Studies of photokilling of bacteria using titanium dioxide nanoparticles

Abstract:  Metal pins used to apply skeletal traction or external fixation devices protruding through skin are susceptible to the increased incidence of pin site infection. In this work, we tried to establish the photokilling effects of titanium dioxide (TiO₂) nanoparticles on an orthopedic implant with an in vitro study. In these photocatalytic experiments, aqueous TiO₂ was added to the tested microorganism. The time effect of TiO₂ photoactivation was evaluated, and the loss of viability of five different bacteria suspensions ( Escherichia coli , Pseudomonas aeruginosa , Staphylococus aureus , Enterococcus hirae , and Bacteroides fragilis ) was examined by the viable count procedure. The bactericidal effect of TiO₂ nanoparticle-coated metal plates was also tested. The ultraviolet (UV) dosage used in this experiment did not affect the viability of bacteria, and all bacteria survived well in the absence of TiO₂ nanoparticles. The survival curve of microorganisms in the presence of TiO₂ nanoparticles showed that nearly complete killing was achieved after 50 min of UV illumination. The formation of bacterial colonies above the TiO₂ nanoparticle-coated metal plates also decreased significantly. In this study, we clearly demonstrated the bactericidal effects of titanium dioxide nanoparticles. In the presence of UV light, the titanium dioxide nanoparticles can be applicable to medical facilities where the potential for infection should be controlled.     

Effect of roughness of zirconia and titanium on fibroblast adhesion

Abstract:  The aim of this study was to investigate the adhesion (4 and 24 h) and the morphology of fibroblast Balb/c 3T3 seeded onto polystyrene, partially stabilized (ZrO₂Y₂O₃), stabilized zirconia ceramic (3YTZP), and pure titanium (Ti, grade 2). Initial cell adhesion (4 h) was greater (P < 0.05, analysis of variance and Tukey's Multiple Comparisons Test) onto ZrO₂Y₂O₃ and polystyrene than in Ti and 3YTZ. After 24 h, the number of adhered cells was similar between the biomaterials tested, but smaller than onto polystyrene (P < 0.05). Cells were more spread onto glass surface after 4 h, but after 24 h, the morphology and density of the cells were similar in all groups (SEM). Profilometry showed distinct Ra values for each material: glass coverslips and ZrO₂Y₂O₃ (0.09 µm), Ti (0.88 µm), and 3YTZP (30.93 µm). It was concluded that ZrO₂Y₂O₃ promoted the best initial adhesion, thus indicating that surfaces with Ra values smaller than 0.1 µm could be more favorable to initial adhesion.     

Do changes in carhiac output affect the inspiratory to end-tidal oxygen dfference?

Background. The paramagnetic technique has made it possible to monitor the end-tidal oxygen concentration and P_(I-ET)O₂, i. e. inspiratory to end-tidal oxygen difference, breath-by-breath. Little is known about the implications of a changing P_(I-ET)O₂, but so far studies have shown it to be a quick and sensitive variable to detect hypoventilation. This study was designed to observe the circulatory effects on P_(I-ET)O₂ in an experimental setting but monitored as in a clinical situation.
Methods. We assessed the oxygen difference during changes in cardiac output induced by intravenous ephedrine-hydrochloride in 12 healthy male volunteers. P_(I-ET)O₂ was measured with a fast-response paramagnetic differential oxygen sensor. Cardiac output was measured with non-invasive transthoracie electrical bioimpedance. As stimultaneous changes in metabolism and ventilation will also influence P_(I-ET)O₂, oxygen uptake and expired minute volume were monitored. After a rest period, the subjects had an intravenous injection of ephedrine-hydrochloride 0.1 mg·kg^-1 followed by a 30-min observation period.
Results. Cardiac output increased significantly as did the oxygen uptake and the ventilation. We found no biologically significant correlation between cardiac output and P_(I-ET)O₂. The P_(I-ET)O₂ was influenced by ventilation and metabolism.     

