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The results of this controlled study of the treatment of 57 patients with Gilles de la Tourette's syndrome suggested that both haloperidol and pimozide were more effective than placebo, but that haloperidol was slightly more effective than pimozide. Adverse effects occurred more frequently with haloperidol vs placebo than with pimozide vs placebo, but the frequency was not significantly different for haloperidol compared with pimozide. Clinically significant cardiac effects did not occur at a maximum dosage of 0.3 mg/kg or 20 mg/d for pimozide and 10 mg/d for haloperidol. However, the QTc interval was prolonged during pimozide treatment compared with that during haloperidol treatment, although the values for both medications were not in an abnormal range.

Clinical assessment (disease activity, severity, and extraarticular manifestations) of 101 rheumatoid arthritis patients was correlated with several laboratory tests, including 5 immune complex assays: the bovine conglutinin, 125I-Clq binding, monoclonal rheumatoid factor inhibition, Raji cell, and staphylococci binding assays. Elevated disease activity indices were most closely associated with the presence of immune complexes detected by the 125I-Clq and staphylococci binding assays. There were significant but weak correlations between the level of disease activity and the level of immune complexes as measured by the bovine conglutinin, 125I-Clq binding, Raji cell, and staphylococci binding assays. Articular disease severity, as measured by anatomic stage, was discriminated by the bovine conglutinin, monoclonal rheumatoid factor inhibition, and staphylococci binding assays. Extraarticular manifestations were best discriminated by the Raji cell and staphylococci binding assays. We concluded that the sensitivity, specificity, predictive value, and overlap of the associations were not sufficient to warrant their wide use for the diagnosis and management of rheumatoid arthritis in individual patients. Conversely, the 125I-Clq and staphylococci binding assays were as good as the erythrocyte sedimentation rate and the IgG rheumatoid factor test (the 2 best of many examined) in assessing disease activity. Further prospective studies with these assays will determine their usefulness in following rheumatoid arthritis for a prolonged period.

Membrane marker and cytochemical analyses were carried out on the abnormal cells from 70 adult acute leukemia patients. Such information may (1) supplement standard morphology and serve as a basis for a new classification scheme for acute leukemia, and (2) characterize the surface membranes of granulocyte, lymphocyte, and monocyte "progenitors." Classification of acute lymphoid leukemias solely on the basis of morphology was unsatisfactory. The presence or absence of T- or B-cell markers was helful in classifying lymphoid leukemias. Monocyte progenitors were characteristically nonspecific esterase positive and Fc-receptor and membrane-IgG positive, but poorly phagocytic. Promyelocytes and myelocytes were frequently Fc-receptor positive and consequently positive for surface immunoglobulin. Myeloblasts were characteristically Fc-receptor negative. We conclude that surface membrane markers are essential in diagnosing lymphoid leukemias and helpful in nonlymphoid acute leukemias, and that cytochemistry is essential in delineating lymphoid from nonlymphoid leukemias and in subclassifying the latter.

The effects of the Trendelenburg (TREND) position and passive straight leg raising (PLR) on cardiopulmonary performance in 18 anesthetized patients undergoing myocardial revascularization were studied with a two-dimensional transesophageal echocardiography probe and a thermodilution right ventricular ejection fraction (RVEF) pulmonary artery catheter. The TREND position (at 20 degrees) and PLR (at 60 degrees) were studied in relation to the level-supine position in random order. At 3 min, the TREND caused significant (p less than .05) decreases in heart rate (62 +/- 9 to 58 +/- 10 [SD] beat/min) and RVEF (0.48 +/- 0.11 to 0.44 +/- 0.10), and significant increases in mean arterial pressure (77 +/- 11 to 82 +/- 11 mm Hg), mean pulmonary artery pressure (16 +/- 4.3 to 19 +/- 5.5 mm Hg), wedge pressure (11 +/- 4 to 13 +/- 4 mm Hg), cardiac index (CI) (2.36 +/- 0.79 to 2.52 +/- 0.93 L/min.m2), right ventricular end-systolic volume index (44 +/- 21 to 58 +/- 21 ml/m2), right ventricular end-diastolic volume index (83 +/- 24 to 102 +/- 22 ml/m2), and shunt fraction (0.16 +/- 0.06 to 0.19 +/- 0.06). CVP and left ventricular areas did not change significantly. PLR had similar effects as the TREND position, except CI did not change significantly. Thus, the TREND and PLR resulted in minor hemodynamic improvement with right ventricular dilation, decreased RVEF, and impaired oxygenation in the anesthetized cardiac surgical patient.

