Characterization of human mononuclear cells using reduced pyridine nucleotide fluorescence and flow cytometry

Peripheral blood monocytes undergo an oxidative burst similar to that seen in neutrophils. The basis for this response appears to be an NAD(P)H oxidase that utilizes reduced NAD(P)H to form superoxide anion. We utilized the unique UV-stimulated fluorescence property of reduced pyridine nucleotides to analyze NAD(P)H utilization in monocytes. UV-stimulated fluorescence in mononuclear cell preparations indicated two populations of cells with the highly fluorescent cells having a Coulter volume consistent with that of monocytes. Dual laser analysis with monoclonal antibodies confirmed that these highly fluorescent cells are monocytes by showing them to be OKM1+, Leu DR+, and anti-monocyte 0.2+. Natural killer (NK) cells, as defined by Leu 7, were not found in this highly fluorescent population. Stimulation of mononuclear cells with phorbol myristate acetate caused a fluorescence loss indicative of NAD(P)H oxidation in monocytes but not in lymphocytes. Stimulation with suboptimal concentrations of PMA (1-5 ng/ml) resulted in a dose-dependent fluorescence loss in monocytes that occurred in an all-or-none fashion identical to the pattern observed in neutrophils. Simultaneous measurement of H2O2 production using dichlorofluorescein formation with NAD(P)H fluorescence indicates that oxidant production occurs in a graded manner. This method, then, provides a convenient way to study in single cells the metabolic events involved in depletion and replenishment of NAD(P)H during the oxidative burst and demonstrates an additional means by which to distinguish monocytes from lymphocytes using flow cytometry.     

Osteosarcomatosis

A review of the 690 cases of osteosarcoma in the radiographic file of the Armed Forces Institute of Pathology revealed 29 cases of "osteosarcomatosis" (multiple skeletal sites of osteosarcoma). Fifteen of these patients were 18 years old and under and manifested rapidly appearing, usually symmetric, sclerotic metaphyseal lesions. The remaining 14 patients were more than 18 years old and had fewer, asymmetric sclerotic lesions. In most patients (28 of 29), a radiographically dominant skeletal tumor was seen. Pulmonary metastases occurred in the majority of patients and were detected at the same time as the bone lesions. These 29 patients were studied with regard to demographic data and skeletal distribution and radiographic appearance of their lesions. As a result of the findings, a metastatic origin from a primary dominant osteosarcoma is favored over a multifocal origin as the basis for osteosarcomatosis. Osteosarcomatosis is more commonly encountered in the mature skeleton than has been previously recognized.     

Management of rectal injuries. Dogma versus practice

The current treatment of civilian rectal injuries stems from military practice. Five principles have evolved: 1) complete fecal diversion, 2) debridement and closure, 3) rectal stump irrigation, 4) presacral drainage, and 5) broad spectrum antibiotics. To assess our practice results, the records of 52 consecutive patients with rectal injury seen at Detroit Receiving Hospital from 1980-88 were reviewed. Etiologies were gunshot (40), shotgun (9), anal assault (2), and stab (1). There were no blunt injuries and no deaths. Treatment consisted of celiotomy (52), diverting colostomy (51), presacral drains (35), rectal stump irrigation (26), and primary closure (1). Broad spectrum antibiotics were administered in all patients. Despite lack of universal application of the "standard" principles, only five patients had postoperative complications and none were related to the rectal injury. Our results demonstrate that a single approach may not be justified, as excellent outcome was achieved with low morbidity and no mortality despite selective management. The universal application of colostomy, repair, irrigation, drainage, and antibiotics cannot be supported.     

Intravenous pulsed steroids in rheumatoid arthritis: a comparative dose study

This prospective, double blind study was undertaken to test the efficacy of intravenous "minipulse" (100 mg) methylprednisolone (MP) therapy versus standard pulse (1000 mg) MP therapy in rheumatoid arthritis (RA). Thirty-six patients with RA synovitis flares were randomized to receive either 100 or 1000 mg MP IV QD x 3 doses. These 2 universally comparable groups exhibited no statistically significant differences in their striking prompt and sustained clinical improvement. These data suggest that minipulse MP is as efficacious as conventional pulse MP in the treatment of RA flares.     

The effect of albuterol aerosol on fine-motor performance in children with chronic asthma

A double-blind, crossover trial was conducted to examine the effect of albuterol aerosol on the fine-motor performance of children with chronic asthma. Twenty subjects, 4 1/2 to 14 1/2 years of age, were each tested after both the administration of albuterol aerosol and a placebo. The fine-motor composite of the Bruininks-Oseretsky Test of Motor Proficiency was used to evaluate abilities in response speed, visual-motor control, and speed and dexterity. The degree of tremor was tested with a "steadiness" tester, and postural adjustment was recorded on a clinical rating scale. No significant differences were found in fine-motor skills after albuterol or placebo administration, but a significant increase in tremor as well as in postural adjustment occurred after albuterol. The effect of test order and several other patient characteristics on test scores may be related to these findings. Major educational adjustments to compensate for the increase in tremor are unnecessary.     