Open-Loop Analysis of Circulatory System in Awake Live Animals: Relations between Mixed Venous Saturation (SvO2) and Hemodynamic Parameters

Abstract: In order to derive quantitative relations between the mixed venous hemoglobin saturation (S_vO₂) and hemodynamic and metabolic parameters in awake live animals, biventricular bypass experiments were con-ducted in goats using a pair of pneumatic artificial hearts. The artificial hearts were used to bypass continuously the fibrillating ventricles, to control precisely the pump output, and to derive open-loop responses of the peripheral circulatory system. The specially developed reflection-type optical sensors were mounted on each artificial heart to measure continuously arterial and mixed venous SO₂ and hemoglobin content ([Hb]), which in combination with pump output data oxygen consumption of the animal was derived on-line for a maximum duration of 40 days. The S_vO₂ sensitively reflected the changes in respiratory status, [Hb], pump output, and oxygen consumption. The S_vO₂ was linearly related to the oxygen consumption but showed nonlinear relation to the pump output. The exercise study revealed that the S_vO₂ level may be used to detect the anaerobic threshold. The S_vO₂ is a useful parameter to evaluate circulatory status of the patients.     

An in vitro promoting role for hydrogen peroxide in human sperm capacitation

A complex process of maturation called capacitation is an essential step for spermatozoa to fertilize oocytes. Recent studies have shown that reactive oxygen species (ROS) can enhance the capacitation of human spermatozoa and sperm-zona interaction. We have investigated whether hydrogen peroxide (H₂O₂) could trigger capacitation of human spermatozoa and the acrosome reaction. The addition of catalase, a specific H₂0₂ scavenger, at the beginning of the capacitation process decreased the levels of both hyperactivation and induced-acrosome reaction whereas catalase added 15 min before the induction of the acrosome reaction by the calcium ionophore had no effect. Supplementation of the medium with H₂O₂ resulted in increased levels of hyperactivation and the acrosome reaction, whereas H₂O₂ added 15 min before induction of the acrosome reaction did not have any stimulatory effect. These results suggest that H₂O₂ may be involved in the capacitation process of human spermatozoa but not in the acrosome reaction.     

Ex vivo magnetofection with magnetic nanoparticles: a novel platform for nonviral tissue engineering

Abstract:  Several methods have been described to introduce DNA expression vectors into mammalian cells both in vitro and in vivo. Each system has benefits and limitations, and to date there is still no ideal method for gene transfer. In this study, we introduced a novel method of gene transfer by using Fe₃O₄ nanoparticles. The magnetic nanoparticles composed of Fe₃O₄, and the transfected genes used are Lac Z and enhanced green fluorescence protein gene ( EGFG ). Four different groups of preparations included in this study were homemade liposome-enveloped EGFP –DNA/Fe₃O₄, homemade liposome EGFP –DNA gene without magnetic Fe₃O₄ nanoparticles, lipofectamine 2000-enveloped EGFP –DNA, and EGFP –DNA gene only. Mice osteoblast and He99 lung cancer cell line were used as host cells for gene transfection. The time-dependent EGFP gene expression was monitored and analyzed. The results showed that the diameter of the complex was less than 100 nm. There was no cytotoxicity observed at any of the magnetic Fe₃O₄ nanoparticle concentrations tested. In the presence of magnetic field, the liposome-enveloped EGFP –DNA/Fe₃O₄ complex exhibited a much higher efficiency for transfecting EGFP –DNA into osteoblast cells under external magnetic fields. The gene can be transfected into cells with an aid of magnetic vectors and magnetic force. Under a gradient magnetic field, the efficiency of magnetofection is enhanced as compared to that without magnetic field.     

Does preoperative night pulse oximetry provide a reliable baseline?

Background: Postoperative hypoxaemia may be detected by pulse oximetry monitoring of the arterial haemoglobin saturation (S_pO₂). The S_pO₂-values obtained on the preoperative night are presumed to represent baseline values for the patients. The aim of this study was to evaluate the possible difference between nocturnal S_pO₂-values in the patient's home and in the hospital before operation.
Methods: We included 20 patients, 60 years or older, who underwent continuous monitoring of S_pO₂ on one night in the patient's home and on the night following hospitalisation before surgery. The alarms were turned off and the values were not accessible to the hospital staff. The S_pO₂ value was stored every 10 s. We described the general oxygen saturation level using the median of all valid values and we calculated the number of valid S_pO₂-values at 3 levels: above 90%, between 86 and 90%, and below 86%.
Results: The number of valid S_pO₂-values at home and in the hospital were 2186 and 2330, respectively; the median value was 96% on both occasions. Only 0.36% of the values were below 91% on the two occasions and there were no statistically significant differences between the number of S_pO₂-values at any of the considered levels.
Conclusion: Arterial oxygen saturation values measured on the preoperative night in the hospital seem reliable as baseline values for elderly patients.     