Limited proteolysis of calcineurin, the Ca2+/calmodulin-stimulated protein phosphatase, with clostripain is sequential and defines four functional domains in calcineurin A (61 kDa). In the presence of calmodulin, an inhibitory domain located at the carboxyl terminus is rapidly degraded, yielding an Mr 57,000 fragment which retains the ability to bind calmodulin but whose p-nitrophenylphosphatase is fully active in the absence of Ca2+ and no longer stimulated by calmodulin. Subsequent cleavage(s), near the amino terminus, yield(s) an Mr 55,000 fragment which has lost more than 80% of the enzymatic activity. A third, slower, proteolytic cleavage in the carboxyl-terminal half of the protein converts the Mr 55,000 fragment to an Mr 42,000 polypeptide which contains the calcineurin B binding domain and an Mr 14,000 fragment which binds calmodulin in a Ca2+-dependent manner with high affinity. In the absence of calmodulin, clostripain rapidly severs both the calmodulin-binding and the inhibitory domains. The catalytic domain is preserved, and the activity of the proteolyzed 43-kDa enzyme is increased 10-fold in the absence of Ca2+ and 40-fold in its presence. The calcineurin B binding domain and calcineurin B appear unaffected by proteolysis both in the presence and in the absence of calmodulin. Thus, calcineurin A is organized into functionally distinct domains connected by proteolytically sensitive hinge regions. The catalytic, inhibitory, and calmodulin-binding domains are readily removed from the protease-resistant core, which contains the calcineurin B binding domain. Calmodulin stimulation of calcineurin is dependent on intact inhibitory and calmodulin-binding domains, but the degraded enzyme lacking these domains is still regulated by Ca2+.

A total of 62 patients with unstable pertrochanteric fractures of the neck of the femur was subjected to either nail plating with 130 degrees neck-shaft angle or valgus osteotomy using a neck-shaft angle of at least 140 degrees. The patients were assessed in terms of ability to walk and stability of fracture fixation. A clear superiority of valgus osteotomy over the controls was demonstrated in terms of fracture fixation, but the difference in terms of ability to walk was less remarkable.

In this paper we describe the isolation of a novel strain of Candida albicans which is a mycelium at ambient temperatures. Mutagenesis of C. albicans ATCC 10261 with N-methyl-N-nitro-N-nitrosoguanidine followed by plating on solid media at 28 degrees C yielded colony morphology variants which were characterized by a raised, rough-surfaced colony of irregular outline in marked contrast to the flat, shiny circular colonies of the parental 10261 strain. One mutant colony, hOG301, was studied in detail. Strain hOG301 was stable and exhibited mycelial morphology over a wide temperature range (5 to 40 degrees C) in several media. The hyphae comprising hOG301 mycelium were examined by light microscopy, scanning electron microscopy, and transmission electron microscopy and showed morphological features described in the literature as being typical of both true hyphae and pseudohyphae. In contrast to 10261, hOG301 was not pathogenic after intraperitoneal injection in mice. This is the first report of a mycelial C. albicans that is stable at ambient temperatures.

Most studies of skinned smooth muscle have been performed in whole tissue preparations. In this study, we report the development of a chemically skinned single smooth muscle cell preparation from the toad, Bufo marinus, stomach. Isolated smooth muscle cells were skinned using saponin. The effect of various ionic environments (i.e., changing free Ca2+ and MgATP2-) on skinned cell contractile response was assessed by measuring cell lengths from populations of cells using a computer-assisted length-measuring system. Comparison of cell length histograms were used to determine the extent of cell shortening in response to a given ionic perturbation. Once skinned, the single cells shortened with a sensitivity to free calcium (ED50 = 1.5 microM Ca2+) that was three orders of magnitude lower than potassium depolarized cells (ED50 = 1.5 mM Ca2+). In addition to the calcium sensitivity, the effect of free MgATP2- on the extent of cell shortening was investigated. The extent of cell shortening was dependent on free MgATP2- with the maximum shortening response occurring at MgATP2- greater than 1 mM.

Hemoglobin Atlanta, alpha 2 beta 2 75 Leu-Pro (E19), has been found in several members of three generations of a Caucasian family living in metropolitan Atlanta. The abnormal hemoglobin is one of the nine unstable variants in which either a leucyl or an alanyl residue is replaced by a prolyl residue. These substitutions have been observed in the B, E, F, and G helixes of the beta chain and in the H helix of the alpha-chain. Hemoglobin Atlanta heterozygotes are mildly affected by the presence of this unstable hemoglobin.

