Pharmacokinetics and hemostatic effect of different factor VIII/von Willebrand factor concentrates in von Willebrand's disease type III.

Four different plasma-derived concentrates composed of coagulation factor VIII (FVIII) and von Willebrand factor (vWF) of varying quality (Hemate-P, Behring; Profilate, Alpha; and F VIII-VHP-vWF, C.R.T.S Lille), or almost purified vWF (Facteur Willebrand, C.R.T.S Lille) and one recombinant F VIII concentrate (Recombinate, Baxter) were given, in doses of 30-60 IU VIII: C/kg or 70-110 IU RCof/kg, to five patients with von Willebrand's disease type III, in order to evaluate the role of the vWF in factor F VIII concentrates. All plasma concentrates except Profilate had a multimeric vWF pattern almost similar to that of normal plasma. Bleeding time (b.t.), VIII: C, vWF:Ag, ristocetin cofactor activity, and multimeric pattern of the plasma-vWF were followed for 72 h. Both Duke b.t. and the multimeric pattern in plasma normalized after infusion of Hemate-P, F VIII-VHP-vWF, and Facteur Willebrand and, to a lesser extent, after Profilate. As expected, in response to Recombinate there was no effect on primary hemostasis, and the half-life of F VIII procoagulant activity (VIII: C) was very short. Normalization of the vWF is important not only for improving the primary hemostasis, but also for maintaining the plasma F VIII concentration on a high level, both by reducing the elimination rate of infused F VIII and via a secondary release of endogenous F VIII. If a prompt hemostatic effect is required, we recommend a concentrate containing both F VIII and all vWF multimers, but for prophylactic treatment, pure vWF may be used.


Effect of hypoxic exposure on iron absorption in heterozygous hypotransferrinaemic mice.

Heterozygous hypotransferrinaemic mice show serum iron, haemoglobin and reticulocyte levels similar to those of normal (+/+) controls but the plasma apotransferrin level is significantly reduced. Male heterozygous hypotransferrinaemic Balb/cJ mice and control (+/+) littermates were exposed to 1-3 days' hypoxia at 0.5 atmospheres. Controls increased their haemoglobin and iron absorption significantly during the first day of exposure; serum transferrin levels were increased and iron absorption had returned to normal levels by the third day or serum transferrin above normal levels and the elevated iron absorption persisted for longer than in controls. Serum iron levels were lower following hypoxic exposure than in controls. Liver iron loading was significantly enhanced by hypoxia in heterozygotes, but not in controls. These observations demonstrate the importance of the reservoir of plasma apotransferrin in the initial phase of enhanced erythropoiesis, and that the ability to enhance plasma levels of iron-bound transferrin is required in order to increase haemoglobin levels under the hypoxic stress.


Iron stores and serum transferrin receptor levels during recombinant human erythropoietin treatment of anemia in rheumatoid arthritis.

Ten rheumatoid arthritis (RA) patients with anemia of chronic disorders (ACD) were treated with recombinant human erythropoietin (r-Hu-Epo) using a dose of 250 U/kg s.c. 3 times a week for 6 weeks, in order to evaluate its effects on the anemia, iron stores, and serum-soluble transferrin receptor (sTfR) levels. All patients showed a rise in hemoglobin (Hb). Median Hb increased from 5.9 (5.5-7.0) at baseline to 6.7 (5.8-7.8) at 3 weeks and to 7.2 (5.9-8.5) mmol/l at 6 weeks during treatment. Ferritin levels decreased significantly during the 6 weeks, and five patients were iron deficient after 6 weeks of treatment. TfR levels increased significantly at 3 and 6 weeks during treatment. These preliminary findings may indicate that r-Hu-Epo is effective in improving ACD in RA. The sTfR rise may be explained by an increase in erythroid precursor cell mass or increased TfR expression and a decrease in tissue iron stores, although direct effects of Epo on TfR regulation cannot be excluded. Large double-blind studies with r-Hu-Epo in patients with RA and ACD are warranted.


Some storage characteristics of buffy coats used for preparation of platelet concentrates.

The present study examined biochemical storage lesions in 58 buffy coats (BCs) intended as the raw material for platelet concentrates. The work was conducted in three series; the aims were to investigate: (a) the storage quality of BCs obtained from the routine production (series I), (b) whether improvement in platelet quality could be achieved by continuous agitation during storage (series II), and (c) whether macroscopic aggregate formation indicates platelet damage (series III). Series I and II consisted of 20 BCs each. Series III compared ten BCs with visible clumping with eight BCs not demonstrating macroscopic aggregates. In series I and II platelet counts and platelet factor 4 (PF4) release were determined after 1, 3, 5, and 24 h; lactate dehydrogenase (LDH) extracellular activity was measured after 1 and 24 h. In addition, in series I elastase concentrations were analyzed after 3, 5, and 24 h. Finally, in series III extracellular beta-thromboglobulin (beta-TG) levels were determined after 18-24 h. Low platelet counts, most likely reflecting the presence of aggregate formation, were found in 20% of the BCs (series I); in this series a total of 25% demonstrated increasing platelet counts, together with elevated PF4 and elastase, over the study period. Significant biochemical storage lesions were found after 5 and 24 h of storage. The study also demonstrated that continuous agitation during storage does not improve platelet quality (series II) and that BCs demonstrating visible clumping (series III) had augmented levels of extracellular beta-TG (p < 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)


A case of congenital leukemia with monosomy 7.

A case of congenital leukemia with monosomy 7 is reported. Immunological study of the blast cells using monoclonal antibodies was suggestive of both myelomegakaryocytic and T-lymphoblastic leukemia. Chromosomal analysis of the bone marrow cells showed monosomy 7. Chemotherapy was initiated with a combination of adriamycin, cytosine arabinoside, 6-mercaptopurine, and prednisolone. The patient obtained complete remission, which has been maintained for 4 years and 1 month. He receives no chemotherapy now. Our case shows that monosomy 7 in congenital leukemia is rare, but the presence of monosomy 7 in congenital leukemia does not necessarily indicate a poor prognosis.


[Effect of superlow doses (10(-18)-10-(-14) M) of biologically active substances: general rules, features, and possible mechanisms].

The effects of ultra-low (10(-18)-10(-14) M) doses (ULD) of biologically active substances have been reviewed in terms of common regularities of ULD effects and peculiarities of action of various groups of compounds. The most common and at the same time paradoxical regularities of ULD action are bi- or polymodal patterns of dose dependence, absence or presence of an inverse effect at higher doses, and instability of ULD effect. Possible mechanisms of ULD action including the mechanism based on the adaptation theory are discussed.


[An unusual leucine motif in amino acid sequences of subunit 9 of ATP-synthase from plant mitochondria, modified by mRNA editing].

The amino acid sequences of subunit 9 of plant mitochondrial ATP-synthase contain a leucine motif which differs from the leucine repeat earlier detected within the composition of proteinaceous products of Myc, Fos, and Jun proto-oncogens. The structural organization of this repeat in proteolipid sequences can be modified via transformation of serine triplets into leucine ones as a result of editing of an appropriate mRNA.


[Structural changes of chromatin during proto-oncogene activation by cycloheximide: dose-effect].

The level of expression of cellular proto-oncogens c-myc and c-fos in rat liver has been studied as a function of protein synthesis rate (cycloheximide dose). Activation of proto-oncogens has been established to be initiated by 50% inhibition of nuclear protein synthesis. This promotes a certain level in chromatin structural rearrangements which is manifested, in particular, in decreasing activity of chromatin cleavage by Ca2+, Mg(2+)-DNAase and increasing degree of chromatin condensation. A role of topoisomerase II in chromatin structural rearrangements during proto-oncogen activation is postulated.


[Membrane ATPase of Vibrio alginolyticus. Ion transport activity and homology with F0F1-ATPase from E. coli].

F0F1-ATPase has been isolated from the marine alkali-resistant bacterium Vibrio alginolyticus. The enzyme subunits cross-reacted with antibodies against subunits alpha, beta, gamma, epsilon, and b of E. coli ATPase. The purified ATPase was reconstituted into liposomes effecting an ATP-dependent uptake of H+. Proton transport was inhibited by the ATPase blockers DCCD, triphenyltin, and venturicidin. Na+ ions had no effect on ATP-dependent proton transport. No ATP-dependent transport of Na+ was detected in proteoliposomes.


[Metabolism of extracellular phosphocreatine during changes in the ionic composition of the medium in the perfused rat heart].

Perfusion of isolated rat hearts with a phosphocreatine (10(-4) M) containing solution to which strophanthin or KCl had been added up to a concentration of 27 mM as well as Ca2+ depletion decreased phosphocreatine concentration in the perfusate with a simultaneous increase in creatine and phosphocreatine concentrations in the myocardium. Neither high extracellular concentrations of Na+ (200 mM), nor phosphocreatine increased creatine and phosphocreatine levels in the myocardium. The effect of high sodium perfusion media was completely reversed by strophanthin. Phosphocreatine decreased the lactate content in the perfusate. Strophanthin or potassium chloride enhanced the effect of phosphocreatine on the lactate release. Conversely, creatine augmented the lactate content in the perfusate. A high specificity of the phosphocreatine effect on the myocardium independently of the ionic composition of the perfusate was postulated. A mechanism of protective effects of phosphocreatine and high sodium perfusion media on "calcium paradox" is proposed.


[Significance of extracellular concentration of sodium ions in the protective effect during the "calcium paradox"].

Increasing of extracellular sodium concentration up to 200 mM diminishes heart damage under "calcium paradox". Phosphocreatine (10(-4) M) potentiates the effect of high sodium perfusion media; in this case myoglobin release from the myocardium is minimal (5-9% of control). An the same time, ATP and phosphocreatine concentrations and oxidation to phosphorylation coupling in mitochondria remain at a sufficiently high level. Elevation of osmotic pressure by the effect of 120 mM sucrose enhances heart damage under "calcium paradox" both in the presence and absence of phosphocreatine. The protective effects of superhigh (200 mM) sodium concentrations and phosphocreatine are completely reversed by strophanthin or decreasing K+ concentration down to 0.5 mM.


[Hydrocortisone and partial hepatectomy activate a proteinase associated with histones].

Proteinase activity has been shown to be associated with histones of rat thymus and liver nuclei. Hydrocortisone increases the activity of proteinases associated with thymus nuclear histones. Increasing activity of histone-associated proteinases is also observed during intensive transcription and replication in regenerating rat liver.


[Correlation between the stability of alpha-chymotrypsin at high temperatures and "salting in" of a strong solution].

