Search

A wound biofilm model was created by adapting a superficial infection model. Partial-thickness murine wounds were inoculated with methicillin-resistant Staphylococcus aureus (MRSA). Dense biofilm communities developed at the wound surface after 24 h as demonstrated by microscopy and quantitative microbiology. Common topical antimicrobial agents had reduced efficacy when treatment was initiated 24 h after inoculation compared to 4 h after inoculation. This model provides a rapid in vivo test for new agents to treat wound biofilm infections.

Data supporting the concept that microbial biofilms are a major cause of non-healing ulcers remain limited. A porcine model was established where delayed healing resulted from methicillin-resistant Staphylococcus aureus (MRSA) infection in full-thickness wounds. At the end of one study a wound remaining open was sampled and a MRSA strain was isolated. This pig-passaged strain was used as the inoculating strain in several subsequent studies. The resulting MRSA wound infections exhibited a greater, more stable tissue bioburden than seen in studies using the parent strain. Furthermore, wounds infected with the passaged strain experienced a greater delay in healing. To understand whether these changes corresponded to an increased biofilm character of the wound infection, wound biopsy samples from studies using either the parent or passaged MRSA strains were examined microscopically. Evidence of biofilm was observed for both strains, as most samples at a minimum had multiple isolated, dense microcolonies of bacteria. However, the passaged MRSA resulted in bacterial colonies of greater frequency and size that occurred more often in concatenated fashion to generate extended sections of biofilm. These results provide a model case in which increasing biofilm character of a wound infection corresponded with a greater delay in wound healing., (© 2012 by the Wound Healing Society.)

Background: Health care handwashes/sanitizers help prevent the spread of infection. Many are alcohol-based, providing immediate microbial kill. Few contain persistence factors for residual antimicrobial effects. We conducted multiple studies on Viacydin-Containing Alcohol Sanitizer (VCAS) to evaluate antimicrobial properties and skin friendliness., Methods: The potential of VCAS to cause use related skin problems was examined by repeated applications over a 3-week period. Excessive handwashing compared effects of VCAS on the skin barrier with that of other handwash products. Antimicrobial range, immediate kill rates, and resistance development were assessed as was the potential for continued antimicrobial activity over an extended period following product use., Results: Our data showed the VCAS formula has broad, rapid antimicrobial efficacy without promoting microbial resistance. VCAS is mild to skin. Antimicrobial persistence testing showed that VCAS remained effective up to 6 hours postapplication., Conclusion: VCAS was superior to or at parity with on-market products, exhibited substantial residual effects and persistence up to 6 hours, and was safe and well tolerated. These results provide strong evidence for the value of VCAS to help prevent and eliminate pathogen contamination of the hands., (Copyright © 2011 Association for Professionals in Infection Control and Epidemiology, Inc. Published by Mosby, Inc. All rights reserved.)

Purpose: Anastomotic vessels in exudative age-related macular degeneration (AMD) represent a serious clinical feature that reportedly does not respond well to either photocoagulation or photodynamic therapy (PDT). Anastomoses also occur in various animal models of choroidal neovascularization (CNV). In the present study, anastomotic vessels and their patency were evaluated in two primate CNV laser-trauma models after PDT, by using two novel photosensitizers., Methods: In cynomolgus (Macaca fascicularis) and squirrel (Saimiri sciureus) monkey eyes (n = 20), matrix placement of laser photocoagulation sites elicited CNV as a component of the development of fibrovascular tissue (FVT). FVT sites received PDT according to specific drug infusion and laser light treatment parameters. FVTs and anastomoses were evaluated by fundus photography, fluorescein angiography, and histologic examination., Results: Anastomoses averaged approximately 48% of FVT sites, with greatest occurrence in the macaque. Although PDT with each photosensitizer effectively produced FVT closure, both retinal vessels and anastomoses remained patent., Conclusions: Although PDT is effective in closing the choroidal neovascularization in FVT, this technique was ineffective in occluding anastomotic vessels and their associated tributaries within the mid- to proximal retina. Various factors (vascular diameter, composition, blood flow, orientation) may contribute to continued anastomotic patency. By convention, such vessels would typically be defined as chorioretinal anastomoses (CRAs); however, continuing studies suggest the possibility that these neovessels constitute dual-origin hybrids. Regardless of origin, viable anastomoses provide one potential mechanism for revascularization to occur after PDT and may help to explain why CRAs are considered a poor prognostic sign in patients with AMD.