Five lots (100 ml or more) of heterologous antiserums specific for human T lymphocytes were prepared using human or Rhesus monkey thymocytes as immunogens. After appropriate adsorptions, these antiserums reacted by immunofluorescence with 68% of human peripheral blood mononuclear cells and 98% of human thymocytes, with E-rosette--positive cells but not with EAC-rosette--positive cells or five human B-lymphoblastoid-cell lines. Blocking experiments showed that Rhesus monkey thymocytes share thymic antigenic determinant(s) with humans. E-rosette receptors modulated independently from T-cell heteroantigens. Non-E--rosetting neoplastic T cells were identified in several patients with lymphoproliferative malignancies. Applying both the E-rosette assay and the anti-T-cell serum provides a better method of defining the biologic properties of normal and neoplastic T lymphocytes. Standardization of immunofluorescent conjugates for human T- or B-cell enumeration is simplified if large lots of well-characterized antiserums are available.

The reasons for a different incidence of osteoporotic fractures in white and black women are unknown. Previous racial comparisons of bone mass have been limited by racial differences in body weight and socioeconomic, health, and nutritional status. This cross-sectional study examined bone density in 105 black and 114 white healthy nonobese women, 24-65 yr old, using dual photon absorptiometry of the lumbar spine and single photon absorptiometry of the distal radius. Bone density at both sites was higher in blacks at all ages than in whites. When adjusted for age and body mass index, mean bone density was 6.5% higher in blacks at both spine and radius (P less than 0.0001). The cross-sectional rate of decline of vertebral bone density was similar between races; however, radial density increased 3.8%/decade (P = 0.03) in premenopausal blacks under age 46 yr, while it declined 3.2%/decade (P = 0.09) in premenopausal whites. The racial difference in slopes in these premenopausal women is significant (P = 0.002). These findings suggest that attainment of higher peak bone mass and delayed onset of bone loss contribute to the lower incidence of osteoporotic fractures in black women.

The glycogen-associated form of protein phosphatase-1 (PP-1G) is a heterodimer comprising a 37-kDa catalytic (C) subunit and a 161-kDa glycogen-binding (G) subunit, the latter being phosphorylated by cAMP-dependent protein kinase at two serine residues (site 1 and site 2). Here the amino acid sequence surrounding site 2 has been determined and this phosphoserine shown to lie 19 residues C-terminal to site 1 in the primary structure. The sequence in this region is: (sequence; see text) At physiological ionic strength, phosphorylation of glycogen-bound PP-1G was found to release all the phosphatase activity from glycogen. The released activity was free C subunit, and not PP-1G, while the phospho-G subunit remained bound to glycogen. Dissociation reflected a greater than or equal to 4000-fold decrease in affinity of C subunit for G subunit and was readily reversed by dephosphorylation. Phosphorylation and dephosphorylation of site 2 was rate-limiting for dissociation and reassociation of C subunit. Release of C subunit was also induced by the binding of anti-site-1 Fab fragments to glycogen-bound PP-1G. At near physiological ionic strength, PP-1G and glycogen concentration, site 2 was autodephosphorylated by PP-1G with a t0.5 of 2.6 min at 30 degrees C, approximately 100-fold slower than the t0.5 for dephosphorylation of glycogen phosphorylase under the same conditions. Site 2 was a good substrate for all three type-2 phosphatases (2A, 2B and 2C) with t0.5 values less than those toward the alpha subunit of phosphorylase kinase. At the levels present in skeletal muscle, the type-2A and type-2B phosphatases are potentially capable of dephosphorylating site 2 in vivo within seconds. Site 1 was at least 10-fold less effective than site 2 as a substrate for all four phosphatases. In conjunction with information presented in the following paper in this issue of this journal, the results substantiate the hypothesis that PP-1 activity towards the glycogen-metabolising enzymes is regulated in vivo by reversible phosphorylation of a targetting subunit (G) that directs the C subunit to glycogen--protein particles. The efficient dephosphorylation of site 2 by the Ca2+/calmodulin-stimulated protein phosphatase (2B) provides a potential mechanism for regulating PP-1 activity in response to Ca2+, and represents an example of a protein phosphatase cascade.