A correlation was found between the thermal stability of alpha-chymotrypsin and the coefficient Ks of the Sechenov equation as a quantitative measure of the "salting-in" or "salting-out" capacity of solutes. At high temperatures, an increase in the concentration of "salting-in" agents (KSNC, GuHCl, urea, formamide) resulted in thermal stabilization of alpha-chymotrypsin. The maximal (about 100-fold) stabilizing effect in concentrated solutions of salting-in agents was comparable with those induced by covalent modification with hydrophilic reagents or immobilization. Conversely, an increase in the concentration of "salting-out" agents stabilized the enzyme only marginally at high temperatures. An additivity of solutes' action on the thermal stability of the protein has been demonstrated. The observed correlation was explained in terms of the solutes' action on the reversible conformational transition of the enzyme native form into a much more stable form existing at high temperatures.


[The effect of pyridine and pyridine-N-oxide on the monooxygenase system of rat liver microsomes].

Effects of pyridine and pyridine-N-oxide on the monooxygenase system of rat liver microsomes have been studied. Pyridine (200 mg/kg) increased total cytochrome P-450 content and activated metabolism of some specific substrates 24 hours after injection. There was an increase in the degree of p-nitrophenol and chlorzoxazone hydroxylation due to increasing ethanol-induced cytochrome P-450IIE1 content. Pyridine was also able to induce cytochrome P-450IIB1 in rat microsomes; this reaction was accompanied by acceleration of 7-pentoxyresorufin 0-dealkylation. Cytochrome P-450IA1 appearance in liver microsomes was associated with increasing content of cytochrome P-450IA2. Dealkylation rates for specific substrates (7-ethoxyresorufin and 7-methoxyresorufin) were also increased. Similar to pyridine, pyridine-7-oxide induced cytochromes P-450IIE1, P-450IIB1/B2, and P-450IA1/A2, resulting in activation of specific substrate metabolism. Hence, pyridine and its derivative pyridine-N-oxide can be regarded as effective inducers of cytochrome P-450.


Cell death induced by vincristine in the intestinal crypts of mice and in a human Burkitt's lymphoma cell line.

Although vincristine is widely used clinically in the treatment of some human cancers, its mechanism of action has not been clearly established. In this study, the patterns of cell death induced by vincristine in the intestinal crypts of mice and in a human Burkitt's lymphoma cell line were investigated by light and electron microscopy. Vincristine was found to enhance apoptosis of interphase cells in both systems and also to cause the arrest of cells in mitosis, the latter effect being more pronounced in the intestinal crypts. Arrested mitotic cells went on to die by a process that had a number of features in common with apoptosis. These include compaction of chromatin (following coalescence of chromosomes), condensation of the cytoplasm, initial preservation of organelle integrity, and eventually the fragmentation of the cell into a number of membrane-enclosed bodies which are morphologically similar to conventional apoptotic bodies. The results suggest that the cytocidal effect of vincristine is not solely dependent on metaphase arrest but is a cumulative one, resulting both from apoptosis of interphase cells and the 'apoptotic-like' death of cells arrested in metaphase.


The role of cell death in the growth of preneoplastic lesions: a Monte Carlo simulation model.

A variety of experimental and clinical examples of preneoplasia demonstrate that regression of early lesions is common. This paper examines the hypothesis that early lesions operate under the identical growth kinetics of 'late' lesions (neoplasms), but that kinetic features favouring continuous growth in established lesions tend to favour extinction of lesions composed of small numbers of cells. Growth simulations of early lesions were produced using the Monte Carlo method, a technique demanding intensive computations. With the advent of powerful personal computers, this technique is now widely available to biologists. Simulating growth under conditions of cell loss similar to those observed in established tumours, the model predicts that the great majority of initiated cell clusters are expected to reach extinction within a few cell doubling times, and most early (promoted) lesions would not likely progress to the size of a clinically detectable lesion within the life span of the host organism. These Monte Carlo simulations provide a model of initiated cell growth consistent with the recently demonstrated role of early lesion cell death in the development of human lymphomas and in transgenic mice expressing the bcl-2 oncogene. The model demonstrates that small increments in the intrinsic cell loss probability in even the earliest progenitors of malignancy can strongly influence the subsequent development of neoplasia from initiated foci.


A model of the control of cellular regeneration in the intestinal crypt after perturbation based solely on local stem cell regulation.

The control mechanisms involved in regeneration of murine intestinal crypts after perturbations are presently not well understood. The existence of some feedback signals from the cells on the villus to the cells in the crypt has been suggested. However, some recent experimental data point to the fact that regeneration in the crypt starts very early after perturbation, at a time when the villus cell population has hardly changed. In particular, this early cell proliferative activity is seen specifically at the bottom of the crypt, i.e. in the presumed stem cell zone and furthest from the villus. The objective of this study was to investigate whether a new concept of regulation operating solely at the stem cell level could explain the present mass of accumulated data on the post-irradiation recovery, which is an extensively studied perturbation from the experimental point of view. In order to check its validity, the new concept was formalized as a mathematical simulation model thus enabling comparison with experimental data. The model describes the cellular development from stem cells to the mature villus cells. As a basic feature it is assumed that the self-maintenance and the cell cycle activity of the stem cells are controlled by the number of these cells in an autoregulatory fashion. The essential features of the experimental data (i.e. the recovery with time and the consistency between different types of measurements) can be very well reproduced by simulations using a range of model parameters. Thus, we conclude that stem cell autoregulation is a valid concept which could replace the villus crypt feedback concept in explaining the early changes after irradiation when the damage primarily affects the crypt. The question of the detailed nature of the control process requires further investigation.


Ferritin-iron increases killing of Chinese hamster ovary cells by X-irradiation.

Stationary-phase Chinese hamster ovary cells were cultured in medium containing ferritin (approximately 19% iron by weight) added at concentrations ranging from 0 to 128 micrograms/ml. One set of cultures was unirradiated, and another set was exposed to 4.0 Gy of X-ray. Clonogenic cell survival was assessed in each set of cultures. In the absence of added ferritin, 4.0 Gy killed approximately 50% of the cells. In the absence of radiation, ferritin was not toxic at less than 48 micrograms/ml; above 48 micrograms/ml, toxicity increased with concentration. Apoferritin was not toxic at any concentration tested (up to 1000 micrograms/ml). Although 32 micrograms/ml ferritin, reflecting only a 3-6 fold increase in iron concentration over normal serum, was not toxic, it reduced the survival of X-irradiated cells by an additional 75%. These results indicate that a sublethal concentration of ferritin can be a potent radiosensitizer. This suggests the possibility that high body iron stores may increase susceptibility to radiation injury in humans.


Haemopoietic long-term bone marrow cultures from adult mice show osteogenic capacity in vitro on 3-dimensional collagen sponges.

Adult murine bone marrow cells, cultured under conditions for long-term haemopoietic marrow cultures, produce bone matrix proteins and mineralized tissue in vitro, but only after the adherent stromal cells were loaded on a 3-dimensional collagen sponge. Provided more than 8 x 10(6) cells are loaded, mineralization as measured by 85Sr uptake from the culture medium, occurred in this 3-dimensional configuration (3-D) within 6 days. In contrast if undisrupted marrow fragments (containing more than 10(7) cells) are placed directly on a collagen sponge, then it requires more than 10 days before significant mineralization can similarly be detected. The 2-dimensional (2-D) long-term marrow culture system allows prior expansion of the stromal cells and some differentiation in an osteogenic direction within the adherent stromal layer. This is suggested by the presence of type I collagen and alkaline phosphatase positive cells. However; synthesis of osteonectin and a bone specific protein, osteocalcin, as well as calcification are only observed in 3-D cultures. Electron microscopy demonstrated hydroxyapatite mineral on collagen fibres, osteoblast-like cells, fibroblasts, cells which accumulated lipids, and macrophages which were retained on the collagen matrices. Irradiation of confluent long-term bone marrow cultures, prior to their loading on the collagen sponge showed that haemopoietic stem cells are not necessary for the mineralization.


Autoradiographic studies of rat astroglial cell proliferation in vitro with and without treatment with basic fibroblast growth factor.

Using specific autoradiographic methods, cell cycle parameters of untreated and basic fibroblast growth factor (bFGF)-treated astroglial cells from newborn rats grown in primary culture were directly measured. The mode of proliferation was also analysed. In untreated cultures, S phase duration (Ts = 6.9-13.1 h) and cell cycle time (Tc = 10-18 h) can be modified by about a factor of 2 depending on the culture conditions (serum-supplemented or defined medium, thyroid hormone concentration). However, growth fraction (GF = 0.15) and the ratio Ts/Tc remain stable. With increasing days in vitro (DIV) (DIV 7-DIV 20), Ts (7.8-10.6 h) and Tc (10-21 h) are prolonged and GF (0.14-0.06) decreases, probably due to cell maturation. In general, astroglial cells proliferate exponentially with a GF < 1, but stop proliferating about 30-36 h after the last feeding, probably caused by exhaustion of the medium. However, after refeeding they continue to proliferate. As opposed to in vivo, no transition of non-proliferating cells into the GF occurs. After addition of bFGF, GF increases (e.g. GF at DIV 7 = 0.43), but Ts and Tc are not influenced at DIV 7 and 12. At DIV 20, bFGF additionally shortens Ts and Tc, thereby producing values of Ts, Tc and GF like 'younger' cultures. However, the revitalizing effect on 'mature' cells is only transitory. In general, bFGF leads to a single re-entry of G0 cells into the GF. Thereafter, bFGF does not affect the mode of proliferation.


The mechanism of action of the tetrapeptide acetyl-N-Ser-Asp-Lys-Pro (AcSDKP) in the control of haematopoietic stem cell proliferation.

The mechanism of action of the haemoregulatory tetrapeptide Acetyl-N-Ser-Asp-Lys-Pro (AcSDKP, M(r) = 487 amu), was investigated using an in vitro assay of a murine high proliferative potential colony-forming cell (HPP-CFC) which responds to proliferation regulators of the haematopoietic stem cell population. AcSDKP had no direct inhibitory effect on the number, or the proportion in S phase, of the committed granulocyte-macrophage progenitor (GM-CFC), or cycling HPP-CFC populations. However, AcSDKP blocked the action of a stimulator of haematopoietic stem cell proliferation, preventing the switching of quiescent HPP-CFC into cell cycle. It would appear that AcSDKP exerts its inhibitory haemoregulatory role indirectly, by preventing stimulator action on haematopoietic stem cells.


