasma growth hormone concentration when administered intravenously. Apomorphine did not produce a statistically significant increase in plasma growth hormone concentration when administered directly into the the third ventricle, and pimozide failed to abolish the increase in plasma growth hormone produced by L-dopa. The increase in plasma growth hormone concentration produced by intravenous L-dopa and clonidine was prevented by administration of phentolamine or phenoxybenzamine directly into the third ventricle. The response to L-dopa was also abolished by intraventricular procaine. In dogs in which central beta-adrenergic blockade was produced by intraventricular L-propranolol, the growth hormone response to L-dopa was greater than it was in control dogs treated with intraventricular D-propranolol. The data indicate that in pentobarbital anesthetized dogs, the increase in growth hormone secretion produced by L-dopa is mediated by norepinephrine, rather than dopamine, that the site of action of the norepinephrine is central, above the median eminence and inside the 'blood-brain barrier', and that the norepinephrine acts via alpha-adrenergic receptors.     

Periodic limb movements in sleep in community-dwelling elderly.

The prevalence of periodic limb movements in sleep (PLMS) in a randomly selected elderly sample is reported. In San Diego, 427 elderly volunteers aged 65 yr and over were recorded in their homes. Forty-five percent had a myoclonus index, MI greater than or equal to 5. Correlates of PLMS included dissatisfaction with sleep, sleeping alone and reported kicking at night. Although statistically significant, the strengths of the associations between interview variables and myoclonus indices were all small. No combination of demographic variables and symptoms allowed highly reliable prediction of PLMS.     

Overproduction and purification of the luxR gene product: Transcriptional activator of the Vibrio fischeri luminescence system

Expression of Vibrio fischeri luminescence genes requires an inducer, termed autoinducer, and a positive regulatory element, the luxR gene product. A plasmid containing luxR under control of a tac promoter was engineered to overproduce this gene product. The overproduced luxR gene product was active in vivo, and its apparent monomeric molecular weight was indistinguishable from that of the protein encoded by luxR under control of its own promoter (M(r) 27,000). The new tac-luxR construct directed the synthesis of large quantities of the luxR gene product in induced Escherichia coli cells lacking other lux genes. In the presence of the other lux genes, overexpression of the tac-luxR construct was not detected. The overproduced luxR gene product, which formed cytoplasmic inclusion bodies, was purified and used in subsequent studies. Nonequilibrium pH gradient electrophoresis indicated that the protein was basic, and the amino-terminal 15 amino acids were sequenced. DNA-binding activity was detected by membrane filter binding assays; under the conditions used, the binding was not lux DNA-specific. Binding of tritium-labeled autoinducer to the luxR gene product was not detected, and autoinducer enhancement of the binding of the luxR gene product to DNA could not be detected reproducibly.     

Osteopetrosis: MR characteristics at 1.5 T

Four patients with proved osteopetrosis (three with the infantile malignant form and one with the benign form) were examined with magnetic resonance imaging at 1.5 T. All patients were studied in the coronal and sagittal planes using both short and long repetition time/echo time sequences. The infantile malignant form was characterized by a complete lack of signal from the marrow alternating with a signal intensity equivalent to that of the intervertebral disks, resulting in a "stepladder" appearance. In the benign form or after successful marrow transplantation in the infantile malignant form, intermediate or high signal intensity in the vertebrae was noted, suggesting the presence of some marrow elements.     

Phase I clinical and pharmacokinetic study of cyclophosphamide administered by five-day continuous intravenous infusion

Summary A total of 14 patients, 7 male and 7 female, received in all 21 evaluable courses of cyclophosphamide administered by 5-day continuous infusion. Cyclophosphamide doses were escalated from 300 to 400 mg/m² per day for 5 days and repeated every 21–28 days. The patient population had a median age of 55 years (range 38–76) and a median Karnofsky performance status of 80 (range 60–100). Only 1 patient had not received prior therapy; 5 patients had received only prior chemotherapy, 1 had received only prior radiotherapy, and 7 had received both. Tumor types were gastric (1), lung (2), colon (4), urethral adenocarcinoma (1), cervical (2), chondrosarcoma (1), melanoma (1), uterine leiomyosarcoma (1), and pancreatic (1). The dose-limiting toxicity was granulocytopenia, with median WBC nadir of 1700/l (range 100–4800) in 8 heavily pretreated patients treated at 350 mg/m² per day for 5 days. One patient without heavy prior treatment received two courses at 400 mg/m² and had WBC nadirs of 800/l and 600l. WBC nadirs occurred between days 9 and 21 (median 14). Drug-induced thrombocytopenia occurred in only one patient (350 mg/m² per day, nadir 85000/l). Neither hyponatremia nor symptomatic hypoosmolality was observed. Radiation-induced hemorrhagic cystitis may have been worsened in one patient. Nausea and vomiting were mild. Objective remissions were not observed. The maximum tolerated dose for previously treated patients is 350 mg/m² per day for 5 days. This dose approximates the doses of cyclophosphamide commonly used with bolus administration. Plasma steady-state concentrations (Css) of cyclophosphamide, measured by gas liquid chromatography, were 2.09–6.79 g/ml. Steady state was achieved in 14.5±5.9 h (mean ±SD). After the infusion, cyclophosphamide disappeared from plasma monoexponentially, with a t^1/2 of 5.3±3.6 h. The area under the curve of plasma cyclophosphamide concentrations versus time (AUC) was 543±150 g/ml h and reflected a cyclophosphamide total-body clearance (CL_TB) of 103±31.6 ml/min. Plasma alkylating activity, assessed by p-nitrobenzyl-pyridine, remained steady at 1.6–4.3 g/ml nor-nitrogen mustard equivalents. Urinary excretion of cyclophosphamide and alkylating activity accounted for 9.3%±7.6% and 15.1%±2.0% of the administered daily dose, respectively. The t^1/2 and AUC of cyclophosphamide associated with the 5-day continuous infusion schedule are similar to those reported after administration of cyclophosphamide 1500 mg/m² as an i.v. bolus. The AUC of alkylating activity associated with the 5-day continuous infusion of cyclophosphamide is about three times greater than the AUC of alkylating activity calculated after a 1500-mg/m² bolus dose of cyclophosphamide. Daily urinary excretions of cyclophosphamide and alkylating activity associated with the 5-day continuous infusion schedule are similar to those reported after bolus doses of cyclophosphamide.