Purpose: Bacteremia in the elderly is associated with a different clinical course and a higher mortality rate when compared with that in younger age groups. In order to examine these issues in the aged, we reviewed the clinical course and factors involved in the outcome of 100 episodes of bloodstream infections in patients over 65 years of age., Patients and Methods: The hospital records of all patients over 65 years of age at The Mount Sinai Hospital with a positive blood culture result during the period October 1984 to October 1986 were reviewed. Place of residence before hospital admission, site of acquisition of infection, source of bloodstream infection, and microorganism were analyzed. Antimicrobial therapy was defined as appropriate if initial therapy included one agent to which the isolate was sensitive, or inappropriate if the isolate was resistant. The following factors affecting survival were analyzed: age, sex, underlying diseases, clinical parameters on admission, white blood cell count, mental status, source of infection, microorganism isolated, antibiotic toxicity, and appropriate versus inappropriate antibiotic therapy., Results: Most patients were female (63 percent), were febrile (90 percent), had an altered mental status (52 percent), and had a neutrophilic response (61 percent). Eighty-three percent of patients were admitted from the community (home), 14 percent were from long-term-care facilities, and 3 percent were transferred from other hospitals. Fifty percent of infections were nosocomial, and 44 percent were community (home and nursing home)-acquired. Gram-negative organisms accounted for 60 percent of isolates, with Escherichia coli (22 percent) and Klebsiella species (11 percent) predominating; 30 percent were gram-positive organisms, with Staphylococcus aureus (13 percent) and Streptococcus faecalis (10 percent) the most common. The overall survival was 60 percent; the survival rate was 65.8 percent for community-acquired (home) bacteremia, 75 percent for nursing home-acquired bacteremia, and 52.8 percent for hospital-acquired bacteremia. Survival for gram-negative isolates was 65 percent, versus 51.7 percent for gram-positive isolates. Survival was greatest in patients whose source of bacteremia was either the genitourinary tract (70 percent) or an intravascular device (78 percent) and poorest in patients with lower respiratory tract source (42 percent); all three patients with endocarditis died. Increased survival was observed in patients treated with appropriate antimicrobial agents regardless of age, source of infection, or bloodstream isolates.(ABSTRACT TRUNCATED AT 400 WORDS)

A morphometric analysis of Candida albicans yeast cells utilizing scanning electron microscopy showed that the cell volume and the DNA content of a tetraploid strain (derived by cell fusion) were 2.4 to 3.0 and 2.0 times, respectively, those of the progenitor diploid strain, ATCC 10261. The pathogenicities of both strains were similar.

A retrospective study conducted from 1975-1987 at Vanderbilt University Medical Center and affiliated hospitals identified 28 patients, ranging in age from 22 to 74 years, who were noted to have "nonfunctioning" adrenocortical tumors. Eighteen (64%) were men, and ten (36%) were women. A left adrenal mass was present in 16 (57%) patients, a right in 11 (39%) patients, and one (4%) patient had bilateral adrenal enlargements. Nineteen patients underwent an adrenalectomy. Ten were found to have adrenocortical adenomas, ranging is size from 2.5-4.0 cm in greatest diameter. Three were adrenocortical carcinomas (3.0, 5.5, and 8.0 cm). A necrotic mass (14.0 cm) was found in one patient. Two patients had myelolipoma, one had ganglioneuroma, and one had a suspected primary melanoma. Five patients were not operated upon and were followed by serial computed tomographic scans for variable periods; two died of unrelated cardiac problems, and one was lost to follow-up. Three patients underwent biopsy with benign pathology. The data indicate a high incidence of adenoma and carcinoma in patients with nonfunctioning adrenal tumors that measure more than 3.0 cm in diameter. These tumors have the potential to progress to malignant degeneration without any recognizable clinical or endocrine abnormalities. The authors thus conclude that all patients with "nonfunctioning" adrenal tumors that are larger than 3.0 cm in diameter should undergo surgical exploration and excision.

Radiation injury to the gastrointestinal tract is an infrequent but major clinical problem. Results of previous studies have shown that prostaglandins provide cytoprotection of the gastrointestinal mucosa against a variety of noxious agents, although, prior to this study, the protection against radiation exposure had not been documented. Exteriorized segment of Sprague-Dawley rat ileum was radiated with 10 and 15 Gy (137Cs). One group of rats was pretreated with prostaglandin E2 one hour before and 24 hours after radiation injury. The rats were sacrificed three and five days following radiation injury. Morphometric measurement of mucosal thickness, villous height, crypt of Lieberkühn height and number of mitosis per square millimeter swath of tissue were analyzed. Also, 125IUdR and 3HTdR were injected in a group of rats radiated with 15 Gy (137Cs). 125IUdR counts per minute per milligram of dry weight and 3HTdR labeled cells were counted and analyzed. The morphometric measurements and radioactive labeled tissue counts suggest that prostaglandin E2 has a cytoprotective effect upon irradiated rat ileum. Speculations about the possible mechanism and usefulness of this observation are included.

