Formation of viscoelastic protein layers on polymeric surfaces relevant to platelet adhesion

The hemocompatibility of biomaterials is highly dependent on the adhesion and activation of platelets. Surface-adsorbed fibrinogen has a dominant role in promoting platelet adhesion to artificial surfaces by binding glycoprotein IIb-IIIa (GPIIb-IIIa), the major platelet membrane receptor. Using quartz crystal microbalance with dissipation monitoring (QCM-D), we have investigated the material-dependent binding kinetics of purified GPIIb-IIIa to polymer-adsorbed fibrinogen. The following ranking of polymer-adsorbed mass (fibrinogen and GPIIb-IIIa) to test polymers could be established: poly[desaminotyrosyl-tyrosine ethyl (DTE) carbonate]/poly(lactide-co-glycolide)>poly[DTE co-5% poly(ethylene glycol) carbonate]. The QCM-D fibrinogen adsorption data were confirmed using an immunofluorescence assay. A synthetic RGD-containing peptide, but not a control peptide, inhibited GPIIb-IIIa binding to polymer-adsorbed fibrinogen, demonstrating the specificity of binding. Importantly, the binding efficiency of purified GPIIb-IIIa to polymer-adsorbed fibrinogen correlated with increased platelet adhesion in an in vitro model. Theoretical simulations using a Voight-based model provided quantitative data on the thickness and viscoelastic properties of the polymer-adsorbed protein layers. The precision of the modeling technique was limited with respect to the shear moduli values, leading to large variations. However, the other modeling parameters showed reproducible results. The thickness of both protein layers was polymer-dependent and ranged from 5 to 35 nm and the viscosity from 0.001 to 0.005 kg/ms, whereas the protein layer densities showed little differences between the test polymers. These results suggest that material-dependent changes in the thickness and viscoelastic properties of adsorbed fibrinogen-GPIIb-IIIa layers are crucial factors in the binding behavior of platelets to biomaterials.     

Hemocompatibility of PMEA coated oxygenators used for extracorporeal circulation procedures

An inflammatory response to cardiopulmonary bypass (CPB) caused by bioincompatibility of extracorporeal circuits is one of the major clinical issues in cardiac surgery. Recently a new coating material, poly-2-methoxyethylacrylate (PMEA), was developed to improve the biocompatibility of blood contacting surfaces. In a simulated cardiopulmonary bypass model, using fresh human whole blood, 15 membrane oxygenators (Capiox SX18, Terumo Corp., Tokyo, Japan) were compared. Five of them had the PMEA coating, five had a heparin-coated surface, and five had no surface treatment. Blood samples were taken at several time-points during a 90 minute circulation period. Changes in coagulation, complement, and blood cell alteration factors were measured by ELISA methods, plasma bradykinin levels were measured by radioimmunoassay, and expression of genes encoding cytokines TNF-alpha, interleukin-1beta, interleukin-6, and interleukin-8 was determined by semiquantitative real time RT-PCR. Platelet adhesion was significantly reduced in both the PMEA and the heparin coated circuits. Release of platelet activation marker beta-thromboglobulin was significantly higher in the uncoated control group (p < 0.01). After 5 minutes of blood circulation bradykinin levels significantly increased in all three groups (p < 0.01); however, the group with the PMEA coated oxygenators showed the lowest values. Expression of genes encoding proinflammatory cytokines in monocytes was increased in all groups, with the lowest being in the PMEA coated group. PMEA coated CPB surfaces in an in vitro experimental model showed an improved thrombogenicity, reduced bradykinin release, less platelet activation and less proinflammatory cytokines gene expression in comparison with a noncoated group. The authors assume that PMEA coating may ameliorate some of intra- and postperfusion syndromes, particularly hypotension, unspecific inflammation, hyperfibrinolysis, and blood loss.     

Long-term effects of transdermal and oral estrogens on serum lipids and lipoproteins in postmenopausal women

The transdermal and oral administration of estrogens for one year were compared with respect to the effects on lipid metabolism. Eighty-one postmenopausal women (1.5-3 years after menopause) were randomly divided into three groups. The first two groups received sequential estrogen treatment with either transdermal estradiol (Estraderm TTS, Ciba Geigy; 50 μg/day; 24 women) or 0.625 mg/day conjugated estrogens (Premarin, Wyeth; 20 subjects), respectively. In both groups medroxyprogesterone (10 mg/day per os) was added for 12 days of each cycle. Thirty-five subjects served as control group without therapy. No significant changes in the lipid profile was observed in control subjects after 1 year of follow-up. Serum triglycerides decreased significantly (-10.9 ± 26% S.D.; P < 0.05) in transdermal treated women, whereas it slightly rose in oral estrogen group. Comparable significant decreases in total and low density lipoprotein (LDL) cholesterol (mean range -6.5/-18.0%) were observed in women on estrogen replacement therapy. High density lipoprotein (HDL) cholesterol significantly diminished in transdermal estradiol group, but it rose slightly in the oral estrogen group. Thus the fraction of HDL cholesterol over LDL cholesterol did not change in the transdermal group whereas it significantly rose in subjects treated with oral estrogens. It remains to be established to what extent these differences on lipid metabolism are relevant for the prevention of cardiovascular diseases.     