Enhanced expression of glutathione peroxidase protects islet beta cells from hypoxia-reoxygenation

Abstract:  The survival of pancreatic islet β –cell xenografts and allografts may be affected by damaging reactive oxygen and nitrogen species generated during hypoxia-reoxygenation. Peroxynitrite, which is formed from superoxide and nitric oxide, appears to be an important mediator of β -cell destruction. The intracellular antioxidant enzymes glutathione peroxidase-1 (Gpx-1) and copper–zinc superoxide dismutase (CuZn SOD) detoxify peroxynitrite and superoxide, respectively. The aim of this study was to examine whether enhanced expression of Gpx-1 and/or CuZn SOD protected NIT-1 mouse insulinoma cells from hypoxia–reoxygenation injury. Stable transfectants expressing human Gpx-1 or CuZn SOD were isolated and tested for their resistance to hydrogen peroxide (H₂O₂) and menadione, which generates superoxide intracellularly. Clones expressing one or both enzymes were subjected to hypoxia in glucose-free medium for 18 h, followed by reoxygenation in complete medium for 1.5 h. Cell viability was measured using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazoliumbromide (MTT) reduction assay. Increases of up to two fold in Gpx or total SOD activity protected NIT-1 cells from H₂O₂ and menadione. Expression of Gpx-1 significantly increased NIT-1 survival following hypoxia-reoxygenation (viability 65 ± 9% vs. control 15 ± 3%, P  < 0.001) but CuZn SOD expression had no effect (15 ± 1%). Expression of both enzymes was no more protective (60 ± 6%) than expression of Gpx-1 alone. Genetic manipulation of islet β cells to increase expression of Gpx-1 may protect them from oxidative injury associated with the transplantation procedure.     

Evaluation of Platelet Adhesion and Activation on Materials for an Implantable Centrifugal Blood Pump

A totally implantable centrifugal artificial heart has been developed in which a pivot bearing supported centrifugal pump is used as a blood pump. The following have been adopted as blood contacting materials in our pump: titanium alloy (Ti-6Al-4V) for the housing and impeller, alumina ceramic (Al₂O₃) for the male pivots, and ultrahigh molecular weight polyethylene (PE) for the female pivots. Greater antithrombogenicity is required for an implantable blood pump. To examine the thrombogenicity of these materials, we evaluated in vitro platelet adhesion and activation, which may play key roles in thrombogenesis on foreign surfaces. Ti-6Al-4V, Al₂O₃, and PE were compared with polycarbonate (PC), silicone carbide (SiC), and pure titanium (pTi). Platelet adhesion was assessed using monoclonal antibody (CD61) directed against glycoprotein IIIa. Platelet activation was evaluated by measuring P-selectin (GMP-140) released from irreversibly activated platelets. Each material with a surface area of 16.6 cm² was incubated with 2.5 ml of plasma or 2.5 ml of heparinized fresh whole blood for 3 h at 37°C. The optical density (OD) at a wavelength of 450 nm for CD61 was 0.93 ± 0.35 in PC, 0.34 ± 0.13 in PE, 0.27 ± 0.13 in pTi, 0.26 ± 0.01 in Al₂O₃, 0.21 ± 0.04 in SiC, and 0.12 ± 0.12 in Ti-6Al-4V. The GMP-140 levels of the tested materials were not significantly different from the control value (45.9 ± 7.2 ng/ml). These results indicate that Al₂O₃, PE, and Ti-6Al-4V, which are incorporated into our implantable centrifugal pump, have satisfactory antithrombogenic properties in terms of platelet adhesion. However, platelet activation by any material was not observed under the static condition in this study.     