Effects of growth media on cell cycle progression in CHO cells exposed to the radioprotector WR-1065.

WR-1065 (2-[(aminopropyl)amino]ethanethiol) reduces cytotoxic and mutagenic effects caused by exposure of cells to radiation and chemotherapeutic drugs, but the mechanisms involved are not fully known. We have observed an accumulation of cells in G2 in WR-1065 treated Chinese hamster ovary cells grown in alpha-minimal essential medium, while others have found no cell cycle effects in WR-1065 treated Chinese hamster ovary cells grown in McCoy's 5A medium. To determine if the two types of media had an effect on cells treated with WR-1065, we examined survival and cell cycle progression. Population doubling times of 12 h were observed for cells grown in both media. Incubation of AA8 cells grown in McCoy's 5A medium with 4 mM WR-1065 30 min prior to and during irradiation with 137Cs gamma-rays resulted in a protection factor of 2.2, in close agreement with the value of 2.0 we previously obtained for AA8 cells grown in alpha-minimal essential medium. Treatment with WR-1065 caused an alteration in the cell cycles of cells grown in both media. An increase in the G2 population and a decrease in the G1 population was observed in cells incubated up to 3 h in the presence of 4 mM WR-1065, with a redistribution of the cells throughout the cell cycle occurring following removal of the drug. These data suggest that exposure of cells to WR-1065 is the cause of perturbations in cell cycle progression, and is not affected by the type of medium the cells are grown in.


Comparison and characterization of retinal pericytes and retinal pigment epithelial cells on subcellular IP3-sensitive Ca2+ pools.

A comparative study of inositol 1,4,5-trisphosphate (IP3)-induced Ca2+ mobilization in bovine retinal capillary pericytes (BRCP) and bovine retinal pigment epithelial cells (BRPE) was carried out. Both cells were permeabilized with saponin. The two cell types had similar basal levels of [Ca2+]i (130 nM for BRCP, 132 nM for BRPE) and responded to IP3 in a dose-dependent manner. However, when stimulated by various concentrations of IP3 (1-10 microM), the increase in [Ca2+]i of BRCP was always two- to threefold higher than that in BRPE. Subcellular-fractionation studies showed that a single population of IP3 binding site with a high affinity and high specificity of IP3 mainly localized to plasma membrane in these two cell types. Although the dissociation constant of specific [32P]-IP3 binding sites (Kd 1.9-2.8 nM) was similar, the profile of maximal binding capacity (Bmax) of each fraction was markedly different. In comparison, plasma membrane fractions of BRCP were with Bmax of 165 fmol/mg protein versus 90 fmol/mg protein for BRPE membranes. The ATP-dependent Ca2+ uptake and IP3-dependent Ca2+ release were observed in the both plasma membrane fractions. With quantitative correlation, the membrane fraction (2 mg) of BRCP released 0.2 nmol Ca2+ whereas BRPE only released 0.07 nmol Ca2+ with the same dose of IP3 (5 microM). The selectively higher density of IP3 binding sites in coupling to the larger Ca(2+)-release in the membrane of BRCP suggests that the quantity of Ca2+ mobilized is determined by the spatially preferential distribution of membrane-associated IP3 binding sites. These findings may provide an explanation for the differences observed between BRCP and BRPE in IP3-induced DNA replication.


The relationship of chemical modification of membrane proteins and plasma lipoproteins to reduced membrane fluidity of erythrocytes from diabetic subjects.

The significance of the two most common hallmarks of the diabetic state, hyperglycaemia and hyperlipidaemia, was investigated in terms of disorders of cell membrane dynamics. In order to examine whether the alterations in cell membrane lipid bilayer dynamics are somehow related to protein chemical modifications in plasma low-(LDL) and high-density lipoproteins (HDL) and blood cell membranes, we compared 19 poorly controlled diabetic subjects with 19 age- and sex-matched controls. The extent of (non-enzymatic) glycation, lipid peroxidation and the cholesterol/phospholipid ratio were increased in plasma low density lipoproteins and high density lipoproteins from diabetic patients. The mean steady-state fluorescence polarization values in 1,6-diphenyl-1,3,5-hexatriene-labelled isolated erythrocyte membranes from diabetic subjects were significantly greater than from control subjects (0.186 +/- 0.008 vs 0.173 +/- 0.006, p < 0.001); the fluorescence polarization values in erythrocyte membranes from diabetic and control subjects positively correlated with the extent of membrane protein glycation, lipid peroxidation and the cholesterol content. The cholesterol to phospholipid molar ratios in low density lipoproteins and high-density lipoproteins from diabetic and control subjects correlated significantly with the fluorescence polarization values in erythrocyte membranes from these subjects. Furthermore, the extent of glycation of low density lipoproteins appears to be strongly correlated with the extent of lipoprotein lipid peroxidation (r = 0.789, p < 0.001). The atherosclerotic potential of plasma lipoproteins in diabetes mellitus was discussed in terms of membrane and plasma protein chemical modifications.


Determination of 2'-5'-oligoadenylate synthetase in serum and peripheral blood mononuclear cells before and after subcutaneous application of recombinant interferon beta and gamma.

The interferon-inducible enzyme, 2'-5'-oligoadenylate synthetase, was estimated in healthy donors and in patients before and after subcutaneous application of recombinant interferon beta and gamma. Tests were carried out with lysates of peripheral blood mononuclear cells, using an established radioenzymatic assay, and in serum samples, using a new radioimmunoassay. Both test systems substantially yielded the same results: after a single injection of interferon beta (1-5 x 10(6) IU), 2'-5'-oligoadenylate synthetase increased in a dose-dependent manner reaching maximal catalytic concentrations in most patients after 24-48 hours (leukocytes) and 48-72 hours (serum). In contrast, interferon gamma (2-4 x 10(6) IU) caused only a small induction of 2'-5'-oligoadenylate synthetase. However, daily application of interferon gamma for 7 days led to a distinct time-dependent increase of 2'-5'-oligoadenylate synthetase activity concentration during this observation period. Characteristically, even during daily application, the 2'-5'-oligoadenylate synthetase activity concentration dropped just 48-72 hours after the first injection of interferon beta. The determination of 2'-5'-oligoadenylate synthetase proved to be useful for optimizing and monitoring subcutaneous therapy with interferon. The new radioimmunoassay which allows the determination of this enzyme in serum is superior to other methods used in the past.


Affinity of europium-labelled proteins A and G for immunoglobulin G from seven mammalian species.

Proteins A and G were each labelled with two different europium chelates (p-isothiocyanatophenyl-ethylenediaminetetraacetic acid and diethylenetriaminepentaacetic acid). Their affinities for IgG from rabbit, goat, horse, sheep, mouse, pig, and rat were then measured using time-resolved fluorescence. Protein G labelled with the second chelate was found to be especially effective in binding to goat and horse IgG.


A fully automated enzyme immunoassay for the measurement of cortisol in biological fluids.

This paper describes a fully automated assay on the Serono SR1 for the measurement of cortisol in serum, heparinised plasma and urine. The assay incorporates a specific polyclonal antibody to cortisol and cortisol conjugated to alkaline phosphatase as a label. Following immunoincubation, bound and free labelled cortisol are separated by magnetic sedimentation of the antibody complex. Phenolphthalein is liberated by the enzymatic hydrolysis of the substrate (phenolphthalein monophosphate) and the absorbance generated is measured as the assay end-point. All processing steps are performed by the SR1. The assay has good analytical performance with respect to precision (within and between run CVs less than 10 and 11.5% respectively), detection limit (less than 5 nmol/l) and recovery of added cortisol (100 +/- 10%). The assay agrees well with cortisol concentrations determined by ID-MS and by established immunoassays (r > 0.96). Reference ranges of normal urine samples (pretreated by solvent extraction) are in good agreement with accepted values. The study demonstrates that the SR1 Cortisol assay on the SR1 analyzer is suitable for the routine determination of cortisol in serum, heparinised plasma and urine samples.


Which is the appropriate coenzyme for the measurement of ammonia with glutamate dehydrogenase?

The determination of ammonia in plasma, using glutamate dehydrogenase, is complicated by non-specific oxidation of the coenzyme, promoted by components of the sample matrix. Measurements performed with appropriate plasma blanks show that 2'-phosphorylated coenzymes (NADPH, deamino-NADPH) are much less oxidized than NADH. By adding lactate dehydrogenase, NADH oxidation by endogenous pyruvate can be completed within a short time. Considerable consumption of coenzyme occurs, however, and endogenous L-alanine aminotransferase also represents a possible source of interference. The results of ammonia determinations using deamino-NADPH (y) or NADPH (x) were identical (a = 0.0 mumol/l, b = 1.00; r = 0.996, n = 62). With NADH as the coenzyme, the method displays adequate specificity only at high sample dilution, e.g. in the measurement of urea after conversion to ammonia.


Free epinephrine, norepinephrine and dopamine in saliva and plasma of healthy adults.

To investigate whether salivary catecholamine levels reflect short term changes of sympathoadrenal activity, we simultaneously measured plasma and saliva epinephrine, norepinephrine and dopamine concentrations at rest, during bicycle ergometry and during epinephrine infusion in 12 healthy adults, using a radioenzymatic method. Whereas all plasma catecholamines significantly increased during bicycle ergometry and epinephrine infusion, no changes were observed in the salivary catecholamine concentrations and the salivary catecholamine flow rate after stimulation by rhythmic chewing. Moreover, salivary catecholamine concentrations were related neither to heart rate nor to blood pressure. Obviously, free catecholamines are rapidly inactivated by enzymatic degradation, neuronal reuptake or extraneuronal uptake in tissues before they diffuse through the tight junctions into the saliva. Therefore, stimulated saliva catecholamine levels do not reflect short term changes in the activity of the sympathoadrenal nervous system.


Influence of reagent composition on atypical pseudocholinesterase activity measurement: comparison of a manual and an automated method and implications for routine.

In determining activities of atypical pseudocholinesterase using a Hitachi 737 and a Cobas Bio analyser, respectively, we repeatedly found discrepancies between the enzyme activities measured by both instruments which were presumably due to differences in the reagent composition (Hitachi 737: Boehringer Mannheim Automated Analysis for BM/Hitachi Systems; Cobas Bio: Boehringer Mannheim Monotest). We consider the present results as a strong hint for manufacturers towards generally declaring all components of the chemical systems they produce and as a warning for users when interchanging apparently identically composed sets of reagents.


A multicentre evaluation of the Ektachem DT60-, Reflotron- and Seralyzer III systems.