Enhanced proliferation of endogenous virus in Chinese hamster cells associated with microtubules and the mitotic apparatus of the host cell.

Chinese hamster ovary cells harbour intracytoplasmic virus-like particles of type A which are closely associated with sites of microtubule formation. We report here the enhanced proliferation of these particles and their release at the cell membrane by using either 5-bromodeoxyuridine or dibutyryl cyclic AMP. The extracellular mature particles are similar in morphology to retroviruses of type B. Close association of the type A virus precursors with microtubule organizing centres, i.e. kinetochores, centrioles and basal bodies, and with microtubules per se, is confirmed by studying the effects of the microtubule inhibitors Colcemid and vincristine sulphate. The role of microtubules in the activation and transport of the intracytoplasmic type A particles is discussed.     

Surgical treatment of portal hypertension in schistosomiasis

Patients with mansonic schistosomiasis and portal hypertension are usually young, with good liver functional reserve, huge splenomegaly, and hypersplenism detectable only by alterations seen in laboratory tests. Patients who are not operated on do not develop portosystemic encephalopathy (PSE). Angiographic alterations are characteristic and totally different from those observed in patients with liver atrophy or cirrhosis. Digestive tract hemorrhage is the most severe complication. The patient with schistosomiasis is, therefore, a good model for the study of the consequences caused by the surgical treatment of portal hypertension. The evaluation, however, of results published in the literature regarding schistosomiasis is practically impossible. In a review of 130 publications referring to 4,516 surgically treated patients, the absence of prospective or retrospective studies with adequate controls was observed, and only 3.2% of the patients were followed for 5 years. According to available data, a total of 29 different surgical techniques has been employed, 81.6% of which are represented by simple splenectomy, esophagogastric devascularization (EGDS), and splenorenal shunt (SRS). Splenectomy was the surgical procedure showing the highest rate of recurrent hemorrhage (54.3%), and the portocaval shunt had the highest PSE incidence (60%). Portosystemic encephalopathy incidence after SRS was 29%. Thus, in view of a lack of objective information, in 1977 our group started a prospective, randomized study comparing the results of the surgical procedures most widely used in our country: selective portal decompression (SPD), EGDS, and SRS. The randomization was interrupted after 94 patients had been operated on (32 SRS, 30 SPD, and 32 EGDS) because of a highly significant PSE incidence in the SRS group. Preliminary results as of January, 1984, refer to an average follow-up period of 53 months (minimum 29, maximum 77 months). In the SRS group, the late mortality rate was 18.7%, and the incidence of PSE was 31.2%. Among the patients included in this group, 18.7% had arterial hypertension. In the SPD group, late mortality occurred in 10% of the cases, and PSE in 13.3%, while a mild hyperbilirubinemia was seen in 43.3% of patients. Recurrent hemorrhage and ascites had an approximately similar incidence in the 3 groups.

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Resumen Los pacientes con esquistosomiasis mansónica e hipertensión portal generalmente son jóvenes con buena reserva funcional hepática, enorme esplenomegalia e hipersplenismo detectable solamente por las pruebas de laboratorio. Los pacientes nó operados no desarrollan encefalopatia portosistémica (EPS). Los cambios angiográficos son característicos y totalmente diferentes de los que se presentan en pacientes con cirrosis y atrofia hepática. La complicación más grave es la hemorragia digestiva. Por consiguiente, el paciente con esquistosomiasis es un buen modelo para el estudio de las consecuencias que se derivan del tratamiento quirúrgico de la hipertensión portal. Sinembargo, la evaluación de los resultados en la esquistosomiasis que han sido publicados en la literatura es prácticamente imposible. En una revisión de 130 publicaciones relativas a 4516 pacientes operados, se pudo comprobar ausencia de estudios prospectivos o retrospectivos con controles adecuados, y apenas el 3.2% de los casos fué seguido por 5 años. De acuerdo con los datos disponibles, se empleó un total de 29 técnicas quirúrgicas diferentes, 81.6% de los cuales estuvo representado por simple esplenectomía, devascularización esofagogástrica (DEG) y shunt espleno-renal (SER). La esplenectomía fué el procedimiento quirúrgico que tuvo la más alta incidencia de hemorragia recurrente (54.3%) y el shunt portacava la más alta incidencia de EPS (60%). La incidencia de EPS después de SER fué de 29%.Por consiguiente, en vista de la ausencia de información objetiva, nuestro grupo inición en 1977 un estudio prospectivo y aleatorizado para comparar los resultados de los procedimientos quirúrgicos mayormente utilizados en nuestro país: descompresión portal selectiva (DPS), DEG y SER. La aleatorización fué interrumpida después de haber operado los primeros 94 pacientes (32 SER, 30 DPS y 32 DEG) debido a una muy significativa incidencia de la EPS en el grupo con SER. Los resultados preliminares (enero de 1984) se refieren a un período promedio de seguimiento de 53 meses (mínimo 29, máximo 77 meses). En el grupo de SER la mortalidad tardía fué de 18.7% y la incidencia de EPS fué de 31.2%.El 18.7% de los pacientes incluidos en este grupo exhibió hipertensión arterial. En el grupo de DPS la mortalidad tardía fué de 10% y la incidencia de EPS de 13.3%; se observó hiperbilirrubinemia leve en 43.3% de los pacientes. La incidencia de hemorragia recurrente y de ascitis fué aproximadamente igual en los tres grupos.