Hemodialysis with Cellulose Membranes Primes the Neutrophil Oxidative Burst

Abstract: Hemodialysis with cellulose membranes causes a complement-mediated neutropenia. Changes in neutrophil function have also been reported; however, it is unclear if these changes indicate a direct effect of the membrane on neutrophils or if they are a consequence of the neutropenia. We tested the hypothesis that neutrophil oxidative burst activity is enhanced during dialysis with cellulose membranes. Resting and Staphylococcus aureus -stimulated H₂O₂ production were determined predialysis and in blood entering and leaving the dialyzer during the first 30 min of dialysis and in blood leaving the membrane module in a single-pass on-line model of hemodialysis. Resting H₂O₂ production increased slightly but significantly during the first 30 min of dialysis. Transit of neutrophils through the dialyzer caused a marked increase in stimulated H₂O₂ production, indicating priming of the oxidative burst. However, priming was limited to the first 5 min of dialysis before the onset of neutropenia. In contrast, stimulation and priming of H₂O₂ production persisted throughout 30 min of single-pass on-line perfusion. These results indicate that cellulose membranes both stimulate and prime neutrophil oxidative burst activity but that these effects are partially obscured by neutropenia.     

Liposome-Encapsulated Hemoglobin Reduces the Size of Cerebral Infarction in Rats: Effect of Oxygen Affinity

Liposome-encapsulated hemoglobin (LEH) with a low oxygen affinity (l-LEH, P ₅₀ = 45 mm Hg) was found to be protective in the rodent and primate models of ischemic stroke. This study investigated the role of LEH with a high O₂ affinity (h-LEH, P ₅₀ = 10 mm Hg) in its protective effect on brain ischemia. The extent of cerebral infarction was determined 24 h after photochemically induced thrombosis of the middle cerebral artery from the integrated area of infarction detected by triphenyltetrazolium chloride staining in rats receiving various doses of h-LEH as well as l-LEH. Both h-LEH and l-LEH significantly reduced the extent of cortical infarction. h-LEH remained protective at a lower concentration (minimal effective dose [MED]: 0.08 mL/kg) than l-LEH (MED: 2 mL/kg) in the cortex. h-LEH reduced the infarction extent in basal ganglia as well (MED: 0.4 mL/kg), whereas l-LEH provided no significant protection. h-LEH provided better protection than l-LEH. The protective effect of both high- and low-affinity LEH may suggest the importance of its small particle size (230 nm) as compared to red blood cells. The superiority of h-LEH over l-LEH supports an optimal O₂ delivery to the ischemic penumbra as the mechanism of action in protecting against brain ischemia and reperfusion.     

Low-flow anaesthesia at a fixed flow rate

Aims and Objectives: This study attempts to assess the safety of low-flow anaesthesia (LFA) at fixed flow rates with particular reference to the incidence of a decline in FiO₂ below safe levels of 0.3 and to determine whether LFA can be used safely in the absence of an FiO₂ monitor.
Methods: A total of 100 patients undergoing procedures under general anaesthesia at fresh gas flows of 300 ml/min of O₂ and 300 ml/min of N₂O were monitored while maintaining the dial setting of isoflurane at 1.5% for 2 h. The changes in gas composition were analysed and even a single recording of FiO₂ of <0.3 was considered sufficient to render the technique unsafe in the absence of gas monitors.
Results: The lowest recorded value of FiO₂ was 31% (v/v%). There was no incidence of adverse events necessitating the conversion from low flows to conventional flows.
Conclusions: We conclude that low flows of 300 ml/min of N₂O and 300 ml/min of oxygen can be used safely for a period of 2 h without the use of monitors for gas analysis of oxygen and agent in adult patients weighing between 40 and 75 kgs.     

Heat stress produces an early phase of protection against oxidative damage in human muscle

