Microsecond-pulsed dielectric barrier discharge plasma stimulation of tissue macrophages for treatment of peripheral vascular disease.

Angiogenesis is the formation of new blood vessels from pre-existing vessels and normally occurs during the process of inflammatory reactions, wound healing, tissue repair, and restoration of blood flow after injury or insult. Stimulation of angiogenesis is a promising and an important step in the treatment of peripheral artery disease. Reactive oxygen species have been shown to be involved in stimulation of this process. For this reason, we have developed and validated a non-equilibrium atmospheric temperature and pressure short-pulsed dielectric barrier discharge plasma system, which can non-destructively generate reactive oxygen species and other active species at the surface of the tissue being treated. We show that this plasma treatment stimulates the production of vascular endothelial growth factor, matrix metalloproteinase-9, and CXCL 1 that in turn induces angiogenesis in mouse aortic rings in vitro. This effect may be mediated by the direct effect of plasma generated reactive oxygen species on tissue.

Quantitative assessment of cell adhesion molecule gene expression in endomyocardial biopsy specimens from cardiac transplant recipients using competitive polymerase chain reaction.

BACKGROUND: Adhesion of leukocytes to vascular endothelium is an early step in cardiac allograft rejection leading to migration of lymphocytes into parenchymal tissues. Cell adhesion molecule (CAM) protein expression appears to increase as a result of rejection. The relationship of CAM gene expression to rejection is less well defined. The goal of this study was to define cell adhesion molecule gene expression in relation to the presence of acute cellular rejection in endomyocardial biopsies from cardiac transplant recipients. METHODS: To quantitatively assess intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1) and E-selectin gene expression, we developed a competitive PCR system using nonhomologous DNA fragments (MIMICs) with complementary sequences to CAM gene-specific primers as internal standards. MIMIC fragments with known concentrations were mixed in serial dilutions with constant amounts of cDNA from the biopsy specimens and amplified with common primers under the same polymerase chain reaction conditions. The relative CAM cDNA concentrations were determined by comparing the density of MIMIC to target cDNA bands on agarose gel. ICAM-1, VCAM-1, and E-selectin mRNA concentrations were analyzed from 38 cardiac transplant biopsies divided into 3 groups according to ISHLT rejection grade: group 1-grade 0 (n=13); group 2-grade 1A or 1B (n=13); group 3-grade 3A (n=12). Glyceraldehyde-3-phosphate dehydrogenase (a constitutive gene) was quantified in the same way as CAMs to normalize the relative levels of CAMs. RESULTS: The results expressed as mean (1x10(-3) pM) (+/-SEM) in groups 1, 2, and 3, respectively, were: ICAM-1; 5+/-1; 57+/-4*; 64+/-13*, VCAM-1; 0.8+/-0.1; 6+/-1**; 9+/-1*, E-selectin; 0.4+/-0.2; 0.8+/-0.2; 0.4+/-0.1 (*P<0.001 versus group 1; **P<0.01 versus group 1). CONCLUSIONS: ICAM-1 and VCAM-1 gene expression was increased during rejection in endomyocardial biopsy specimens. Competitive polymerase chain reaction can be used to quantitatively assess gene expression in biopsy specimens from patients.

The anticoagulant properties of a modified form of protein S.

Protein S (PS) is a vitamin K-dependent anticoagulant that acts as a cofactor to activated protein C (APC). To date PS has not been shown to possess anticoagulant activity in the absence of APC. In this study, we have developed monoclonal antibody to protein S and used to purify the protein to homogeneity from plasma. Affinity purified protein S (PSM), although identical to the conventionally purified protein as judged by SDS-PAGE, had significant anticoagulant activity in the absence of APC when measured in a factor Xa recalcification time. Using SDS-PAGE we have demonstrated that prothrombin cleavage by factor Xa was inhibited in the presence of PSM. Kinetic analysis of the reaction revealed that PSM competitively inhibited factor Xa mediated cleavage of prothrombin. PS preincubated with the monoclonal antibody, acquired similar anticoagulant properties. These results suggest that the interaction of the monoclonal antibody with PS results in an alteration in the protein exposing sites that mediate the observed anticoagulant effect. Support that the protein was altered was derived from the observation that PSM was eight fold more sensitive to cleavage by thrombin and human neutrophil elastase than conventionally purified protein S. These observations suggest that PS can be modified in vitro to a protein with APC-independent anticoagulant activity and raise the possibility that a similar alteration could occur in vivo through the binding protein S to a cellular or plasma protein.