The analytical performance of the three analytical systems Reflotron, Ektachem DT60 and Seralyzer III was studied according to ECCLS guidelines (1) and partly according to a protocol of the Société Francaise de Biologie Clinique (SFBC) (2) in a multicentre evaluation involving four laboratories. The determination of 11 analytes led to more than 180,000 data. With the Ektachem DT60, imprecision was acceptable for all analytes except for sodium. With the Reflotron, imprecision for glucose and creatinine was wider than the acceptable limits. With the Seralyzer III, imprecision for glucose, uric acid, cholesterol, creatinine and potassium was not within acceptance limits. The recovery of system assigned control sera values was acceptable for all analytes, except for glucose, creatinine and sodium with the Ektachem DT60 system, and for all but glucose and cholesterol with the Reflotron. On the Seralyzer III, the limits of acceptance were exceeded only with the creatinine assay. The recovery of the reference method values with Kontrollogen L caused problems with all three systems and for all analytes. Only 30% of the mean values of Kontrollogen L measured with the Ektachem and with the Seralyzer III were within the limits of acceptance. 40% of the mean values determined with the Reflotron were inside these limits. The upper limits of linearity as claimed by the manufacturers were obtained with all analytes and systems with the exception of cholesterol on all systems and the creatinine assay on the Seralyzer III. The systems under test and several different comparison methods showed good agreement for the analysis of patient samples, except in one laboratory for the analysis of sodium and aspartate aminotransferase with the Ektachem system and potassium with the Seralyzer III. Turbidity showed no significant influence on the measurements of all analytes and all systems. Haemolysis, hyperproteinaemia, and bilirubinaemia affected several methods on all three systems. A start time delay of up to 60 s did not affect the results of the Reflotron, except in the case of the triacylglycerol assay, which was affected by start time delays greater than 45 s. The results of 4 assays on the Seralyzer III were decreased considerably by a delayed start time (triacylglycerols and creatinine above 5 s, aspartate aminotransferase above 35 s and creatine kinase above 15 s). For reliable results from all the assays in each of the three analytical systems, it was necessary to use the prescribed sample volume within certain limits. The practicability of all analytical systems tested was found to be very good. A field study was conducted with the Reflotron system. The analyte concentration was determined in venous blood from various patients. In 25 out of 30 experiments, the results of the "field" laboratories showed a greater spread about the fitting line than those obtained in the "expert" laboratories.


Event-related potential correlates of impaired selective attention in children at high risk for schizophrenia.

Auditory event-related potentials (ERPs) associated with selective attention were recorded in 21 children at high-risk for schizophrenia and in 21 matched controls. The subjects performed a selective listening task. For behavioral evaluation, target counts on the selective listening task and on cognitive performance were assessed. Group-specific differences of ERP components could be demonstrated, as reflected by significant amplitude reductions of the frontally located negative difference wave (Nd) and of the P3 component, following selectively attended stimuli, in the high-risk children. P3 latencies tended to be prolonged in the high-risk group. Reduced Nd was found in 14 and reduced P3 in 16 high-risk children among the 21 matched pairs. Significant correlations between the ERP reductions and psychometric deficit (counting accuracy) were observed. Mismatch negativity (MMN), an ERP component associated with automatic processing of physically deviant stimuli, did not differentiate significantly between groups, but was distinctly reduced in the high-risk group. The Nd and P3 reductions suggest deficits of selective attention in a considerable number of the subjects genetically at risk for schizophrenia. The present findings are discussed with respect to their relevance as indicators of a predisposition to schizophrenia.


Periodicity of episode occurrences in rapid cycling affective disorders.

We analyzed the medical records of nine patients with severe rapid cycling affective disorders (RCAD), and determined the cycle of mania occurrences by calculating the period between two successive onsets of mania for each patient. Using the cycle as the index we devised a cycle-oriented diagram by dividing the observation period by the index cycle, and visually studied the mode of episode occurrences. In seven patients, the onset of mania generally followed the index cycle, but sometimes shifted from the day estimated by the index cycle. The shift seemed to be caused by a cycle other than the index cycle, which appeared temporarily in the course. The period between two successive onsets of mania was often several times longer than the index cycle length. In patients with RCAD, some sort of periodic rhythms may possibly control the course of episode occurrences on a continuing basis.


EEG mapping and psychopharmacological studies with denbufylline in SDAT and MID.

Computed tomography (CT), electroencephalograms (EEG), clinical and psychometric data were obtained in 96 mildly to moderately demented patients (72 women, 24 men), aged 61-96 years (mean 82), diagnosed according to DSM-III criteria. Patients were off drugs for at least 2 weeks and subdiagnosed according to the modified Marshall-Hachinski ischemic score and CT in 45 senile dementia of the Alzheimer type (SDAT) and 51 multiinfarct dementia (MID) patients. Evaluations were carried out before and 12 weeks after treatment with either 100 mg denbufylline BID or placebo and included EEG mapping, the Sandoz Clinical Assessment Geriatric (SCAG) score/factors, the Clinical Global Impression (CGI), the Digit Symbol Substitution Test (DSST), the Trail-Making Test (TMT) and the Digit Span Test (DS). Descriptive data analysis including confirmatory statements found delta/theta activity enhanced, alpha and beta activity reduced, total power augmented, and the centroid slowed down over various brain regions in patients as compared with controls. The two subtypes of dementia could be differentiated in some conventional EEG variables but mostly by means of power asymmetry indices. Denbufylline induced a statistically significant and clinically relevant improvement in both SDAT and MID patients, whereas after placebo this was not the case in CGI, the TMT, and the DS, with interdrug differences being significant in all primary target variables such as the CGI, MMS, SCAG, and DSST. Thus, both the degenerative and vascular type of dementia exhibited a therapeutic benefit that could be objectified at the neurophysiological level by EEG mapping in an improvement of vigilance.


Brainstem auditory evoked responses (BAERs) during the menstrual cycle in women with and without premenstrual syndrome.

Brainstem auditory evoked responses (BAERs) were recorded at three menstrual cycle phases (menstrual, late follicular, and late luteal) in a sample of healthy control women (n = 21) and in a sample of women (n = 30) diagnosed as suffering from Late Luteal Phase Dysphoric Disorder (LLPDD). The latter were divided on the basis of retrospective and prospective self-ratings into moderate (PMS+) and severe (PMS++) symptom groups. Results showed (1) no change in BAER latencies at different phases of the menstrual cycle; (2) increased BAER latencies for wave III in women with moderate PMS symptoms compared with healthy controls; (3) increased BAER latencies for waves III and V in women with severe PMS symptoms compared with healthy controls. These results raise the possibility of brainstem dysfunction in PMS women, and support the idea of a neurobiological predisposition to this order.


Blood flow response to auditory stimulations in normal, mentally retarded, and autistic children: a preliminary transcranial Doppler ultrasonographic study of the middle cerebral arteries.

Using the noninvasive transcranial ultrasonic Doppler method, flow dynamics of the middle cerebral arteries were investigated in relation to auditory stimulations in 12 children with autistic behavior compared with 12 normal controls and 10 mentally retarded children. In normal children, auditory stimulation evoked lateralized modifications: blood flow increased and resistance index decreased on the left side; such modifications were not recorded on the right side. This pattern should indicate vasodilatation mechanisms induced by changes in the metabolism of the brain areas supplied by the left middle cerebral arteries (MCA). Although less asymmetrical, this pattern was also found in the mentally retarded children. Autistic children significantly differed from these two groups. They displayed a symmetric pattern of responses with a blood flow decrease and resistance-index increase on both sides; this could suggest abnormal metabolic mechanisms induced by auditory stimulation in autistic children and could be related to the previous hypothesis of impairment in the development of cerebral lateralization in autism. These preliminary results show that transcranial Doppler ultrasonography may be a valuable and practicable tool for the noninvasive study of evoked blood flow responses in psychopathology.


Changes in norepinephrine output following light therapy for fall/winter seasonal depression.

Recurrent fall/winter depressions that remit during spring and summer have been called Seasonal Affective Disorders (SAD) (Wehr and Rosenthal 1989). The pathophysiology of SAD, its relationship to nonseasonal affective disorders, and the mechanism of action of light therapy, which is effective in treating SAD, remain to be elucidated (Depue et al 1989; Jacobsen et al 1987; James et al 1986; Joseph-Vanderpool et al 1991; Skwerer et al 1988, Terman et al 1989). Norepinephrine (NE) may play a role in the mechanisms of action of many antidepressant treatments (Schildkraut 1965) that alter NE metabolism (Schildkraut et al 1964 and 1965) and decrease the urinary output of NE and its metabolites, i.e., "whole-body NE turnover" (WBNET) (Golden et al 1988; Potter et al 1988). The present study explored whether light therapy also reduces the urinary output of NE and its metabolites.


Melatonin and jet lag: confirmatory result using a simplified protocol.

This study replicates the alleviation of jet-lag with melatonin in a simplified protocol for eastward flight. At 22-n hr (n is the time-lag between the North American departure point and France), subjects took either melatonin (8 mg, n = 15), or placebo (n = 15) on the day of the return flight and for 3 consecutive days. On day 8, self-ratings significantly discriminated between melatonin and placebo for global treatment efficacy, morning fatigue, and evening sleepiness.


Evolving role of flucytosine in immunocompromised patients: new insights into safety, pharmacokinetics, and antifungal therapy.

Flucytosine is an antifungal agent useful in combination with amphotericin B in the treatment of several deeply invasive mycoses. The potentially dose-limiting, hematologic, gastrointestinal, and hepatic toxicities of flucytosine lead to a reluctance to use it in myelosuppressed patients. To investigate the safety and tolerability of flucytosine in this setting, we evaluated its use in 17 patients with cancer or aplastic anemia during a 2 1/2-year period at our institution and reviewed the literature describing mechanisms of action, resistance, in vitro and in vivo antifungal activity, clinical antifungal activity, pharmacokinetics, and toxicity. The combination of amphotericin B plus flucytosine eradicated the mycosis in 12 (71%) of 17 patients, whereas 3 (18%) of 17 died of progressive fungal infection. Serial serum levels of flucytosine measured by a creatinine iminohydrolase assay permitted reliable dosage adjustment. During therapy, only 2 (12%) of 17 patients had elevated mean serum levels of flucytosine (> 100 micrograms/mL) and 3 (18%) other patients had transiently elevated levels. Paired serum samples (n = 45) obtained at steady state during therapy with orally administered flucytosine showed similar peak and trough levels. Adverse effects of flucytosine therapy included one case each of reversible nausea, diarrhea, elevated transaminase levels, and thrombocytopenia. No cases of bone marrow aplasia, enterocolitis, hepatitis, or death due to flucytosine toxicity were encountered. We conclude that flucytosine in combination with amphotericin B is well tolerated in myelosuppressed patients when serum flucytosine levels are serially monitored.