Background: It has been rarely reported that heat stress induces an early phase of protection against oxidative damage, whereas a delayed phase of protection is shown in heat stress. To explore the early effect of heat stress against oxidative damage, we evaluated the changes in contractility, lipid peroxidation, and ultrastructure induced by hydrogen peroxide (H₂O₂) with or without heat stress (HS) in human skeleton muscle.
Methods: Thirty-two muscle samples were obtained from the vastus lateralis muscle of 7 subjects. These specimens were divided into three groups based on form of treatment: HS (n=13), non HS (n=14), and control group (n=5). The control group was performed under identical conditions without H₂O₂. Specimens in the HS group were incubated at 42°C for 20 min, while those in the non-HS and control groups were maintained at 37°C.
Results: The control group showed no significant change in contractile force. Although contractile force significantly decreased 30 min after H₂O₂ administration in both the HS and non-HS groups, only the HS group showed apparent recovery of contractile force 60 min after H₂O₂ administration. Lipid peroxidation was lower in the HS group than in the non-HS group. Ultrastructural examination revealed less mitochondrial damage in the HS group compared with the non-HS group.
Conclusion: We found that human skeleton muscle escaped cellular damage induced by H₂O₂ in the early phase after heat stress. These data suggest evidence for an early effect of heat stress against ischemia/reperfusion injury in human muscle.     

Clinical assessment of oxygen delivery and consumption during hypotensive anaesthesia - A clinical application of The Deep Picture™

Usual evaluation of the relationship between oxygen delivery (DO₂) and oxygen uptake (VO₂) is based on the arterial oxygen tension (pO₂), oxygen saturation (sO₂), haemoglobin concentration (ctHb), the same indicators in mixed venous blood and cardiac output, sometimes supplemented by the expiratory carbon dioxide concentration. And, so far, the relationship among DO₂, VO₂ and the new parameters for an evaluation of oxygen status (oxygen extraction tension: p_K, concentration of extractable oxygen: c_x, oxygen compensation factor: Q_x) (1) has not been discussed enough.
Therefore, this study was designed to evaluate whether the new parameters give the clinically significant information to analyse the relationship between DO₂ and VO₂ during the acute haemodynamic change with intentionally induced hypotension in anaesthetized adult patients.     

Extracorporeal shock wave treatment modulates skin fibroblast recruitment and leukocyte infiltration for enhancing extended skin-flap survival

Extracorporeal shock wave (ESW) treatment has a positive effect of rescuing ischemic skin flaps. This study assessed whether ESW treatment rescues the compromised flap tissue by suppressing the apoptosis of ischemic tissue and recruiting tissue remodeling. We used a random-pattern extended dorsal–skin-flap (10 × 3 cm) rodent model. Thirty-six male Sprague–Dawley rats were divided into three groups. Group I, the control group, received no treatment. Group II received one session of ESW treatment (500 impulses at 0.15 mJ/mm²) immediately after surgery. Group III received two sessions of ESW treatment, immediately and the day after the surgery. Results indicated that the necrotic area in the flaps in group II was significantly smaller than that of the flaps in group I ( p <0.01). Transferase dUTP-nick end labeling (TUNEL) analysis revealed a significant decrease in the number of apoptotic cells in group II. Hydrogen peroxide (H₂O₂) expression in circulation blood was significantly decreased in group II on the day after ESW treatment. Immunohistochemical staining indicated that compared with no treatment, ESW treatment could substantially increase proliferating cell nuclear antigen (PCNA), endothelial nitric oxide synthase, and prolyl 4-hydroxylase (rPH) expression, reduce CD45 expression, and suppress 8-hydroxyguanosine (8-OG) expression in the ischemic zone of the flap tissue. In conclusion, ESW treatment administered at an optimal dosage exerts a positive effect of rescuing ischemic extended skin flaps. The mechanisms of action of ESWs involve modulation of oxygen radicals, attenuation of leukocyte infiltration, decrease in tissue apoptosis, and recruitment of skin fibroblasts, which results in increased flap tissue survival.     

Central venous oxygen saturation for the diagnosis of low cardiac output in septic shock patients

Background: Simple diagnostic tests are needed to screen septic patients for low cardiac output because intervention is recommended in these patients. We assessed the diagnostic value of central venous oxygen saturation in the superior vena cava (ScvO₂) for detecting low cardiac output in patients with septic shock.
Methods: We conducted a prospective observational study in three general intensive care units (ICUs) of adult patients with septic shock, who were to have a catheter inserted for thermodilution measurement of cardiac index (CI_TD). Paired measurements of CI_TD and central venous oximetry values were obtained when the clinician first measured CI_TD.
Results: We included 56 patients with septic shock and a mean sequential organ failure assessment score of 12 (range 3–20). Baseline CI_TD was 3.5 l/min/m² (1.0–6.2) and ScvO₂ of 70% (33–87). The best cut-off of ScvO₂ for CI_TD>2.5 l/min/m² ( n =42) was a value ≥64% with positive and negative predictive values of 91% (95% confidence interval 79–98) and 91% (59–100), respectively. The diagnostic values were not improved by using instead central venous O₂ tension or the difference between arterial and central venous O₂ saturation.
Conclusions: This prospective, observational study found that a ScvO₂ measurement of ≥64% indicated CI_TD>2.5 l/min/m² in ICU patients with septic shock.     