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Résumé Les malades qui accusent une hypertension portale d'origine bilharzienne sont généralement jeunes. Leurs fonctions hépatiques sont bonnes mais ils présentent une splénomégalie très importante et une hypersplénisme qui se manifeste seulement par des altérations des tests biologiques. Lorsqu'ils ne sont pas opérés ils ne présentent pas de manifestations d'encéphalopathie. Les caractères angiographiques sont très particuliers et totalement différents de ceux qui sont observés chez les sujets dont le foie est atrophié ou cirrhotique. L'hémorragie digestive représente la complication la plus sévère. Le malade atteint de bilharziose hépatique est par conséquent un bon modèle d'étude des conséquences provoquées par le traitement chirurgical de l'hypertension portale. Cependant les résultats du traitement publiés dans la littérature sont ininterprétables. Dans une revue de 130 publications concernant 4516 opérations, l'absence d'études prospectives ou rétrospectives bénéficiant d'un contrôle de qualité a été constatée, 3,2% seulement des malades ayant été suivis pendant 5 ans. En tenant compte des données disponibles 29 techniques chirurgicales ont été employées, 81,6% desquelles sont repésentées par les opérations suivantes: splénectomie, devascularisation et anastomose spléno-rénale. La splénectomie a été suivie du taux le plus élevé de récidive hémorragique (54,3%) et l'anastomose portocave du taux le plus important d'encéphalopathie (60%), l'anastomose spléno-rénale n'étant suivie d'encéphalopathie que dans 29% des cas. En raison du défaut d'information objective notre équipe a entrepris en 1977 une étude prospective faite selon la méthode du choix au hasard des 3 techniques chirurgicales les plus employées au Brésil: l'anastomose spléno-rénale, la décompression portale sélective et la dévascularisation. L'étude fut interrompue alors que 94 malades avaient été opérés (32 anastomose spléno-rénales, 30 décompressions portales sélectives et 32 dévascularisations oesophagogastriques) en raison du taux particulièrement élevé de l'encéphalopathie postopératoire. Les résultats de cette étude menée jusqu'en janvier 1984 répondent à une période postopératoire moyenne de 53 mois (minimum 29, maximum 77 mois). Dans le groupe traité par anastomose spléno-rénale le taux de la mortalité tardive fut de 18,7% et celui de l'encéphalopathie de 31,2%. Parmi les malades de ce groupe, 18,7% présentaient une hypertension artérielle. Dans le groupe traité par décompression portale selective le taux de la mortalité tardive fut de 10%, celui de l'encéphalopathie de 13,3%, cependant qu'une hyperbilirubinémie fut constatée chez 43,3% de ces opères. La récidive hémorragique et l'ascite atteignent un taux très voisin pour les trois opérations.