Human babesiosis in New York State: an epidemiological description of 136 cases.

Epidemiological data on 136 cases of human babesiosis reported from laboratories and clinicians in the state of New York from 1982 to 1991 were reviewed. All but two patients, who had traveled to Nantucket Island in Massachusetts, acquired disease in Suffolk County, Long Island. The highest average age-group-specific annual incidence rates occurred among men > or = 80 years of age (7.72 per 100,000) and among women 70-79 years of age (3.61 per 100,000). Seven patients (5%) had previously undergone splenectomy, and 31 (23%) had evidence of concurrent Lyme disease. One hundred three patients (76%) were hospitalized, and seven (5%) died. Although the geographic distribution of this disease has remained constant, the recent introduction of babesiosis in mainland Connecticut and the spread of Lyme disease in New York suggest that the geographic distribution of babesiosis could also spread in New York.


Culture isolation of Acanthamoeba species and leptomyxid amebas from patients with amebic meningoencephalitis, including two patients with AIDS.

Acanthamoeba species and leptomyxid organisms are free-living amebas that cause meningoencephalitis, primarily in immunocompromised patients. We report the isolation and culture of Acanthamoeba species and leptomyxid amebas from four patients with fatal amebic meningoencephalitis. Acanthamoeba species were cultured from brain abscess specimens from three immunocompromised patients (including two patients with AIDS). In the case of the fourth patient, who had no identifiable immunodeficiency, leptomyxid amebas were cultured from a specimen from a subcutaneous nodule and were identified in amebic granulomas in brain tissue by the indirect immunofluorescence test. Persons with advanced infection due to the human immunodeficiency virus may be at increased risk for amebic meningoencephalitis, but the diagnosis should be considered in the differential diagnosis of any immunocompromised patient with cerebral abscesses.


The management of pregnancies complicated by genital infections with herpes simplex virus.

In this review we summarize current knowledge related to the identification of pregnancies that may be complicated by genital herpes and describe the consequences of maternal infections with genital herpes. We address the implications of this information for the management of genital herpes during pregnancy and at delivery and for the care of neonates exposed to herpes simplex virus at delivery. On the basis of the current data, we cannot make specific recommendations concerning many of the clinical problems that are caused by herpes simplex virus infections in pregnant women. We identify and discuss unresolved questions about optimal management.


Successful treatment of disseminated nocardiosis complicated by cerebral abscess with ceftriaxone and amikacin: case report.

We report the case of an 85-year-old female patient who suffered from disseminated Nocardia asteroides infection complicated by a cerebral abscess. Treatment with amikacin for 2 weeks and ceftriaxone for 6 weeks led to complete recovery, and there was no recurrence of disease over a follow-up period of 12 months after therapy. The use of ceftriaxone in combination with amikacin might significantly shorten the duration of treatment for patients with disseminated nocardiosis. This combination of antibiotics merits further investigation with use of a larger sample of patients.


Cryptosporidiosis in patients infected with the human immunodeficiency virus.

Infection caused by Cryptosporidium species has proved to be one of the most taxing and frustrating conditions faced by clinicians caring for patients with AIDS. Unfortunately, this unique organism, which was identified as a human pathogen only shortly before the AIDS epidemic began to manifest itself, has received only minimal attention during the past decade. Dr. Carolyn Petersen, an assistant professor of medicine at the University of California, San Francisco, and a member of the Division of Infectious Diseases at San Francisco General Hospital, is a molecular parasitologist whose investigative career is focused on elucidating the biology of Cryptosporidium species. In this AIDS Commentary Dr. Petersen provides an update on recent developments in this field.


Disseminated candidiasis in addicts who use brown heroin: report of 83 cases and review.

From November 1983 to April 1990, disseminated candidiasis was diagnosed in 83 heroin addicts at our institution. All patients had consumed brown heroin diluted in fresh lemon juice. Sixty-two (75%) had skin lesions, 41 (49%) had ocular lesions, and 35 (42%) had one or several costochondral tumors. Candida albicans was grown in culture or histopathologically identified in 34 cases (41%). The patients who had only cutaneous lesions were treated with ketoconazole, and they were all cured. The patients with ocular involvement received systemic amphotericin B with or without oral flucytosine; 29 of these patients developed varying degrees of vision loss. The method of treatment of costochondral tumors was not uniform; in 14 cases the lesions were resected. The one patient who died developed endocarditis involving the aortic valve. Cases of pleuropulmonary involvement, spondylitis, and large-joint arthritis have also been described among the 300 cases reported in the reviewed literature. This is a new syndrome of candidal infection in drug addicts who use brown heroin; ocular lesions are the most harmful manifestation, and loss of vision is the major sequela.


Cutaneous leishmaniasis: review of 59 cases seen at the National Institutes of Health.

Fifty-nine cases of cutaneous leishmaniasis seen at the National Institutes of Health in Bethesda, Maryland, are reviewed. The group of patients involved was unique in that the majority were American civilians, their disease was acquired in many different endemic areas of the world, and their illnesses represented all points on the clinical spectrum of cutaneous disease. The majority of American patients acquired leishmaniasis while engaging in activities related to their occupations. Cutaneous disease acquired in the New World usually consisted of one or two lesions, while multiple lesions often characterized Old World infections with Leishmania major. Patients with chronic relapsing or diffuse cutaneous leishmaniasis were native to endemic areas and were infected at an early age. Even the localized form of cutaneous leishmaniasis was often extensive and difficult to treat. Diagnosis with culture and identification of the parasite to the subspecies level is instrumental in the selection of optimal therapy. Cutaneous leishmaniasis may be encountered increasingly often in the United States because of the frequent international travel of U.S. residents and the influx of immigrants from endemic areas of the world.


Agrobacterium tumefaciens peritonitis mimicking tuberculosis.

Agrobacterium species have been previously implicated in the development of clinical disease. We report what we believe to be the first case of ascites caused by Agrobacterium tumefaciens in a cirrhotic patient. Since the correct diagnosis was made only after laparoscopy-guided collection of specimens from two different tissues, we suggest that Agrobacterium may be an underdiagnosed pathogen in clinical situations in which tuberculosis is considered to be the cause of high-protein ascites.


Epidemiology of infections with Pseudomonas aeruginosa in burn patients: the role of hydrotherapy.

Pseudomonas aeruginosa remains a cause of serious wound infection and mortality in burn patients. By means of restriction fragment length polymorphism analysis and a DNA probe for the pilin gene of Pseudomonas, a lethal strain of nosocomial P. aeruginosa was identified as the cause of an outbreak of wound infections among burn patients. Environmental surveys suggested an association of the outbreak with hydrotherapy provided to many patients in a common facility. In a trial of burn wound care without hydrotherapy, overall mortality was reduced significantly, mortality associated with pseudomonas sepsis was eliminated, and the strain of P. aeruginosa associated with earlier mortality was eradicated. Moreover, fewer nosocomial pseudomonas infections, lower levels of pseudomonas resistance to aminoglycoside antibiotics, significantly fewer pseudomonas infections of skin graft donor sites, and later appearance of Pseodomonas species in burn patients were found during the period when hydrotherapy was not used.


Clostridium sordellii bacteremia: case report and review.

Clostridium sordellii is a gram-positive, anaerobic bacillus that has rarely been implicated as a human pathogen. It produces several exotoxins, which contribute to the progressive edema and refractory shock frequently seen with human infection. There have been eight prior reports of bacteremic C. sordellii infection and seven prior reports of nonbacteremic infections not due to myonecrosis of skeletal muscle. Mortality was 50% in the bacteremic group and 71% in the nonbacteremic group. Mortality correlated with both shock and leukemoid reaction at presentation. We present a case of C. sordellii sepsis in an asplenic patient with sickle beta thalassemia and inflammatory bowel disease, and we review the literature.


Aerobic and anaerobic microbiology of external otitis.

Microbiological and clinical data from 46 patients with external otitis were retrospectively evaluated. Specimens were processed for isolation of aerobic and anaerobic bacteria. A total of 42 aerobic bacteria, 22 anaerobic bacteria, and 3 Candida albicans organisms were recovered. Aerobic bacteria only were isolated from 31 patients (67%), anaerobic bacteria only were isolated from 8 (17%), and mixed aerobic and anaerobic bacteria were isolated from 4 (9%). C. albicans alone was recovered from two patients (4%), and the organism was mixed with aerobic bacteria in one case (2%). The most common isolates were Pseudomonas aeruginosa (19 instances), Peptostreptococcus species (11), Staphylococcus aureus (7), and Bacteroides species (5). One isolate was recovered from 30 patients (65%), 2 isolates were recovered from 11 (24%), and 3 isolates were recovered from 5 (11%). beta-Lactamase activity was detected in 32 isolates recovered from 27 specimens (59%). These data illustrate the polymicrobial nature of external otitis in one-third of the patients and the role of anaerobic bacteria in one-quarter of them. Further prospective studies are warranted for evaluating the role of anaerobic bacteria in this infection and the therapeutic implications of these findings.


Protothecosis complicating prolonged endotracheal intubation: case report and literature review.

Prototheca species are ubiquitous, aerobic, unicellular algae closely related to the green algae Chlorella. Their involvement in human disease--in both immunocompetent and immunocompromised patients--has been reported with increasing frequency. The wide array of presentations has included cutaneous, subcutaneous, mucosal, bursal, catheter-related, and (in rare instances) systemic disease. We report a case of protothecosis complicating prolonged endotracheal intubation presenting as a nasopharyngeal ulceration with a soft-tissue mass, and we review the presentation, treatment, and outcome of the 59 previously reported cases of protothecosis. Optimal therapy for protothecosis includes excision (where possible) followed by systemic administration of amphotericin B; the sole exception is in the case of olecranon bursitis, where excision alone appears curative. The role of the newer imidazoles is yet to be determined.


Fever of unknown origin: review of 86 patients treated in community hospitals.

This study describes the clinical features of fever of unknown origin (FUO) in 86 patients in a community setting from 1984 to 1990. Infectious diseases remain the most common category of illnesses causing FUO; in this study, infectious diseases including recently described diseases--such as AIDS (three cases) and Lyme disease (one case)--caused FUO in 28 patients. Although percutaneous computed tomography-guided procedures were useful for obtaining diagnostic specimens (15 cases), a noninvasive approach established the diagnosis in many instances (37 cases). In all but nine cases, diagnostic testing was guided by abnormalities detected during the physical examination or routine laboratory tests.