Thoughts and Progress: Electromotive Administration of a New Morphine Formulation: Morphine Citrate

Abstract: Two formulations of morphine citrate were synthesized: trimorphine citrate, 3(MH)+(C₆H₅O₇)^3– and morphine sodium citrate, 3(MH)+ 3Na+2(C₆H₅O₇)^3–. Four healthy individuals volunteered to undergo electromotive administration of the two formulations. Application of electric current (2 mA) to solutions of trimorphine citrate for 1 h resulted in iontophoretic transcutaneous administration of therapeutic quantities of morphine, without deleterious reduction in the pH of the drug solutions. Application of a 2–mA current to solutions of morphine sodium citrate for 2 h resulted in combined iontophoretic and electrophoretic delivery of morphine with increased administration rates and an improved buffering capacity of the drug solutions.     

Dermal Heat Transport Analysis for Transcutaneous O2 Measurement

Heat from a transcutaneous oxygen electrode is transmitted locally to the blood beneath it causing a shift in the HbO₂ dissociation curve. This increases the local PO₂, and allows a measurable PO₂, at the skin surface. The temperature effect on the HbO₂, curve must be accounted for in in vivo calibration of Ptco₂, data. To do this, the capillary blood temperature beneath the electrode must be known. A heat balance is written around the capillary blood with heat being conducted in from the electrode and carried out by two means: conduction to deep tissue; and transport away by the flowing capillary blood. The following equation is the steady state solution of the heat transport problem:
T₈ = ±
where Z = ± = 0.17
T₈, = capillary blood temperature
T₁ = electrode temperature
T_o = body temperature
ρ = blood density
P = cutaneous perfusion
δ = dermal capillary depth
k = thermal conductivity of skin
C± = heat capacity of blood
This solution shows the capillary blood temperature may be calculated if the T₁ and T_o are measured and the physiologic constants in 2 are known. 2 is a dimensionless heat transport number which represents the relative importance of perfusion to conduction effects on the deterring T₈, and may be used as a data correlating parameter. Z = 0.17 is obtained using literature values for the physiological constants. This analysis used in conjunction with a mass transport analysis for oxygen will produce a theoretically based correlation scheme for in vivo calibration of heated transcutaneous oxygen electrodes.     

Changes in delivered tidal volume with the addition of carbon dioxide to mechanical ventilation

Objective:  To determine discrepancies in delivered tidal volumes induced by the addition of carbon dioxide (CO₂) during mechanical ventilation.
Design:  Prospective, experimental, lung model study.
Setting:  Research laboratory at a University hospital.
Subjects:  Ventilator set-up using a mechanical lung model.
Interventions:  Tidal volumes were measured during pressure and volume limited ventilation at various pressures and volumes with the addition of inspired CO₂ concentrations of 0, 1, 2, and 4%. Three ventilator set-ups were used including the addition of CO₂ to the inspiratory limb (M1), the use of two external blenders to mix air, oxygen and CO₂ prior to entry to the ventilator (M2), and the use of one external blender to mix air and CO₂ prior to addition to the ventilator.
Measurements and main results:  Statistically significant increases in delivered tidal volume were noted with the addition of CO₂ with all three of the ventilator set-ups (M1, M2 and M3). However, the maximum increase was 2.4% above that of baseline (no CO₂ added). With the M2 set-up, there was also a significant discrepancy noted between the set and the delivered tidal volumes (16–17%) when using the volume limited mode even without the addition of CO₂.
Conclusions:  Either the M1 or the M3 set-up functioned efficiently without clinically significant alterations in ventilator performance. We prefer the M1 set-up as it is the one that is used most commonly in clinical practice and does not require significant alterations in the working configuration of the ventilator.