Purpose: To evaluate and characterize the New-World squirrel monkey as a primate model for experimental choroidal neovascularization (CNV) studies and to compare it with the current Old-World macaque monkey model., Methods: Fibrovascular tissues (FVT) were elicited in 12 maculae of seven squirrel monkeys by laser photocoagulation using optimized laser parameters (532 nm, 0.05 second, 75 micro m, 650 mW). Follow-up fundus and fluorescein angiography (FA) examinations were conducted on postlaser days 30 and 35, followed by euthanasia and histologic analysis of tissues. For comparative evaluations, FVT development also was induced and analyzed in eight maculae of four macaque monkeys with laser parameters previously used in this species (514 nm, 0.1 second, 50 micro m, 390 and 455 mW)., Results: FVT developed in both primate species, consisting of fibrous tissue that contained vessels that ranged from sparse but identifiable capillaries to well-established neovascular networks. Overall, 65% of the photocoagulation sites in the squirrel monkey and 37% of sites in macaque monkey elicited development of FVT. Localized FVT ranged from modest to extensive thickenings of the choriocapillaris layer. Unexpectedly, 76% of the FVT sites in squirrel monkey eyes and 27% of the sites in macaque eyes showed diffuse FVT that expanded beyond the original photocoagulation sites, accompanied by neovascular infiltration of the retina., Conclusions: Like the macaque, the squirrel monkey can be considered a useful primate model for experimental CNV investigations, while additionally offering certain species-specific advantages. Diffuse FVT permit studies of antiangiogenic therapies in areas distant from laser photocoagulative trauma sites.

Rimexolone is a potent anti-inflammatory corticosteroid with a lower potential for elevating intraocular pressure, relative to other ophthalmic steroids, and is indicated for postsurgical inflammation and uveitis. Fertility and peri/postnatal toxicities were evaluated at oral gavage doses of 50, 150 or 500 mg/kg, and developmental toxicity at 100, 500, or 1000 mg/kg. In the fertility study, male rats were treated daily beginning 4 weeks prior to mating and females were treated daily beginning 2 weeks prior to mating, and through gestation day 6. Females were necropsied on gestation day 15 and males were necropsied after 10 weeks of exposure. In males, dose-related reductions in mean body weights, body weight gains, and food consumption occurred in all groups. In the 500 mg/kg females, mean body weights were reduced during gestation, and there was an increase in early resorptions and concomitant decrease in viable fetuses at this level. There were no effects on copulation or fertility indices, or on the number of corpora lutea and implantation sites. The no-observed-effect level (NOEL) for fertility and reproductive effects was 150 mg/kg. In the developmental toxicity study, female rats were treated daily from gestation days 6 through 17, necropsied on gestation day 20 and fetuses were evaluated. Maternal toxicity occurred at 500 and 1000 mg/kg as indicated by reduced maternal body weights and body weight gains. However, there was no indication of a developmental effect on fetuses due to rimexolone. The NOEL was 1000 mg/kg for the developing fetuses. In the peri/postnatal toxicity study, female rats were treated daily from gestation day 6 through lactation day 20 and necropsied. F1 developmental and behavioral parameters were evaluated. Selected F1 animals were mated at 12 weeks, allowed to deliver, and necropsied on lactation day 21. At 500 mg/kg, F0 maternal body weights were reduced during gestation and lactation, and F1 pup weights were reduced during lactation and the growth phase. There were no effects on the F1 fertility or reproductive capabilities, or on F2 developmental parameters. The NOEL for the F0 females and F1 offspring was 150 mg/kg. Together, these studies indicate that, unlike some corticosteroids, rimexolone does not produce developmental or reproductive toxicity in rats.