Systemic manifestations of invasive amebiasis.

Eighty-four patients with serious infection due to Entamoeba histolytica were evaluated for systemic complications by objective criteria for dysfunction of the organ systems normally assessed in surgical sepsis. Of 71 patients with amebic liver abscess (ALA), 41% had systemic complications and 13% had more than one organ system involved. Patients > or = 40 years of age and those being treated with steroids were at significantly increased risk of developing complications (P < or = .05). The erythrocyte sedimentation rate and the levels of the acute-phase markers C-reactive protein (CRP) and serum amyloid A (SAA) were significantly elevated in patients with ALA over values in those without ALA (P < or = .05). ALA patients with complications had lower CRP and SAA concentrations than those without complications (P < or = .05). Blood and liver aspirates in ALA patients were usually bacteriologically sterile. The pathogenesis of systemic complications and the associated acute-phase response requires further study, and ways of predicting disease severity and intervening therapeutically must be devised.


Persistent fever in association with infective endocarditis.

Fever persisting despite adequate antimicrobial therapy for endocarditis can be an ominous sign. To evaluate the significance of persistent fever in this situation, we reviewed the records of patients at three hospital affiliates of Albert Einstein College of Medicine. Twenty-six patients with 27 episodes of endocarditis and fever lasting for > or = 2 weeks despite appropriate antimicrobial therapy were identified and compared with a matched cohort of 26 patients with endocarditis but without prolonged fever. The median duration of fever in the former group was 35 days. Cardiac infection caused fever in 13 of these patients, seven of whom had myocardial abscesses. Additional causes of infection included drug treatment, nosocomial transmission of pathogens, and pulmonary emboli. Sixteen patients required cardiac surgery (seven on an emergent basis), whereas only two controls underwent such a procedure (P < .001). Twenty-two patients with persistent fever and five controls developed nosocomial complications (P < .001). Six patients with fever died, five from endocarditis-related complications. Thus persistent fever often indicates complicated endocarditis. We present an approach for the evaluation of the patients affected by this condition.


Septicemia in 980 patients at a university hospital in Berlin: prospective studies during 4 selected years between 1979 and 1989.

A total of 980 episodes of clinically and bacteriologically proven septicemia were included in four prospective 1-year studies at a 1,300-bed university hospital in Berlin between 1979 and 1989. The incidence was 8.1 per 1,000 admissions. The percentage of patients with severe underlying diseases increased significantly from 67% to 95% over the decade. Septicemia due to gram-positive bacteria decreased from 47.3% in 1979 to 43.7% in 1986 and increased again to 51.2% in 1989. Septicemia due to gram-negative organisms decreased constantly from 45.0% in 1979 to 39.8% in 1989. The most frequently isolated species were Escherichia coli (26.4%), Staphylococcus aureus (18.9%), coagulase-negative staphylococci (10.2%), enterococci (7.7%), viridans streptococci (6.4%), Klebsiella species (5.5%), and pneumococci (5.0%). The overall mortality rate decreased significantly from 33.6% in 1979 to 20.8% in 1989. Mortality for episodes of septicemia due to gram-positive bacteria (25.5%) was higher than that for septicemia due to gram-negative bacteria (18.3%). Mortality rates associated with polymicrobic and fungal septicemia were higher than the overall mortality rate.


Biocompatibility of particulate polymethylmethacrylate bone cements: a comparative study in vitro and in vivo.

The biocompatibility of particles of four different commercial polymethylmethacrylate (PMMA) bone cements was evaluated by exposing human synovial fibroblasts and mouse peritoneal macrophages to particles of cement. Cell integrity and inflammatory potential were assessed using enzyme release and microscopical examination. Results suggested the occurrence of cell damage in both cell types and macrophage studies indicated inflammatory potential. The response in vivo was investigated by intra-articular injection of the particles into mouse knee joints. Clinical and histological evaluation was performed over 2-52 wk. Particles of all four cements were well tolerated in the joints.


Cellular reaction on the intraperitoneal injection of four types of polylactide particulates.

Four types of polylactide particulates, P-L-LA 100, 250, 550 KD and a P-DL-LA 400 KD were injected into the peritoneal cavity of mice. The inflammatory reaction showed an increase in cell number (mainly neutrophilic granulocytes) up to 48 h after which the cell numbers decreased below the control (phosphate-buffered saline). All four polylactide particulates aggregated and intermingled with inflammatory cells. The aggregates remained throughout the investigation period of 6 months. Quantitative measurements showed that standardization of the particle form and size is essential. From this study and other experiments in which calcium phosphates and asbestos were injected intraperitoneally, it is concluded that the inflammatory response observed in the peritoneal cavity is related to the type of material injected and probably to form and size of the individual particles, but not to molecular weight.


Permucosal implantation pilot study with HA-coated dental implant in dogs.

The histological response of transmucosal one-stage titanium dental implants coated with hydroxyapatite is described. The gingival adhesion to the implant was examined with regard to coated, partially coated or non-coated surfaces in the cervical region. From each coating type, 9 implants were inserted into dogs. Six months after the insertion, 19 implants could be evaluated, but 8 implants were lost. From these 19 implants, 6 implants showed severe pockets with inflammation up to the bony tissue. The 13 successful implants showed direct bone bonding with the hydroxyapatite coating and adhesion of submucosal connective tissue to the implant surface, with inflammation. The marginal gingiva showed slight inflammation. A totally coated implant will probably introduce inflammation by debris formation against the rough implant surface more easily. The hydroxyapatite coating often disappeared in the soft tissue or in the oral cavity. Bone which directly adapted to the coating seemed to prevent it from resorption.


Modification of the biocompatible and haemocompatible properties of polymer substrates by plasma-deposited fluorocarbon coatings.

The polymerization of gases present in a low temperature plasma is a technique particularly well suited for biomedical material processing. Therefore, the possibilities this technique offers to increase the biocompatibility and haemocompatibility of polysulphone and poly(hydroxybutyrate) membranes to be used in a new bioartificial pancreas device were studied. The deposition of thin fluorocarbon coatings from an argon plasma containing perfluorohexane gave very smooth and hydrophobic surfaces without affecting the filtering properties of the treated membranes. Adding hydrogen increased the reaction yield, but gave rougher and less hydrophobic coatings. We characterized the biological properties of the treated surfaces and discussed the influence of the modified surface properties on the biological behaviour of the treated polymers. The good biocompatibility of the deposited coatings was established by following in vitro the insulin secretion of Langerhans islets cultured on the treated membranes and by examining the fibrous capsule that developed on plasma-treated polymer disks after three months of in vivo incubation in the peritoneum of Wistar rats. Rough and haemocompatible films of poly(hydroxybutyrate) and smoother, but more thrombogenic, polysulphone films were treated by perfluorohexane and perfluorohexane + H2 plasmas to study the relative influence of surface roughness and surface energy on polymer thrombogenicity. In vitro protein adsorption and total blood clotting tests proved that the surface roughness influences the thrombogenicity more than the other surface properties. This study seems to show that the plasma deposition of smooth and hydrophobic fluorocarbon coatings can increase the biocompatibility and reduce the surface thrombogenicity of the treated membranes without affecting their filtering properties.


Human serum albumin as a probe for surface conditioning (opsonization) of block copolymer-coated microspheres.

The adsorption of human serum albumin to polystyrene microspheres sterically stabilized with block copolymers, was investigated using photon correlation spectroscopy and laser doppler anenometry. The block copolymers used were non-ionic surfactants of the poloxamer and poloxamine series made of polyoxyethylene and polyoxypropylene chains. Photon correlation spectroscopy and laser doppler anenometry showed that the coating reduced the adsorption of the protein to the polystyrene microspheres surface. Quantitative studies using 125I-labelled human serum albumin and sodium dodecyl sulphate-polyacrylamide gels (in combination with densitometry), were employed to evaluate the adsorption of human serum albumin to uncoated and coated polystyrene microspheres. They confirmed that the hydrophilic polyoxyethylene steric layer, created by coating with the block copolymers, reduced the adsorption of human serum albumin. Moreover, the amount of human serum albumin adsorbed was related to the polyoxyethylene content of the block copolymers.


Controlled drug release from implantable matrices based on hydrophobic polymers.

Reports on the controlled release of drugs, including macromolecular drugs, from silicone elastomers and ethylene-vinyl acetate copolymers, based on the formation of channels and cracks in the polymer, are reviewed. Aqueous interconnected pores are produced by osmotically active additives or by using loads of water-soluble drugs exceeding the percolation threshold. The release is generally proportional to the square root of time (t1/2). Nevertheless, pseudo-zero-order release kinetics can be obtained by adequately controlling the formulation variables. The factors controlling the release pattern and rate are discussed. In vivo applications of these types of systems are also considered.


Antibacterial activity and mutagenicity studies of water-soluble phosphazene high polymers.

Eight water-soluble phosphazene high polymers, [NPR2]n (R, organic, water-solubilizing side-group; n, approx: 15,000) and the small-molecule counterparts of the polymers were examined for antibacterial activity against six different strains of bacteria (Escherichia coli, Salmonella typhimurium (TA 100), Salmonella pullorum, Streptococcus faecalis, Bacillus subtilis and Pseudomonas aeruginosa). Antibacterial testing was carried out by measuring zones of inhibition and changes in solution turbidity over time. In addition, the antibacterial activity of the surfaces of cross-linked poly[di(methoxyethoxyethoxy)phosphazene] (MEEP) hydrogels were investigated. A number of the high polymers, as well as the MEEP hydrogels, impeded bacterial growth. Only E. coli was unaffected by the phosphazenes. A possible explanation for the antibacterial character of the polymers is presented. The same compounds were monitored for potential mutagenic activity using the Salmonella typhimurium tester strains TA 100 and TA 98. None of the high polymers or their small-molecule analogues showed mutagenic activity in either strain of Salmonella at the concentrations tested. The use of these materials as coatings for artificial implants is discussed.


Poly(ethylene oxide)-graft-poly(L-lysine) copolymers to enhance the biocompatibility of poly(L-lysine)-alginate microcapsule membranes.