The kinetics of ATP hydrolysis and cation effects on ATPase activity in plasma membrane from Candida albicans ATCC 10261 yeast cells were investigated. The ATPase showed classical Michaelis-Menten kinetics for the hydrolysis of Mg X ATP, with Km = 4.8 mM Mg X ATP. Na+ and K+ stimulated the ATPase slightly (9% at 20 mM). Divalent cations in combination with ATP gave lower ATPase activity than Mg X ATP (Mg greater than Mn greater than Co greater than Zn greater than Ni greater than Ca). Divalent cations inhibited the Mg X ATPase (Zn greater than Ni greater than Co greater than Ca greater than Mn). Free Mg2+ inhibited Mg X ATPase weakly (20% inhibition at 10 mM). Computed analyses of substrate concentrations showed that free Zn2+ inhibited Zn X ATPase, mixed (Zn2+ + Mg2+) X ATPase, and Mg X ATPase activities. Zn X ATP showed high affinity for ATPase (Km = 1.0 mM Zn X ATP) but lower turnover (52%) relative to Mg X ATP. Inhibition of Mg X ATPase by (free) Zn2+ was noncompetitive, Ki = 90 microM Zn2+. The existence of a divalent cation inhibitory site on the plasma membrane Mg X ATPase is proposed.

Two experiments demonstrated that self-perceptions and social perceptions may persevere after the initial basis for such perceptions has been completely discredited. In both studies subjects first received false feedback, indicating that they had either succeeded or failed on a novel discrimination task and then were thoroughly debriefed concerning the predetermined and random nature of this outcome manipulation. In experiment 2, both the initial outcome manipulation and subsequent debriefing were watched and overheard by observers. Both actors and observers showed substantial perseverance of initial impressions concerning the actors' performance and abilities following a standard "outcome" debriefing. "Process" debriefing, in which explicit discussion of the perseverance process was provided, generally proved sufficient to eliminate erroneous self-perceptions. Biased attribution processes that might underlie perserverance phenomena and the implications of the present data for the ethical conduct of deception research are discussed.

In N-acetylglucosamine induced germ tube formation by Candida albicans, multiple (up to five) protuberances appeared within 90 min at 37 degrees C on each yeast cell. The protuberances were extensions of the cytosol and contained vesiclelike structures. Usually only one protuberance subsequently developed into a germ tube. The germ tubes emanated from all aspects of the cell surface but seldom from the budding (long axis) poles. Pseudohyphae, which originated from the budding pole, exhibited a marked constriction at the site of emergence and were 0.6-2.5 microns in diameter compared with a diameter of 0.6-0.8 micron for germ tubes. The presence of septa confirmed that germ tubes are precursors of septate mycelia. Ultrathin-section transmission electron microscopy of aldehyde plus osmium fixed cells revealed electron-lucent walls with a thin electron-dense outer layer. A fibrillar border was also routinely associated with germ tubes. Poststaining with potassium permanganate revealed, in addition, a previously invisible fuzzy layer on the outer region of the cell wall which extended over bud scars and germ tubes and which coalesced at sites of contact between cells.

This paper discusses computer simulation of the differential equations which describe javelin dynamics in flight. It is shown that the use of experimental aerodynamic forces and moments in the equations is preferable to theoretical approximations for these forces and moments which have been used in previous studies. An example which is characteristic of a good throw is presented and analyzed and many interesting features of the trajectory are pointed out.

We have developed a system to measure initial conditions in the javelin throw rapidly enough to be used by the thrower for feedback in performance improvement. The system consists of three subsystems whose main tasks are: (A) acquisition of automatically digitized high speed (200 Hz) video x, y position data for the first 0.1-0.2 s of the javelin flight after release (B) estimation of five javelin release conditions from the x, y position data and (C) graphical presentation to the thrower of these release conditions and a simulation of the subsequent flight together with optimal conditions and flight for the sam release velocity. The estimation scheme relies on a simulation model and is at least an order of magnitude more accurate than previously reported measurements of javelin release conditions. The system provides, for the first time ever in any throwing event, the ability to critique nearly instantly in a precise, quantitative manner the crucial factors in the throw which determine the range. This should be expected to much greater control and consistency of throwing variables by athletes who use system and could even lead to an evolution of new throwing techniques.