Phase I clinical and pharmacokinetic trial of dextran conjugated doxorubicin (AD-70, DOX-OXD)

Coupling of anthracyclines to high-molecular-weight carriers may alter drug disposition and improve antitumor effects. We have performed a clinical phase I trial of doxorubicin coupled to dextran (70000 m.w.). The drug was administered as single dose i.v. every 21–28 days. Thirteen patients have received a total of 24 courses (median 2; range 1–3). At the starting dose of 40 mg/m² doxorubicin equivalent (DOXeq), WHO grade IV thrombocytopenia was noted in 2/2 patients. WHO grade IV hepatotoxicity and WHO grade III cardiotoxicity were noted in a patient with preexisting heart disease. Five patients were treated with 12.5 mg/m² DOXeq. Maximal toxicity at this dose level was WHO grade III thrombocytopenia and local phlebitis (WHO grade II) in 1/5 patients, elevation of alkaline phosphatase (WHO grade III) and WHO grade III vomiting in another patient. Subsequently, five patients received 20 mg/m² DOXeq. Hepatotoxicity was noted in 5/5 patients (1 × WHO grade IV, 1 × WHO grade III). Thrombocytopenia was noted in 3/5 patients (1 × WHO grade IV, 2 × WHO grade III). At 12.5 mg/m² DOXeq, a patient diagnosed with a malignant fibrous histiocytoma had stable disease for 4 months. Pharmacokinetic analyses of total and free doxorubicin were performed in plasma and urine. The maximum peak plasma concentration (ppc) for total DOX was 12.3 g/ml at 40 mg/m² DOXeq. The area under the plasma concentration time curve (AUC) ranged from 28.83–80.21 g/ml^*h with dose-dependent elimination half lives (t_1/2: 0.02–0.87 h;_1/2: 2.69–11.58 h;_1/2: 41.44–136.58 h). We conclude that the maximal tolerated dose (MTD) of AD-70 using this schedule is 40 mg/m² DOXeq. The recommended dose for clinical phase II studies is 12.5 mg/m² DOXeq.Abbreviations ALT Alanine Aminotransferase - AST Aspartate Aminotransferase - DOX Doxorubicin - DOXeq Doxorubicin Equivalent - ECG Electrocardiogram - HPLC High Pressure Liquid Chromatography - LD₁₀ Lethal Dose for 10% of individuals - MTD Maximal Tolerated Dose - ppc Peak Plasma Concentration - WHO World Health Organisation     

The organization of the medical practice and implications for delivering quality care

The attention devoted to quality in health care has focused primarily on hospitals. With the shift in care from hospitals to outpatient clinics and physicians' offices, there has emerged a need to consider how quality should be approached in these settings. This article explores the structural barriers in the typical medical practice that must be removed in order to improve the many dimensions of quality. The typical practice must concentrate on enhancing clinical outcomes, patient satisfaction, and practice productivity. One effective strategy is to form teams of physicians and other staff to work on relevant issues. Another is to focus on how the practice can be defined to better meet the needs of patients.     

Lipoprotein(a) stimulates growth of human mesangial cells and induces activation of phospholipase C via pertussis toxin-sensitive G proteins

BACKGROUND: Renal disease is commonly associated with hyperlipidemia and correlates with glomerular accumulation of atherogenic lipoproteins, for example, lipoprotein(a) [Lp(a)], and mesangial hypercellularity. Specific binding of Lp(a) to mesangial cells and induction of c-myc and c-fos expression has been demonstrated. Therefore, in this study, we investigated a possible growth stimulatory effect and mode of action of Lp(a) in human mesangial cells. METHODS: Lp(a) was purified from the regenerate fluid of a dextran sulfate column-based low-density lipoprotein apheresis system. Human mesangial cells were isolated by a sequential sieving technique from patients undergoing tumor nephrectomy. DNA synthesis was measured by [3H]-thymidine incorporation. The intracellular calcium concentration ([Ca2+]i) was determined by Fura 2-fluorescence, and inositol 1,4,5-trisphosphate (1,4,5-IP3) concentration was measured by a radioreceptor assay. RESULTS: The data show that Lp(a) bound to the cells with a Kd of 17.0 micrograms/ml and increased DNA synthesis and cell proliferation. Lp(a) caused a rapid increase in 1,4,5-IP3 and [Ca2+]i via a pertussis toxin-sensitive mechanism. The phospholipase C (PLC) inhibitor U73122 abolished Lp(a)-induced cell proliferation. In contrast, vasopressin-induced increase in 1,4,5-IP3 and [Ca2+]i was pertussis toxin insensitive. CONCLUSION: This study revealed that Lp(a) stimulates growth of human mesangial cells. Lp(a)-induced signaling involves binding to a receptor and stimulation of PLC via Gi proteins. Stimulation of PLC appears to be essential for the growth stimulatory effect of Lp(a). Whether these effects of Lp(a) contribute to the pathophysiology of renal disease needs to be determined.