     

Clearing the clouds: Case‐report and review of the literature

In peritoneal dialysis (PD), a cloudy dialysate is an alarming finding. Bacterial peritonitis is the most common cause, however, atypical infections and non‐infectious causes must be considered. A 46‐year‐old man presented with asthenia, paraesthesia, foamy urine and hypertension. Laboratory testing revealed severe azotaemia, anaemia, hyperkalaemia and nephrotic‐range proteinuria. Haemodialysis was started through a central venous catheter. Later, due to patient preference, a Tenckhoff catheter was inserted. Conversion to PD occurred 3 weeks later, during hospitalization for a presumed central line infection. A month later, the patient was hospitalized for neutropenic fever. He was diagnosed an acute parvovirus infection and was discharged under isoniazid for latent tuberculosis. Four months later, the patient presented with fever and a cloudy effluent. Peritoneal fluid (PF) cytology was suggestive of infectious peritonitis, but the symptoms persisted despite antibiotic therapy. Bacterial and mycological cultures were negative. No neoplastic cells were detected. Mycobacterium tuberculosis eventually grew in PF cultures, despite previous negative molecular tests. Directed therapy was then initiated with excellent response. Thus, facing a cloudy effluent, one must consider multiple aetiologies. Diagnosis of peritoneal tuberculosis is hampered by the lack of highly sensitive and specific exams. Here, diagnosis was only possible due to positive mycobacterial cultures.     

Binocular interactions and steady-state VEPs. A study in normal and defective binocular vision (Part II)

Background: Recent evidence indicates that an index of binocular activity may be found in some properties of steady-state visual evoked potentials (VEPs), such as amplitude facilitation and phase shortening. We evaluated binocular interactions with steady-state VEPs in normal subjects as well as in patients with concomitant strabismus and defective binocular vision. Methods: Steady-state (8-Hz) VEPs to counterphased sinusoidal gratings (1.2 c/deg spatial frequency) of low contrast (3.2%) were recorded in 19 esotropic patients and in 18 age-matched controls. Patients had either anomalous retinal correspondence (ARC, n =10) or suppression (n=9) in casual seeing conditions (striated glasses). In all subjects, both binocular and monocular VEPs displayed a major component at twice the stimulation frequency (second harmonic), whose amplitude and phase were measured. A binocular interaction index was obtained by comparing binocular VEPs (BVEPs) with the sum (vectorial) of the two monocular VEPs (SMVEPs). Results: In normal subjects, BVEPs were larger in amplitude than SMVEPs (facilitation), and shortened in latency (phase). On average, both ARC and suppression patients displayed loss in amplitude facilitation and absence of phase shortening. However, 50% of ARC patients showed clear VEP facilitation. In both ARC and suppression patients, the amplitude ratio BVEP/SMVEP was negatively correlated with the amount of the angle of deviation. Conclusion: These results suggest that losses in amplitude facilitation and phase shortening of binocular steady-state VEPs reflect abnormal binocular interactions associated with different forms of sensorial adaptation in concomitant strabismus.     

Regioselectivity and Enantioselectivity of Metoprolol Oxidation by Two Variants of cDNA-Expressed P4502D6

Purpose. The oxidative metabolism of metoprolol was investigated in two human lymphoblastoma cell-lines transfected with variants of cDNA for cytochrome P4502D6. Methods. The regioselective and enantioselective features of the oxidations of deuterium-labeled pseudoracemic metoprolol were characterized by GC/MS analysis of the substrate and products. Results. There were significant differences between the two P4502D6 variants in the formation kinetics of O-demethylmetoprolol and -hydroxymetoprolol. The h2D6-Val microsomes highly favored the formation of the O-demethylmetoprolol regioisomer 6.3:1 and 2.8:1, respectively from (R)-metoprolol-d₀ and (S)-metoprolol-d₂, while the corresponding ratios for h2D6v2 microsomes were much lower. For both variants, O-demethylmetoprolol formation favored the (R)-substrate 1.5 to 2-fold, while -hydroxymetoprolol formation was non-enantioselective. Similar Km values of metoprolol oxidation, 10-20 µM, were observed for the two microsomal preparations. Conclusions. The regioselectivity, enantioselectivity, and Km values for the h2D6-Val microsomes resemble those observed for the native P4502D6 in human liver microsomes, whereas the h2D6v2 microsomes deviated remarkably in regioselectivity.     

Acute obstructive hydrocephalus due to cysticercosis during pregnancy

Background: Cysticercosis, due to the parasite Taenia solium, can involve any organ. When central nervous system infection occurs, signs and symptoms depend on the location of the cerebral lesions. Most patients develop seizures, focal symptoms, or headaches with nausea and vomiting.Case: A case of extraparenchymal (intraventricular) cysticercosis was diagnosed in a patient at term who presented with acute alteration in mental status. Ventriculostomy was performed because of acute obstructive hydrocephalus. Labor ensued and was augmented with oxytocin. Intrapartum management included magnesium sulfate seizure prophylaxis and corticosteroids. Intracranial pressures ranged between 4 and 12 cm H(2)O peripartum with approximately 300 mL of cerebrospinal fluid drained over the first 24 hours. Postpartum management included craniotomy with resection of a larval cyst and oral praziquantel therapy.Conclusion: This case describes an uncommon presentation of neurocysticercosis that should be considered in gravidas with acute mental status changes.