To investigate the relationship between the effects of lidocaine on excitation patterns and its effects on the incidence of arrhythmias, the left anterior descending coronary artery was occluded for 6-min periods separated by 30 min of reperfusion, under control conditions and after injection of lidocaine, at a dose of either 2.5, 5.0, or 10.0 mg/kg i.v., in 29 open-chest anesthetized pigs. Sixty-three unipolar electrograms and a surface lead electrocardiogram were continuously recorded during atrial pacing and spontaneous ventricular arrhythmias. Ventricular fibrillation (VF) occurred only in four of a total of 45 control occlusions. VF occurred in two of five pigs following injection of lidocaine 2.5 mg/kg, in 15 or 17 pigs following injection of a 5 mg/kg dose, and in all three preparations following injection of a 10 mg/kg dose. Just prior to VF during occlusions preceded by injections of lidocaine 5 mg/kg, activation time of ischemic myocardium in atrial-paced beats was delayed by only 30 +/- 17 ms beyond preocclusion values, compared with 18 +/- 11 ms at a similar time during control occlusions and 33 +/- 18 ms at the end of control occlusions (mean +/- SD; n = 8). As ventricular tachycardia (VT) developed in the presence of lidocaine, conduction was further slowed or blocked in ischemic areas, and slowed in nonischemic regions; at the transition from VT to VF, excitation patterns displayed circus movement involving nonischemic regions.

The sulfation of cellular chondroitin sulfate in human promyelocytic leukemia HL60 cells was inhibited by a number of phorbol diesters, which concurrently induced differentiation into monocytic cells. Inhibition was dependent on concentration, and was 90% complete at 10 nM 12-0-tetradecanoylphorbol-13-acetate (TPA), the most active ester. Maximal effects were seen within 2-4 hours following initiation of treatment. The degree of inhibition observed correlated well with the ability of the esters to induce differentiation, and with their reported affinity for a "receptor", identified as protein kinase C associated with certain lipids. Chondroitin sulfation was also inhibited in cells treated with sn-1,2-dioctanoylglycerol, a lipid which is considered to be an endogenous activator of protein kinase C. Our findings therefore indicate that monocytic differentiation of HL60 cells occurs subsequent to reduced glycosaminoglycan sulfation via activation of the calcium-activated, phospholipid-dependent protein kinase.

Transmural conduction time, measured as the difference in arrival time of impulses from a distant stimulating site at endocardial and epicardial electrodes near the left ventricular apex, has been reported to decrease when closely coupled extrasystoles are interpolated, indicating that muscle conduction could be supernormal. We have now determined that reduction in transmural conduction time is accounted for completely by the relatively late arrival time of the extrasystolic wave front at the endocardial recording site. The endocardial recording site was activated later than an immediately adjacent site within the wall in two out of eight animals, which could be interpreted as retrograde conduction. No evidence for supernormal conduction within the ventricular wall could be obtained by multiple electrode recordings. Supernormal conduction throughout the myocardial wall could not be demonstrated on stimulation of the endocardial site. We conclude that supernormal conduction in myocardial muscle cannot be demonstrated and that changes in transmural conduction time do not always measure myocardial conduction velocity.

To investigate the actions of lidocaine and diltiazem on the ischemic alterations associated with the onset of acute ischemic arrhythmias, the left anterior descending coronary artery was occluded for 6-min periods separated by 30 min of reperfusion, under control conditions and after injection of lidocaine (2.4-3.8 micrograms/mL of plasma) or diltiazem (390-510 ng/mL) in open-chest anesthetized pigs. Sixty-one unipolar electrograms were continuously recorded in the ischemic zone. Isochronal maps and isopotential maps were determined by computer analysis. The magnitude of beat-to-beat alternation of unipolar waveforms was described by the difference between the time integrals subtended by electrograms of consecutive beats. Activation times were prolonged by ischemia and the ST segment became elevated. Delay and ST elevation developed at a faster rate in the presence of lidocaine than under control conditions, but were reduced by diltiazem. ST-T alternation was not significantly different between control and lidocaine occlusions, but the incidence of negative T waves and that of ventricular tachycardia degenerating to fibrillation were higher in lidocaine occlusions than in control occlusions. In contrast, unipolar waveform alternation and negative T waves were virtually abolished by diltiazem, even at fast pacing rates (180-210 beats/min) at which diltiazem did not reduce ST elevation. Ventricular arrhythmias also were abolished by diltiazem. Thus, lidocaine and diltiazem produce opposite effects on the ischemic alterations most closely associated with the initiating mechanism of tachycardia. This could be related to differences between these drugs with regard to their actions on transmembrane currents during repolarization.