The choice of jet nebulizer, nebulizing flow, and addition of albuterol affects the output of tobramycin aerosols.

The use of inhaled antibiotics in the treatment of cystic fibrosis has become widespread despite controversy in the literature as to the appropriate dosing regimen and its effectiveness. This study compared two tobramycin (T) preparations (one with and one without the addition of albuterol) using two different jet nebulizers in order to determine if drug output would be affected. Using calibrated flows from a dry compressed gas source of 6 and 8 L/min as well as a specific compressor (Pulmo-Aide), the Hudson 1720 nebulizer was compared with the newer disposable Hudson 1730. The albuterol preparation used in this study was the Ventolin (albuterol) Respirator Solution (VRS). The nebulizers were charged with (1) 2 mL T (80 mg/2 mL) with 0.5 mL VRS (5 mg/mL) and normal saline solution to make the total nebulizer charge of 3 or 4 mL, or (2) 2 mL T and either 1 or 2 mL normal saline solution. A laser diffraction analyzer (Malvern 2600) was used to determine the aerosol particle size distribution. From the distribution, the respirable fraction, which is the fraction of aerosol that could enter and remain in the lungs, was calculated. For all solutions and each particular flow, the Hudson 1730 had a larger respirable fraction of T. The addition of VRS lowered the surface tension of the solution in the nebulizer and resulted in a greater output of T. This effect was most apparent for the 3-mL volume fills of the Hudson 1720. The greatest differences were between the 3-mL nebulizer charges of T using the Hudson 1720 driven by a flow of 6 L/min, which produced 8 mg of T in the respirable fraction, compared with 35 mg produced by the Hudson 1730 driven by a flow of 8 L/min. These results suggest that different nebulizers, different nebulizer solutions, and different techniques of nebulization may result in very different amounts of T aerosol output in the respirable fraction.

A comparison of pulmonary availability between Ventolin (albuterol) nebules and Ventolin (albuterol) Respirator Solution.

The two most common albuterol preparations used for nebulization are: (1) Ventolin (albuterol) respirator solution (Glaxo Canada Inc; Montreal, Canada) of which 2.5 mg (0.5 mL) is diluted with 2 mL of normal saline solution, and (2) the preservative-free, prediluted Ventolin (albuterol) Nebules PF (Glaxo) (2.5 mg/2.5 mL). The two preparations were compared using both a Hudson 1720 "T" up-draft Neb-U-Mist jet nebulizer and a Hudson 1730 "T" up-draft Neb-U-Mist II jet nebulizer (Hudson; Temecula, Calif), which were driven by a compressor (Pulmo-Aide; Devilbiss; Somerset, Pa) and by dry compressed air at 6 and 8 L/min. Particle size distribution was measured with a particle sizer (Malvern 2600; Malvern Instruments; Malvern, UK) and drug output for the nebulizer was calculated from the differences in predrug and postdrug volume and concentration. Drug availability was defined as the amount of drug carried in particles less than 5 microns in diameter. Drug availability was greater with the albuterol respiratory solution, due to the surface activity of the preservative benzalkonium chloride, for both nebulizers but particularly for the 1720. Differences in drug availability between nebulizers exceeded fourfold depending on the preparation, the nebulizer, and the nebulizing flow. These differences could not have been predicted from the manufacturer's specifications. The results suggest that prediction of drug availability must be based on measurements with the specific preparation and the specific nebulizer used.

A comparison of the availability of tobramycin for inhalation from vented vs unvented nebulizers.

