Katinka Jung

Effects of water-filtered infrared-A and of heat on cell death, inflammation, antioxidative potential and of free radical formation in viable skin - First results

The effects of water-filtered infrared-A (wIRA) and of convective heat on viability, inflammation, inducible free radicals and antioxidative power were investigated in natural and viable skin using the ex vivo Bovine Udder System (BUS) model. Therefore, skin samples from differently treated parts of the udder of a healthy cow were analyzed using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) test, by prostaglandin E2 (PGE2) measurement and by electron spin resonance (ESR) spectroscopy. Neither cell viability, the inflammation status, the radical status or the antioxidative defence systems of the skin were significantly affected by wIRA applied within 30 min by using an irradiance of 1900 W m(-2) which is of relevance for clinical use, but which exceeded the maximum solar IR-A irradiance at the Earths surface more than 5 times and which resulted in a skin surface temperature of about 45 °C without cooling and of about 37 °C with convective cooling by air ventilation. No significant effects on viability and on inflammation were detected when convective heat was applied alone under equivalent conditions in terms of the resulting skin surface temperatures and exposure time. As compared with untreated skin, free radical formation was almost doubled, whereas the antioxidative power was reduced to about 50% after convective heating to about 45 °C.

SURF_ER - surface electron spin resonance (ESR) of the surface domain of large objects

SURF_ER is a method for spectral and spatial electron spin resonance measurements on the surface of large objects which extension is only restricted by the width of the pole gap of the magnet and the homogeneity of the magnetic field and not by the cavity dimensions. The application of several techniques like SURF_ER for spectroscopic measurements, SURF_ERM for spatial scanning and SURF_ERI for spatial measurements of the depth of the surface region are discussed and represented for the skin of a human being as an example.

STABILITY OF WATER-SOLUBLE AND LIPID-SOLUBLE PARAMAGNETIC PROBES IN BACILLUS SUBTILIS

Batch cultures of the Gram-positive bacterium Bacillus subtilis PB19 have been investigated for their metabolic action to electron spin resonance (ESR) probes. Five- and six-membered water-soluble and lipid-soluble nitroxides have been used, which were reduced most probably to the corresponding hydroxylamine derivatives. The reduction was followed by the ESR signal intensity and found to be dependent on chemical structure and stability, lipophilic/hydrophilic character, charge, concentration, and temperature. Water-soluble nitroxides did not show apparent toxicity towards B. subtilis, in contrast with n-DXSA (n=5, 12, 16) which were found to be strongly cytotoxic. The cytotoxicity depended on the position of the doxyl unit along the hydrocarbon chain. The hydrophilic nitroxides were reduced at a much slower rate relative to the lipophilic ones. Membrane diffusion was suggested to be a slower process relative to chemical reduction for water-soluble nitroxides. The lipophilic nitroxides were solubilized into the membrane where they were rapidly reduced with a reduction maximum at 303-310 K, which is the optimal growth temperature of B. subtilis, while an inactivation at higher temperatures was observed. Both toxicity and reduction rates of nitroxides strongly indicated that the reduction was an enzyme-mediated process taking place near the outer surface of the periplasmic membrane.

Reduction in the free radical status and clinical benefit of repeated intrathecal triamcinolone acetonide application in patients with progressive multiple sclerosis.

BackgroundPrevious open trials performed repeated intrathecal application of the sustained release steroid triamcinolone acetonide every third day in patients with progressive multiple sclerosis and described enhanced walking abilities. ObjectivesThe objectives of this study were to demonstrate the efficacy of 5 triamcinolone administrations every other day and to describe their effects on the amount of inducible free radicals in cerebrospinal fluid. Subjects/MethodsClinical ratings, determinations of maximum walking distance, and execution of an instrumental peg insertion test were performed at baseline and on each day after a triamcinolone injection in 21 patients with progressive multiple sclerosis. Induction of free radicals was assessed in cerebrospinal fluid before each triamcinolone application by electron spin resonance spectroscopy. ResultsScores for multiple sclerosis improved, walking distance increased, and necessary intervals for the peg insertion procedure were shortened. The amount of inducible free radicals decreased. ConclusionsRepeat triamcinolone application improves dysfunction of upper and lower extremities even when administered 5 times only and in series every other day. The declined potential for free radical synthesis may be caused by the anti-inflammatory effect of triamcinolone. It may contribute to suppress the smoldering, chronic inflammation, particularly in spinal lesions of patients with progressive multiple sclerosis. The enhanced arm function hypothetically reflects the effect on cervical and brain lesions due to the hypobaric features of triamcinolone.

Cu(II) complexes in bacterial growth medium: electron spin resonance study.

In this study we report a spectroscopic investigation on the structure and stability of Cu(II)-complexes that are formed in a minimum growth medium (MM), normally used for Bacillus subtilis cultures. As other transition metals, Cu(II) compounds are toxic to this bacterium and the toxicity depends on the Cu(II) concentration. MM contained NH4+ ions and asparagine (asn) as the source of inorganic and organic nitrogen. Both ESR and electronic spectra demonstrated the very important role played by the amino acid asparagine in the coordinative behaviour of Cu(II). In particular, three different complexes were evidenced: Cu(H2O)6(2+); Cu(asn)+ and Cu(asn)2. The relative amount of these three species strongly depended on pH, on Cu:asn ratio and on the presence of the phosphate ions. They were identified and evaluated quantitatively by extensive simulation of the electron spin resonance (ESR) spectra recorded in different experimental conditions. The bis-complex was found to be more stable in MM than in an asparagine-containing water solution with the same Cu:asn ratio. A comparison of the spectroscopic results with microbiological investigations is also made.

One-time intrathecal triamcinolone acetonide application alters the redox potential in cerebrospinal fluid of progressive multiple sclerosis patients: a pilot study:

Introduction: Cerebrospinal fluid analysis may provide insight into the interplay between chronic inflammation and response to treatment. Objectives: To demonstrate the impact of one intrathecal triamcinolone injection on the redox potential and on ascorbyl radical appearance in the cerebrospinal fluid of chronic progressive multiple sclerosis patients. Methods: A total of 16 patients received 40 mg triamcinolone. Electron-spin resonance spectroscopy measured the oxidation range after copper ion [Cu (II)] addition and ascorbyl-radical bioavailability. Results: There was an increase of Cu (II) ion absorption, which reflects an augmented content of reduced proteins. Ascorbyl radicals were present in contrast to healthy controls according to the literature. Conclusion: Intrathecal steroid application alters the redox potential in cerebrospinal fluid. Our findings support the beneficial role of steroids on oxidative stress generally demonstrated by ascorbyl radical appearance. Reactive oxygen species decline is necessary for an upregulated production of reduced proteins.

Electron spin resonance of copper(II) as a tool for the determination of asparagine concentration in Bacillus subtilis cultures.

A procedure is presented that is based on the detection of Cu(II)-asparagine complexes by quantitative ESR, and allows in a very simple and rapid manner to evaluate the changes of asparagine concentration during the entire time range of the growth of Bacillus subtilis in a typical growth medium. The analysis is carried out in terms of the decrease of the intensity of the ESR-active mono- and di-asparagine copper(II) complexes. It is resulted that at the end of the exponential growth the asparagine concentration was reduced to values as low as 2% of the initial value. The procedure here reported may be the basis of similar methods to be used for other amino acids and prokaryote systems.