The reliability of the Enteric Analyzer for identification of Enterobacteriaceae was evaluated using biochemical results previously obtained for 291 organisms with the conventional, R/B, and Minitek systems. The instrument correctly identified 77.3% of the organisms using conventional system results, 74.2% using R/B results, and 60.5% using Minitek results. The low rate of identification with the conventional system occurs primarily because the instrument is not programmed to consider delayed biochemical reactions. The arbitrary use of 90% and 99% probabilities for test reactions also contributes to a lower percentage of identification. The Enteric Analyzer does not replace the judgment of experienced personnel in the identification of atypical bacteria, but it may prove helpful in speeding up final computer identification of typical microorganisms.

A personal series of twelve wrist arthroplasties performed on ten patients with rheumatoid arthritis of the radio-carpal joint were reviewed in retrospect. Six arthroplasties were of the Meuli metal/plastic/metal design and six were of the Swanson silastic type with an additional silastic ulnar head prosthesis. Both prostheses were successful in giving a pain free stable joint with some degree of useful movement. The Meuli appeared to give a greater range of movement than the more constrained Swanson prosthesis. Although complications were encountered it is concluded that wrist arthroplasty does have a place in the management of patients with rheumatoid arthritis of the radio-carpal joint. Our present policy is to continue with the use of the Swanson design because of the advantages of a cement free prosthesis.

Monoclonal antibodies have been raised against native calcineurin using conventional in vivo immunization and hybridoma procedures. The relatively high affinity of nonimmune IgG for the two subunits of calcineurin resulted in large nonspecific binding values for immunoassays of native, dissociated and denatured calcineurin, which complicated the antibody screening. Monoclonal aCn5, a high-affinity IgG1 that exhibits specific binding, was characterized. Other calmodulin-binding proteins tested were not recognized by aCn5. Simple binding properties were exhibited in solid-phase experiments, Kd = 26 (+/- 4) pM, but the stoichiometry was low. The loss of immunoreactivity after denaturation of calcineurin indicated that the aCn5 epitope is of the assembled topographic, not segmental, type. The epitope was located to the A subunit and affinity was unaffected by the presence of calcineurin B. The epitope remained intact after proteolytic removal of the amino-terminal 20 residues of calcineurin A essential for phosphatase activity, and the carboxyl-terminal inhibitory and calmodulin-binding domains. The calmodulin-binding peptide derived from calcineurin, cA8, was not recognized by aCn5. Addition of Ca2+, Mn2+, Ni2+, chelators or dithiothreitol did not influence the affinity of aCn5 for the holoenzyme. Phosphatase activity of calcineurin, in the presence and absence of calmodulin and after removal of the inhibitory domain, was little affected by aCn5. Thus, the aCn5 epitope defines a previously unidentified structural domain of calcineurin A located in a region of the proteolytically resistant core that is topologically distinct from the catalytic, inhibitory, calmodulin-binding and calcineurin-B-binding domains, and not functionally connected with calcineurin B or the putative metal-binding domain(s).

Conventional radioimmunoassays for IgG rheumatoid factors (RF) detect the binding of IgG RF (or their F(ab')2 fragments) to solid-phase human IgG Fc fragments or rabbit IgG. Binding is detected with a radiolabeled antibody that is IgG-specific but is nonreactive with human Fc (i.e., an alpha-Fd reagent) or with rabbit IgG. Anti-Fd reagents are quite laborious to produce. We developed a murine monoclonal alpha-Fd antibody, confirmed its specificity, and compared it with conventional rabbit alpha-Fd in the IgG RF radioimmunoassay. We also adapted the assay to an enzyme-linked assay with both human Fc and rabbit IgG as antigen. We compared the four assay methods with each other and examined their relationship to certain clinical and laboratory features of rheumatoid arthritis.

Most studies of mammalian vascular smooth muscle cell mechanics and pharmacology have relied upon data from intact tissue preparations. The purpose of this study was to determine how single mammalian vascular smooth muscle cell pharmacological sensitivity to histamine and force development compared to the intact tissue from which it was derived. Thus, techniques were developed for isolating large numbers (5 X 10(5) cells/ml) of cells from bovine carotid arteries. Single cells were spindle-shaped with dimensions of 117 +/- 11 (SE) micron in length by 7 +/- 1 (SE) micron in diameter (n = 27). The effect of histamine concentration on cell length was studied in suspensions of freshly isolated cells. Histamine concentration-response curves revealed that single smooth muscle cells shortened to histamine concentrations in a graded fashion, and that shortening was mediated through histamine H1 receptors. The threshold for 10% of maximum response occurred at 8.0 X 10(-11) M histamine. This threshold was at least three orders of magnitude lower than the threshold for 10% of maximum tension in the intact tissue under isometric conditions. In addition to cellular pharmacology, mechanical studies were performed by tying a single cell to an ultrasensitive force transducer. Active stresses of 0.5 +/- 0.1 X 10(5) N/m2 (n = 4) were measured in response to electrical stimulation for cells in low extracellular calcium (0.5 mM). These data suggest that histamine sensitivity and smooth muscle cell mechanics may best be determined in single cell studies that minimize diffusional barriers to agonist and avoid intercellular interactions that are present in the intact tissue.