STUDY OBJECTIVE: To compare drug output from a vented nebulizer (Pari LC Jet Plus) with a traditional unvented nebulizer (Hudson 1730 T Up-Draft 11) using aerosolized tobramycin, which is frequently used in the treatment of cystic fibrosis. DESIGN: Six nebulizers of each type were filled with a 4 mL tobramycin (80 mg) solution and were driven by a compressor (Pulmo-Aide). Various inspiratory flows (VI) (0, 5, 10, 15, 20 L/min for the Pari LC Jet Plus and 0, 5, and 10 L/min for the Hudson 1730, all at 40% relative humidity) were directed through each nebulizer. Drug output was measured from changes in weight and concentration (assessed by changes in osmometry) within the nebulizer. Particle size distributions were determined by laser diffraction allowing the calculation of the amount of aerosol output in the respirable range (<5 microm). The nebulizers were first run until end-nebulization to establish total drug output and then for either 4 or 5 min to determine the rate of drug output (mg/min) before intermittent aerosol output. RESULTS: The total drug output without VI for both the unvented and the vented nebulizers was not significantly different, 55 (51, 60) mg for the Hudson 1730 vs 51 (49, 53) mg for the Pari LC Jet Plus (mean [95% confidence limits]). Inspiratory flow had no effect on the unvented Hudson 1730 nebulizer but significantly increased the rate of total drug output and the rate of drug output in the respirable range for the vented Pari LC Jet Plus nebulizer (VI=0, 3.35 [2.84, 3.85] and 1.72 [1.48, 1.96] compared with VI=20, 9.87 [9.03, 10.70] and 6.11 [5.33, 6.88] mg/min). CONCLUSIONS: These findings indicate that the increase in the rate of drug output with VI for the vented nebulizer would result in shorter nebulization times and a relative decrease in drug loss during the expiratory phase.

Inhibitory effect of anoxia on 125I-insulin binding by rat hepatocytes.

We have examined the possibility that 125I-insulin binding by isolated rat hepatocytes is modulated by cellular ATP levels. To avoid complications due to ATP-dependent internalization of bound insulin, 125I-insulin binding was determined at 10 degrees C; at this temperature, equilibrium binding was achieved after incubation for 4-6 h. When hepatocytes were incubated at 37 degrees C under anaerobic conditions, ATP levels and 125I-insulin binding were both lowered by about 65%. Anoxia inhibited the association of 125I-insulin with the hepatocyte receptor; the dissociation of insulin from hepatocytes was not affected. Cellular ATP levels and 125I-insulin binding were both restored when anaerobic cells were incubated further at 37 degrees C under aerobic conditions. When anaerobic cells were incubated in air at 10 degrees C during the insulin binding assay, 125I-insulin binding recovered completely, but ATP levels were unaffected. The inhibitory effect of anoxia on 125I-insulin binding was not due to any effect on 125I-insulin degradation or on cell viability. We conclude (1) that the ability of hepatocytes to bind insulin can be modulated on a short-term basis in response to the metabolic status of the cell, and (2) that modulation of the liver cell insulin receptor is not a function of cellular ATP levels.

Changes in cardiac contractility in IDDM and NIDDM diabetic rats.

Differences in myocardial contractility were studied in type I (insulin-dependent, IDDM) and type II (non-insulin-dependent, NIDDM) diabetic rats. Using the streptozotocin-induced diabetes as an experimental model, the contractile properties of left ventricular myocardium of IDDM and NIDDM animals were compared to similar parameters of their age-matched controls. Contraction force was analyzed as a function of the pacing frequency. Paired-pulse stimulation and catecholamine treatment were applied to compare the inotropic responses obtained in the two types of diabetes. Diabetic and control preparations developed equal peak tension at each driving frequency upon the application of paired-pulse stimulation with fixed interpulse interval. The interpulse interval dependence of paired-pulse induced inotropy was altered and the velocity of contraction and relaxation decreased in IDDM, but not in NIDDM muscles. Sensitivity to isoproterenol and norepinephrine was decreased in both types of diabetes, however, the isoproterenol resistance of old diabetic animals was attributable to age rather than to the diabetic state. The results indicate that alterations in the contractile parameters and catecholamine sensitivity in IDDM differ from those observed in NIDDM form of diabetes mellitus.

[Hemodynamic effects of verapamil in pulmonary hypertension caused by heart valve disease]. / Verapamil haemodinamikai hatásainak vizsgálata billentyübetegségek kapcsán kialakult pulmonalis hypertoniában.

The authors have investigated the haemodynamic effects of verapamil on the pulmonary circulation by 24 patients suffering from secondary pulmonary hypertension, caused by mitral and/or aortic valve diseases. For this purpose the numeric and graphometric analysis of intracavitary right ventricle pressure curve and pulmonary artery pressure tracing was applied. It was observed a selective antihypertensive effect on the lesser circulation. The pulmonary and systemic systolic tension decreased comparing in percentage 3:1, in case of diastolic tension this comparison was 2:1. The decrease in pulmonary circulation was strongly significant. The diminishing of heart rate and the improving of right heart function was not significant. The elevation of end-diastolic pressure of the right ventricle, just as the shape-analysis of pressure curves suggested right ventricle overload.