To investigate ventricular tachycardias produced in healthy canine myocardium by stimulation of sympathetic ganglia or cardiac nerves, we simultaneously recorded a surface ECG and 63 ventricular electrograms in anesthetized open-chest dogs. Isochronal and isopotential maps were generated off-line by computer. Ventricular tachycardia with uniform beat-to-beat morphology was induced in 13 or 22 dogs by electrical stimulation of the left stellate ganglion (five experiments), the left middle cervical ganglion (four experiments), the left caudal pole cardiopulmonary nerve (two experiments), or the ventrolateral cardiac nerve (eight experiments). It was not inducible by stimulation of the right-sided major cardiopulmonary nerves or ganglia. In most instances the earliest measured electrical excitation occurred on the posterior aspect of the ventricles. Isochronal maps demonstrated a radial spread of the impulse away from the area of earliest excitation. Changes in the region of earliest excitation and (or) activation pattern were accompanied by changes in QRS morphology. The potential gradients measured between areas displaying positive and negative T waves on the anterior and left lateral aspects of the ventricles were significantly increased by ventrolateral cardiac nerve stimulation. However, the ventricular regions where these potential gradients existed differed from the regions of earliest excitation during ventricular tachycardia. These results demonstrate that the thoracic autonomic nervous system can induce repetitive ventricular excitation originating from consistent loci.

The efficient synthesis of 4-hydroxylated metabolites of various retinoids is described. Allylic bromination of beta-ionone, retinal, and methyl retinoate followed by treatment with aqueous AcOH and saponification afforded the 4-hydroxy analogues in 64-79% yields. With the conjugated polyenes, retinal and methyl retinoate, about 25% of the products were found to be the 13-cis isomers. Purification of isomer mixtures by HPLC permitted stereochemical assignments after 1H NMR analysis.

To investigate the mechanism of uniform ventricular tachycardia induced by programmed stimulation, we recorded His bundle electrograms and unipolar electrograms from 64 subepicardial, subendocardial, and intramural sites in dogs. Isochronal maps were generated off-line by computer. Two groups of dogs were studied 3 days after occlusion of their left anterior descending coronary arteries; one group underwent reperfusion after 2 to 2.5 hr of occlusion and the other methylprednisolone treatment before permanent occlusion. In the former, subepicardial sequences presented either a pattern suggesting circus movement or a radial pattern in which excitation at intramural sites could precede earliest subepicardial excitation. In the latter preparations, subepicardial excitation patterns consistently suggested circus movement in the subepicardial muscle layer surviving over necrotic tissue. Assuming complete circus movement, the "missed" time interval, measured as the interval left unaccounted for by actual recording of local excitation between ventricular tachycardia cycles, ranged from 3% to 64% of the cycle length of ventricular tachycardia. While surviving subepicardial and intramural layers appeared to be involved in the mechanism of ventricular tachycardia, a late second breakthrough on the right ventricle, in conjunction with fixed-coupled H deflections on the His bundle electrograms, suggested the involvement of the conducting system in propagation of the impulse.

The relation between electrical alternans recorded in acutely ischaemic myocardium, ventricular arrhythmias, and changes in the activation sequence was studied. Sixty three unipolar electrograms, simultaneously recorded from the epicardial surface of the in situ porcine heart during 6 min periods of coronary occlusion, were converted to a digital format and analysed by computer. The time integrals of the electrograms during their QRS complex, ST segment, and T wave were measured. Unipolar waveform alternation was then quantified by subtracting the values obtained for two consecutive beats showing alternans. The spatial distribution of unipolar waveform alternation was illustrated by isoarea difference maps. Isochronal maps of the local excitation detected on each electrogram were constructed. Of 52 occlusions in 27 preparations, unipolar waveform alternation was detected in 42 and was promptly followed by ventricular arrhythmias in 37. The magnitude of unipolar waveform alternation increased from the margin to the inner portion of the ischaemic zone but its correlation with activation delay was poor, and there was no change in the activation sequence. These results suggest that beat to beat alternation of the unipolar waveform is related to changes in the action potential configuration rather than to changes in the activation sequence. Furthermore, unipolar waveform alternation appears to be associated with the development of early post-occlusion ventricular arrhythmias.