The combination of a modified Staphylococci binding assay for immune complexes and Western blot analysis is described for the isolation and detection of antigen in immune complexes from human sera. The strategy of the procedure is to preclear immune complexes from other serum components by sequential polyethylene glycol precipitation and incubation with insoluble protein A under conditions in which immune complexes are preferentially bound. Immune complexes are eluted from protein A in sodium dodecyl sulfate buffer and dissociated by acrylamide electrophoresis. Resolved proteins are then transblotted to nitrocellulose, and antigen is detected with specific antibody. Immune complexes were prepared in vitro with an antigen (keyhole limpet hemocyanin) that focuses well on electrophoresis and for which a potent immunologic probe (antibody) was available. In this system, antigen could be detected when immune complexes were present in sera in concentrations as low as 20 micrograms aggregated-IgG eq/ml, regardless of antigen-antibody ratio. We demonstrate the detection of horse globulin in immune complexes from a patient with acute serum sickness and hepatitis B virus antigen in complexes from a patient with vasculitis.

Immunologic and serologic studies were done in 120 homosexual men who reportedly had no symptoms related to the acquired immunodeficiency syndrome. Forty-nine men (41%) had antibody to the retrovirus human T-lymphotropic virus type III/lymphadenopathy-associated virus (HTLV-III/LAV), and 37 (31%) had an abnormal T-cell subset ratio and other immunologic abnormalities. These abnormalities were almost exclusively confined to men seropositive for the retrovirus. Multivariate statistical analysis showed that exposure to HTLV-III/LAV was the single best predictor of T-cell (and other) immunologic abnormalities. Sexual practice, particularly receptive anal intercourse, predicted exposure to HTLV-III/LAV but was not independently related to T-cell abnormalities. The association of other microbial serologic findings with HTLV-III/LAV seropositivity or T-cell changes was a coincidental function of sexual activity. Immunologic abnormalities in asymptomatic homosexual men are mostly related to exposure to HTLV-III/LAV.

A rapid rate of isolation of molar-tooth germs from rodents aged from 19 embryonic days to 7 days postnatal can be achieved. The procedure is of direct application to biochemical studies of odontogenesis where relatively large amounts of tissue are often required. The quality of dissection, assessed by morphological and organ culture criteria, extends the application of this procedure to general use in the isolation of molar-tooth germs from rodents.

Whether the normal pericardium exerts a constraining effect on left ventricular (LV) diastolic compliance and/or systolic function is controversial. Left ventricular filling and performance were studied in 15 patients by two-dimensional transesophageal echocardiography (2D-TEE) measuring end-diastolic area (EDa), end-systolic area (ESa), ejection fraction area (EFa), and hemodynamics immediately pre- and post-pericardiotomy. To diminish the influences of other variables such as surgical stimulation, chest wall constraint, and autoregulation, measurements were performed in deeply anesthetized patients with the chest fully opened immediately before and after pericardiotomy (PC). No significant echocardiographic or hemodynamic changes were observed after PC. Although alterations in compliance cannot be excluded, no significant changes in LV diastolic filling (EDa, pulmonary capillary wedge pressure [PCWP]) or systolic performance (EFa, cardiac output [CO]) were found. Therefore, it is concluded that the normal pericardium does not exert a measurable constraining effect on LV performance.

Freezing lymphocytes in microtest plates instead of vials for the purpose of screening sera for HLA or other cytotoxic antibodies offers the same technical advantages as freezing sera in microtest plates for tissue typing. Plate-loading errors and cell wastage are minimized, and time required for serum screening is greatly reduced. A method for freezing lymphocytes in large numbers of microtest plates for use in serum screening that is simple, reproducible, and inexpensive is described. Cryoprotected lymphocytes dispensed into microtest plates are placed on a metal table that is lowered onto a tray containing 1 cm of liquid nitrogen in a Styrofoam box. The amount of liquid nitrogen in the tray and the height of the metal table above the tray determine the freezing rate of the lymphocytes. Over 100 microtest plates can be frozen per cycle with total cooling uniformity. The materials required for this freezing device can be assembled for less than $400. Commercial freezing units typically cost over $4,000.