A graft copolymer having poly(L-lysine) (PLL) as the backbone and monomethoxy poly(ethylene glycol) (MPEG) as pendent chains was synthesized. This polycationic copolymer was used to form microcapsules with sodium alginate, a polyanion. Microcapsules and model surfaces formed with PLL-graft-MPEG demonstrated reduced protein adsorption, complement binding and cell adhesion in vitro compared to materials with unmodified PLL. Microcapsules with PLL-g-MPEG on the surface were seen to be much more biocompatible than the widely used alginate/PLL/alginate microcapsule in a mouse intraperitoneal implant model. The graft copolymers demonstrated a lower affinity for alginate and increased microcapsule permeability more than PLL. To correct this, pentalayered alginate/PLL/alginate/PLL-g-MPEG/alginate microcapsules were fabricated, and these demonstrated both appropriate permselectivity and enhanced biocompatibility.


Epifluorescent video microscopy (EVM) for platelet-biomaterial interactions: elimination of photoactivation and dye effects.

The use of two intracellular dyes for epifluorescent video microscopy (EVM) in observations of cell-surface interactions is evaluated and discussed. This methodology permits determinations of cell adhesion, detachment and movement at the surfaces of biomaterials in the presence of flow and physiological haematocrit. Two tests, one which examines for the effect of incident light on platelet adhesion and one which checks for sufficient light for accurate observation of cells, have been designed. Evaluations were made of the adhesion of platelets labelled with the fluorescent dyes mepacrine and acridine orange, used singly and in combination. The use of a number of light level-dye level combinations with glass and several polymers and the addition of a plasma level of fibrinogen did not show any photoactivation effects. This methodology paves the way for longer than previous exposures to light with our system, from 1 min up to 30 min now. Washed platelet suspensions are preferred; these allow for the selective labelling of specific cells and the removal of dye from the surface of the cell.


Effect of electrostatic forces on the dynamic rheological properties of injectable collagen biomaterials.

Injectable collagen is a concentrated dispersion of phase-separated collagen fibres in aqueous solution used to correct dermal contour defects through intradermal injection. The effect of electrostatic forces on the rheology of injectable collagen was studied by observation of the birefringence of collagen fibres through a polarizing microscope as well as by oscillatory rheological measurements on dispersions of varying ionic strengths (0.06-0.30). The birefringence of fibres progressively increased as ionic strength was reduced from 0.30 to 0.06. The linear viscoelastic measurements displayed a logarithmic relationship between storage (and loss) moduli and frequency over oscillation frequencies of 0.1-100 rad/s. The associated relaxation time spectra, interpreted using the theory of Kamphuis et al. for concentrated dispersions, show that collagen fibres become more flexible as ionic strength increases. This result was analysed at the molecular level from the perspective that collagen fibres are a liquid-crystalline phase of rigid rod collagen molecules which have phase-separated from solution. Electrostatic forces affect the volume fraction of water present in the collagen fibres which in turn alters the rigidity of the fibres. Flexible collagen fibre dispersions displayed emulsion-like flow properties whereas more rigid collagen fibre dispersions displayed suspension-like flow properties. Changes in fibre rigidity significantly alter the injectability of collagen dispersions which is critical in clinical performance.


Central connections of trigeminal primary afferent neurons: topographical and functional considerations.

This article reviews literature relating to the central projection of primary afferent neurons of the trigeminal nerve. After a brief description of the major nuclei associated with the trigeminal nerve, the presentation reviews several early issues related to theories of trigeminal organization including modality and somatotopic representation. Recent studies directed toward further definition of central projection patterns of single nerve branches or nerves supplying specific oral and facial tissues are considered together with data from intraaxonal and intracellular studies that define the projection patterns of single fibers. A presentation of recent immunocytochemical data related to primary afferent fibers is described. Finally, several insights that recent studies shed on early theories of trigeminal input are assessed.


Recent advances in primary palate and midface morphogenesis research.

During the sixth week of human development, the primary palate develops as facial prominences enlarge around the nasal pits to form the premaxillary region. Growth of craniofacial components changes facial morphology and affects the extent of contact between the facial prominences. Our recent studies have focused on developing methods to analyze growth of the primary palate and the craniofacial complex to define morphological phases of normal development and to determine alterations leading to cleft lip malformation. Analysis of human embryos in the Carnegie Embryology Collection and mouse embryos of cleft lip and noncleft strains showed that human and mouse embryos have similar phases of primary palate development: first, an epithelial seam, the nasal fin, forms; then a mesenchymal bridge develops through the nasal fin and enlarges rapidly. A robust mesenchymal bridge must form between the facial prominences before advancing midfacial growth patterns tend to separate the facial components as the medial nasal region narrows and elongates, the nasal pits narrow, and the primary choanae (posterior nares) open posterior to the primary palate. In mouse strains with cleft lip gene, maxillary growth, nasal fin formation, and mesenchymal replacement of the nasal fin were all delayed compared with noncleft strains of mice. Successful primary palate formation involves a sequence of local cellular events that are closely timed with spatial changes associated with craniofacial growth that must occur within a critical developmental period.


Salivary gland function and aging: a model for studying the interaction of aging and systemic disease.

This review describes an approach to examining the interaction of aging and systemic disease on a key aspect of oral physiology, salivation. The approach requires several steps: defining general health, and a specific physiological function, at different ages; defining a disease of interest and the influence of the disease on the specific physiological function; and determining if the disease can affect performance of the physiological function with increased age.


Mesoderm and jaw development in vertebrates: the role of growth factors.

The head and neck arise during development as the result of a complex series of cellular and molecular interactions that begin in the fertilized egg. In this article, the role of an important class of molecules, growth factors, is examined in two main steps of the developmental sequence: the initial induction of mesoderm and the later induction of jaw cartilage and bone. The article focuses particularly on the roles of members of the transforming growth factor-beta (TGF-beta), fibroblast growth factor (FGF), and epithelial growth factor (EGF) families in these processes and current models of growth factor involvement. Possible experiments for the future are discussed.


Artificial cells in immobilization biotechnology.

Artificial cells contain biologically active materials. Artificial cells containing adsorbents have been a routine form of treatment in hemoperfusion for patients. This includes acute poisoning, high blood aluminum and iron, and supplement to dialysis in kidney failure. Artificial cells are being tested for use as red blood cell substitutes. Artificial cells encapsulated cell culture are being tested in animals for the treatment of diabetes and liver failure. A novel 2 step method has prevented xenograft rejection. Artificial cells containing enzymes are being studied for treatment in hereditary enzyme deficiency diseases and other diseases. Recent demonstration of extensive enterorecirculation of amino acids in the intestine has allowed its oral administration to deplete specific amino acids. Artificial cells containing complex enzyme system convert wastes like urea and ammonia into essential amino acids. Artificial cell is being used for the production of monoclonal antibodies, interferons and other biotechnological products. It is also being investigated for drug delivery, and for use in other applications in biotechnology, chemical engineering and medicine.


Semi-selective immunoadsorption treatment in myasthenia gravis.

We treated some patients with severe generalized myasthenia gravis (MG) with semi-selective adsorption by plasma perfusion(PP) on IM-T350 column. We studied efficiency and undesired effects of the employed material to verify the safety and effectiveness of methodology. Thus a treatment protocol has been prepared: 6 PP per treatment cycle; 3-7 days between PP sessions; plasma to treat per session 1,2 lt; only one adsorbent column for every cycle. Our experience, on 13 cycles of treatment, showed that PP in autoimmune MG has provided good clinical results without any improvement in genetic MG.


Phosphate removal by resin hemoperfusion efficacy and biocompatibility of a new exchange resin.

A new coated anionic exchange resin for blood purification specifically designed to remove phosphates was experimentally employed in animals. 3 pigs, in which uremia had been surgically induced, underwent 6 extracorporeal hemoperfusion sessions (2 per pig) with a cartridge containing 100 gr of resin. The phosphate clearance proved satisfactory, values being 120 ml/min after 10' and around 80 ml/min after 2 hours. The biocompatibility of the resin and of the coating membrane was satisfactory. The negligible variation in pH and plasma bicarbonate during all sessions confirmed the low absorption by the tested resin of other blood anions competing with phosphate.


Adsorption treatment in hyperlipidemia.

Clinical LDL- adsorption has made overwhelming progress in recent years. Thus, immunoadsorption with polyclonal anti-LDL-antibodies and chemoadsorption using the dextransulfate system have been applied successfully for selective LDL-removal in routine clinical plasma treatment of hypercholesterolemic patients suffering from coronary artery disease. Clinical pilot studies showed good results using immunoadsorption with anti-LDL-F(ab)-columns. Promising laboratory results have been achieved using immunoadsorption with monoclonal LDL-antibodies, polyacrylate/fractogel and macroporous cellulose. While these systems need a primary separation step for cells and plasma, the integrated plasma separation/adsorption device and the LDL-hemoperfusion module using E 280590 can even treat whole blood. Thus, these systems hold a great promise for the future.


Extracorporeal elimination of carbamazepine by haemoperfusion.

The clearance of the haemoperfusion cartridge H 200 C Tekin was determined in vitro for blood dissolved carbamazepine to test the efficiency of haemoperfusion treatment. In the first hour of in vitro haemoperfusion test of pig blood (concentration of carbamazepine 1084 mg/l) a clearance of 190 ml/min was obtained, which declined slightly to 150 ml/min after 4 hours. In a second test the elimination of carbamazepine from albumin solution was proven to be more than 1000 mg of carbamazepine after a four hour perfusion period using teflon coated charcoal. These results are in accordance with the clinical course of a 30 years old male patient who presented with severe hypoxaemia due to acute intoxication by 6 g carbamazepine. The carbamazepine blood level was 33.7 mg/l. Charcoal haemoperfusion was started immediately. A dramatic improvement of clinical symptoms was already observed during haemoperfusion. The carbamazepine serum level was 18.6 mg/l after 4 hours of haemoperfusion.


Hybrid artificial pancreas: islet transplantation inside membrane bioreactors.

The use of pancreatic islet transplantation in membrane bioreactors put in vascular circuits aims at resetting the glucose homeostasis in diabetic or pancreatectomized patients, avoiding immune host rejection. Our experience was carried out at following stages: porcine pancreas explantation and enzymatic separation of endocrine tissue from exocrine fraction by collagenase; evaluation of islet functionality (culture tests); in vitro tests of the islets-bioreactor system, to assess the metabolic response to the glucose; in vivo evaluation to assay the haemodynamic behaviour. The trials showed a good metabolic bioreactor functionality and a decreasing incidence of coagulative problems.


Treatment of uraemic hyperphosphatemia with calcium acetate: a safe alternative to calcium carbonate.