Tussiphonographic analysis of cough sound recordings performed by Schmidt-Voigt and Hirschberg and Szende.

The cough sound records published by Schmidt-Voigt and Hirschberg and Szende were submitted to tussiphonographic analysis. It has been established that all the recordings of various types of cough sounds registered in airway disease were of pathological character in the tussiphonographic recordings. It has repeatedly been confirmed that tussiphonography is a suitable means for screening of respiratory diseases.

Information obtained from tussigrams and the possibilities of their application in medical practice.

The physiology of cough was studied using 2008 voluntary coughs. The tussiexpirograms verify that in a cough-series there are "double-flow" phases. The tussitachogram enables a more precise observation, for example, one can recognise five flow phases within one single cough. It proves, furthermore, that in the first five coughs the speed of airflow is always higher in the intermediate deceleration than in the acceleration phase. On the tussiphonogram one can differentiate seven phonetic phases. The simultaneously recorded tussigrams and tussiphonograms allow identification of two sounds, two intervals and noises. There are at least two possibilities for use of these data in clinical practice, 1) the measurement of the volume during the first second on the tussiexpirogram is a suitable control for the forced expiratory volume in one second (FEV1), 2) analysis of the voluntary prolongation of the intermediate deceleration during respiratory physiotherapy.

Graphometric analysis of cough tachogram.

The tachogram shows that the flow of cough is neither continuous nor steady. Within a cough, 5 different phases can be differentiated: 1) acceleration, 2) initial fast decleration, 3) intermediary decleration, 4) terminal fast decleration, and 5) the flow-pause. Only in the course of phases 1) and 3) is the air flowing out from the lungs. In cough series the peak-flows of subsequent coughs are growing smaller and smaller. This decrease follows the form of an isosceles hyperbola. A cough can be short or elongated. From the flow-phases the 1-st, 2-nd and 4-th ones are of fairly stable duration. The wide deviation of the intermediary deceleration shows that this phase makes it possible to elongate voluntarily the cough.

Chemistry of acetylene on platinum (111) and (100) surfaces.

An ultra-high vacuum experimental study of acetylene chemisorption on Pt(111) and Pt(100) and of the reaction of hydrogen with the acetylene adsorbate has established distinguishing features of carbon-hydrogen bond breaking and making processes as a function of pressure, temperature, and surface crystallography. The rates for both processes are substantially higher on the Pt(100) surface. Net acetylene-hydrogen processes, in the temperature range of 20 degrees C to approximately 130 degrees C, are distinctly different on the two surfaces: on Pt(100) the net reaction is hydrogen exchange ((1)H-(2)H exchange) and on Pt(111) the only detectable reaction is hydrogenation. Stereochemical differences in the acetylene adsorbate structure are considered to be a contributing factor to the differences in acetylene chemistry on these two surfaces.

Preclinical trial of the antitumoral therapeutic effectiveness of some natural polyphenolic biopreparations.

We have assessed the antitumoral action of the POLYAS I and POLYAS II vegetal polyphenolic biopreparations--separated and purified from Asclepias syriaca leaves - in rats with various experimental tumoral lines. We studied the therapeutic effect of different doses on the tumor generation process and compared it with the experimental oncostatic action of several standard chemotherapeutic drugs of clinical use (thiotepa, methotrexate, melphalan and cyclophosphamide). In our experimental treatment with the bioactive polyphenolic agents, we have used various doses, both higher and lower than the dose that had conditioned the expression of their antitumoral action upon Guerin T-8 lymphotropic epithelioma and upon Walker 256 carcinosarcoma. We found the antineoplastic effectiveness of those aromatic biopreparations from phytomass to be dose-dependent. We compared the evaluation indices of the antitumoral pharmacodynamic effect we obtained in the treatment with the POLYAS biopreparations with those of reference cytostatic agents. The antitumoral potential of the new natural biopreparations is higher than, equal or close to that of the standard oncochemotherapeutic agents. Antitumoral effectiveness can be improved by an experimental manipulation of the therapeutic doses--which proves the existence of a dose-response relationship. POLYAS I and POLYAS II polyphenolic biopreparations are compatible in point of effectiveness with the standard cytostatic agents, a fact that we considered relevant for the characterization of the POLYAS I and POLYAS II vegetal extracts as potential antineoplastic agents. The quantitative preclinical evaluation of the specific pharmacodynamic effect will be complemented by the investigation of the new polyphenolic biopreparations therapeutic effectiveness in tumors with various degrees of development.