The glycogen-binding (G) subunit of protein phosphatase-1G is phosphorylated stoichiometrically by glycogen synthase kinase-3 (GSK3), and with a greater catalytic efficiency than glycogen synthase, but only after prior phosphorylation by cyclic AMP-dependent protein kinase (A-kinase) at site 1. The residues phosphorylated are the first two serines in the sequence AIFKPGFSPQPSRRGS-, while the C-terminal serine (site 1) is one of the two residues phosphorylated by A-kinase. These findings demonstrate that (i) the G subunit undergoes multisite phosphorylation in vitro; (ii) phosphorylation by GSK3 requires the presence of a C-terminal phosphoserine residue; (iii) GSK3 can synergise with protein kinases other than casein kinase-2.

We isolated normal, nonactivated human monocytes from peripheral blood by four different methods: (1) rosetting with sheep erythrocytes pretreated with 2-aminoethylisothiouronium bromide hydrobromide (AET) followed by monoclonal antibody (OKT3 (CD3), B1 (CD19), Leu7, Leu11 (CD16] and complement treatment; (2) adherence to gelatin/plasma-coated flasks; (3) adherence to plastic dishes; and (4) separation by the Sepracell technique. We monitored these monocyte separations by determining cell recoveries, OKT4A+ lymphocyte contamination, monocyte binding to human immunodeficiency virus (HIV), number of non-specific esterase-positive cells, and proportion of mononuclear cells reactive with a battery of monoclonal antibodies specific for monocytes. Our results indicate that of the four methods compared, adherence to gelatin/plasma-coated flasks produced the highest purity, recovery, and satisfactory binding to HIV with the fewest contaminating CD4+ T cells.

Immune complex assays (and other immunologic tests) were performed on sera from 162 patients with the acquired immunodeficiency syndrome (AIDS) and 275 AIDS-related subjects. Immune complexes were detected in 89% of AIDS patients and 93% of homosexual men with lymphadenopathy. Immune complex levels in AIDS patients were not associated with a particular risk group or with types of opportunistic infection or malignancy; however, they correlated with other laboratory features of the immune defect (depression in T helper cells and T helper/suppressor-cell ratio, and IgG levels). Immune complexes were also detected in a lesser proportion of risk-group controls (homosexual men, hemophiliacs, Haitians). In risk-group controls, immune complex levels were associated with certain features reflecting sexual practice, blood product exposure, or infection, but these features did not account for the higher levels found in AIDS patients. In appropriate situations, immune complex assays may be of value as screening tests or, possibly, as prognostic indicators for AIDS or AIDS-related syndromes.

The interaction of calmodulin with calcineurin, a calcium- and calmodulin-stimulated protein phosphatase, was investigated using a solid-phase assay. Binding of 125I-calmodulin by calcineurin immobilized on nitrocellulose membrane filters was of high affinity, reversible, and calcium-dependent. Complex binding kinetics reflected a time- and calcium/calmodulin-dependent conformational change of calcineurin which was shown to be ligand-induced renaturation. After renaturation and removal of calmodulin, immobilized calcineurin exhibited simple 125I-calmodulin binding kinetics with a single class of independent sites. The maximum stoichiometry of 125I-calmodulin binding to immobilized calcineurin was 0.1 mol/mol. The association rate (K1 = 8.9 x 10(3) M-1 S-1) and the dissociation rate (K-1 = 8.5 x 10(-5) s-1) yielded a dissociation constant of Kd = 10 nM. Equilibrium binding analyses gave a Kd value of 16 nM. The affinity of 125I-calmodulin for immobilized calcineurin was half that of unmodified calmodulin. Using equilibrium competition experiments, we determined, for the first time, the dissociation constant for the binding of native calmodulin by calcineurin in solution, Kd less than or equal to 0.1 nM (Kd for 125I-calmodulin = 0.23 +/- 0.09 nM). The effects of ionic strength and pH on 125I-calmodulin binding to immobilized calcineurin were characterized. The dissociation rate was dependent on free calcium concentration, with half-maximal rate at 700 nM calcium. 125I-Calmodulin equilibrium binding by the immobilized A subunit of calcineurin exhibited half the affinity of the holoenzyme, Kd = 30 nM. The described phenomenon, of reversible denaturation associated with immobilization of a protein on nitrocellulose, may be a general one open to exploitation in other systems.