Clinical usefulness of calcium acetate (CAA) as phosphorus binder was assessed in 19 stable hemodialysis patients with persistent hyperphosphatemia. All were dialysed thrice weekly with a constant dialytic schedule and a dialysate calcium of 3.5 mEq/l. One month prior the study beginning all patients stopped assumption of Ca and vitamin D supplements. In the first period of the study CAA (mean daily doses 2.2 g) was administered for one month followed by 15 days of withdrawal. The mean serum phosphorus decreased from 7.6 +/- 1.4 to 5.8 +/- 0.8 mg% (p < 0.005). After 15 days of withdrawal mean serum phosphorus reached the pretreatment value. Then the patients entered a long term study with personalized doses of CAA (between 1 and 4 g/day) and administration in 8 of them of alpha-calcidol. After a mean follow-up period of 5.4 +/- 1.5 months serum phosphorus was reduced from 7.5 +/- 1.1 to 5.6 +/- 1.1 mg% (p < 0.0005) while calcemia increased from 9.0 +/- 0.7 to 9.6 +/- 0.6 mg% (p < 0.005). Only one patient developed mild hypercalcemia. We concluded that CAA is a safe alternative to calcium carbonate for the control of hyperphosphatemia of uraemic patients for the most efficient phosphorus binding and the lesser absorption of calcium.


DNA-coated carbon adsorbents experimental assessment and results of severe psoriasis treatment.

Newly developed combined adsorbents, containing from 1 to 8 mg of thymic DNA per 1 g of the granulated or the fibrous carbonic matrix, demonstrated good biocompatibility and selectivity for DNA- and DNP-binding substances. In a group of 14 patients with severe psoriasis (uncontrol trial) a single hemoperfusion procedure through DNA-coated granulated synthetic carbons (perfusion volume was 2.5-3.1 L, sorbent quantity was 30 g) resulted in complete remission in 6 patients and in substantial improvement of clinical status in 6 other patients. A positive effect was observed in 4 patients during 7-11 months; in 8 patients it is being observed for more than 29-33 months. The double-blind tests in the group of patients subjected to hemoperfusion through the DNA-coated charcoal (27 people) and uncoated charcoal (9 people) showed the full-scale remission in 55.5% and 11.2% respectively. The authors believe that the DNA-coated activated carbons can be an effective therapeutic procedure for treatment of numerous immuno-dependent diseases and states associated with disorders in the kinetics of cell division.


Removal of beta 2-microglobulin by hemodialysis and hemofiltration: a four year follow up.

Efficient removal of total body burden beta 2-Microglobulin (beta 2-M) in uremia is a continuing challenge, as dialysis-related amyloidosis represents a major complication of chronic renal replacement therapy. To investigate long-term beta 2-M removal we studied 3 groups of stable end-staged renal failure patients over a period of 4 years; we compared low flux (cuprophane) hemodialysis (n = 12), high flux (polysulfone) hemodialysis (n = 12) and hemofiltration using high flux polysulfone (n = 8). In contrast to the cuprophane membrane, the polysulfone membrane eliminated considerable amounts of beta 2-M. This was associated with a sustained reduction of predialysis serum beta 2-M-levels (by 20%). Compared with high flux hemodialysis, hemofiltration provided a 50% higher elimination of beta 2-M. Thus, our long-term evaluation of beta 2-M removal suggests that hemofiltration rather than hemodialysis may be the treatment of choice for delaying the incidence of dialysis-related amyloidosis.


The whole molecule design approach to drug discovery.

An approach to drug design and lead generation is presented which attempts to retain the qualities of traditional drug design and leave control with the bench scientist, whilst harnessing the power of computers to handle the combinatorial explosion of ideas. A computer language, ALEMBIC, is used to collate the ideas of the scientists. The resulting list of potential molecules is then parameterised using whole molecule descriptors. Based on these descriptors, appropriate statistical techniques are used to generate sets of molecules retaining the maximum amount of the information inherent in all possible combinations of the scientists ideas.


Synthesis and ACE inhibitory activity of the stereoisomers of perindopril (S 9490) and perindoprilate (S 9780).

Perindopril, a powerful ACE inhibitor contains 5 chiral carbons, thus there is the possibility of 2(5) = 32 stereoisomers for the general structure 1. These 32 stereoisomers were synthesized by cross-coupling the 8 stereoisomers of perhydroindole 2-carboxylic acid benzylester with the 4 stereoisomers of 2-(1-carbethoxybutylamino) propionic acid 4, then hydrogenating the resulting benzylesters. Each stereoisomer of perindopril furnished by saponification the corresponding diacid stereoisomer 2 of perindoprilate which is the active form of perindopril. For each of the 32 stereoisomers 2 the in vitro ACE inhibitory potency (IC50) was determined. Four of them, including perindoprilate, had activities in the nanomolar range, and four more were ca. 10 x less active. The four acid esters 1 corresponding respectively to the four most active diacids 2 in vitro were studied (1 mg/kg via the oral route) for their in vivo activity in dogs. It could be concluded that p.o. absorption of the active acid esters 1 and their activation to the active diacid 2 depended only on the chiralities of the two ring junction carbons of the perhydroindole ring.


Molecular modeling of adenosine receptors. I. The ligand binding site on the A1 receptor.

The amino acid sequence of the canine adenosine A1 receptor and the atomic coordinates of a structurally related protein, bacteriorhodopsin, were combined to generate a three-dimensional model for the adenosine A1 receptor. This model consists of seven amphipathic alpha-helices, forming a pore that has a rather distinct partition between hydrophobic and hydrophilic regions. Subsequently, a highly potent and selective ligand, N6-cyclopentyladenosine, was docked into this cavity. A binding site is proposed that takes into account the conformational characteristics of the ligand, obtained from indirect modeling studies by the 'active analog approach'. Moreover, it involves two histidine residues that were shown to be important for ligand coordination from chemical modification studies. Finally, the deduced binding site was used to model other potent ligands that could all be accommodated consistent with earlier biochemical and pharmacological findings.


Synthesis and antitumor activity of poly(ethylene glycol)s linked to 5-fluorouracil via a urethane or urea bond.

In order to provide a macromolecular prodrug of 5-fluorouracil (5FU) with reduced side-effects and exhibiting strong antitumor activity, 5FU was covalently linked to poly(ethylene glycol) (PEG) via a urethane or urea bond. For the purpose of evaluating the release behavior of 5FU, the hydrolysis of the urethane or urea bond in the obtained conjugate of PEG-end capped with 5FU was investigated in vitro at 37 degrees C in aqueous solution media. The survival effect for the conjugate was assessed in vivo against p388 lymphocytic leukemia in female CDF1 mice by intraperitoneal (i.p.) transplantation/i.p. injection. The effects of a hydrophobic hexamethylene spacer group, the end group and the number n of ethylene oxide (EO) units in PEG on the release behavior of 5FU and the survival effect were investigated. The release rate of 5FU from the 5FU-terminated PEG conjugates via urethane or urea bond was very fast. However, it became slow with increasing n of EO units in PEG and was depressed by the introduction of hydrophobic spacer group. The 5FU-terminated PEG conjugates obtained exhibited significant survival effects against p388 leukemia mice i.p./i.p. Especially, the methoxy PEG (n = 113)/urethane/hexamethylene/urea/5FU conjugate showed the strongest survival effect among the synthesized 5FU-capped PEG conjugates via urethane or urea bond compared to free 5FU against p388 leukemia mice. These conjugates obtained did not display an acute toxicity even in high dose ranges.


[Functional capacity of patients over 65 according to the Katz index. Reliability of the method].

To find functional capacity of elderly patients by using the Katz Index, which measures the "Basic Activities of Daily Life". To examine how reliable the Katz Index is in the Primary Care field. Descriptive crossover study. Assessment of the measuring instrument's internal consistency and reliability, using a test-retest method. SITE. Otero Health Area, in Oviedo. This was a random sample, ordered by age and gender, of people over 65 who used the Otero Health centre. The elderly population made up 12.86% of our base Health Area. The Katz Index showed that 81.6% of the elderly in the sample were independent in all their functions. A significant age difference was found. Internal consistency was measured by Cronbach's Alpha and was 0.8612 for nurses and 0.8362 for doctors. The correlation coefficient within Sperman's estimated observer was r = 0.936074. Kappa was 0.8436 (p < 0.0000) for the different observers, i.e. nurses or doctors. Most of the elderly patients in our base Health area possess full functional capacity, at the level of the routine activities of their daily life. The Katz Index is a reliable method of evaluating elderly peoples' functional capacity, at the Primary Care level. It can be used by any professional in the health team.


[Calibration and safety of sphygmomanometers in health centers of Murcia].

To find the levels of calibration, safety and physical faults in the sphygmomanometers used in Health Centres. A multi-centre, observational, descriptive and crossover study. SITE. 7 Health Centres in the urban zone of Murcia and nearby towns. All the sphygmomanometers in the 7 Health Centres, that is 80 aneroid and 62 mercury ones. It needs high-lighting that 43.7% of the aneroid sphygmomanometers and 12.3% of the mercury ones were wrongly calibrated. We noted that 20% of the measurements made with wrongly calibrated aneroid sphygmomanometers had a margin of error above +/- 8 mmHg, with a tendency towards under-registering. We found very significant differences between the average arterial pressure measured by aneroid instruments as against mercury ones (p < 0.001). The number of sphygmomanometers which show a margin of error greater than +/- 3 mmHg at every level of pressure is significantly greater in aneroid instruments than in mercury ones (p < 0.001). We found physical faults in 23.9% of the instruments. We consider the number of wrongly calibrated aneroid sphygmomanometers to be excessively high. Therefore mercury ones should be used whenever possible and particularly during diagnosis and treatment. Both aneroid and mercury sphygmomanometers should be regularly checked, using a proper procedure. Mercury instruments set at zero are correctly calibrated and can be used as a model for checking aneroid ones.


[Oral-dental hygiene habits in 12-year-old schoolchildren of the city of Huesca].

To study dental hygiene habits and their relationship to dental caries. A descriptive epidemiological crossover study, based on observation. SITE. Schools in the city of Huesca. Every child in the 7th grade (i.e. 12 years old) of Basic General Education (EGB) in the city of Huesca. We found that only 43.83% of the children (689 in all: 340 male, 349 female) brushed their teeth every day. 49.20% had had a dental check-up during the previous year. We noted a significant link between cariogenic factors, especially those to do with tooth-brushing and the consumption of sweets, and the actual existence of caries. There was a notable lack of dental hygiene, both around the questions of using a tooth-brush and in eating habits. There was little primary (the use of fluorides or dental hygiene) or secondary prevention (programmes of odonto-stomatologic care).