Frank Schweitzer

Serum concentrations of asymmetric (ADMA) and symmetric (SDMA) dimethylarginine in patients with chronic kidney diseases.

BACKGROUND NO synthesis is inhibited by the dimethylarginine (DMA) ADMA, which accumulates, similar to SDMA, in the plasma of patients suffering from chronic renal failure (CRF). ADMA and possibly SDMA contribute to hypertension and atherosclerosis in patients with chronic renal disease: ADMA inhibits directly eNOS, whereas SDMA competes with the NO precursor arginine for uptake into the cells. METHODS In 26 control persons and 221 patients with kidney diseases of different stage as were CRF, end stage renal disease (ESRD), and patients after renal transplantation (RT), the plasma concentrations of ADMA (c(ADMA)), SDMA (c(SDMA)) and 20 endogenous amino acids (AA) were measured by HPLC and correlated to blood pressure, cardiac events, endothelial dysfunction, and diabetes mellitus. RESULTS Both ADMA (1.04+/-0.04 vs. 0.66+/-0.04 microM) and SDMA (2.69+/-0.12 vs. 0.49+/-0.03 microM) were significantly (p<0.001) elevated in all patients compared to healthy controls, whereas arginine concentration (51.4+/-2.3 vs. 76.0+/-5.2 microM) was decreased in dependence on the degree of kidney disease. In RT patients, SDMA levels were significantly decreased, but c(ADMA) remained enhanced. A strong correlation was found between SDMA and both serum urea and creatinine in CRF and RT patients. A linear correlation was found between ADMA and cholesterol concentrations in RT patients. Hypertension in CRF was accompanied by a further increase in the concentration of DMAs. There was no relation between DMAs and the occurrence of peripheral arterial occlusive disease or cerebrovascular diseases. In patients with cardiac diseases, c(SDMA) was additionally increased only in the CRF group. CONCLUSIONS In patients with chronic kidney disease, c(ADMA) and c(SDMA) are significantly increased but cardiovascular diseases are evidently not correlated to changes in DMA concentrations in this group of patients.

In vivo measurement of time-resolved autofluorescence at the human fundus

An experimental setup for measurement of time-resolved autofluorescence of the human eye fundus is demonstrated. The method combines laser scanning technique and time-correlated single photon counting. The light source is a laser diode, delivering pulses of about 100 ps duration at a repetition rate of 40 MHz. The excitation wavelength is 446 nm and the cutoff wavelength of fluorescence detection is at 475 nm. The autofluorescence can be determined with a spatial resolution of 80 x 80 microm2 and 25 ps time resolution. The fluorescence decay is optimally approximated by a biexponential model. The dominating lifetime tau1 is shortest in the macula (320 to 380 ps) and reaches 1500 ps in the optic disk. The lifetime tau2 varies between 2 ns and 5 ns, but the spatial distribution is more homogeneous. Respiration of 100% oxygen for 6 min leads to changes in the fluorescence lifetime pointing to detection of coenzymes. Diagrams of lifetime tau2 versus tau1 are well suited for comparison of substances. Such lifetime clusters of a 20 deg macular field of a young healthy subject and of a patient suffering from dry age-related macular degeneration overlap only partially with tau2-tau1 clusters of lipofuscin.

Spektrale Differenzierung in Eigenfluoreszenzbildern des Augenhintergrunds von Patienten mit altersabhängiger Makuladegeneration

ZusammenfassungHintergrundEine veränderte Eigenfluoreszenz des Augenhintergrunds wird gegenwärtig insbesondere im Zusammenhang mit der altersabhängigen Makuladegeneration diskutiert. Zu ihrer richtigen Bewertung sind jedoch weitere Kenntnisse über das chemische Substrat der Fluoreszenz erforderlich.MethodeDie Autofluoreszenz des Augenhintergrunds wurde mit einer Funduskamera beobachtet. Die Anregung erfolgte im blauen Spektralbereich. Zur Aufzeichnung der Fluoreszenz wurde eine 3-Chip-CCD-Farbkamera benutzt. Zum Vergleich wurden die Fluoreszenzspektren von Lipofuszin, A2-E, FAD, NADH2 und AGE mit dem Jenaer Ophthalmospektrometer aufgenommen.ErgebnisseIm Gegensatz zu einem gesunden Fundus zeigen die Autofluoreszenzaufnahmen an Patienten mit AMD deutlich lokalisierbare Areale spektral unterschiedlicher Fluoreszenzemission. Die jeweils gefundenen Emissionsspektren zeigen starke Überlappungen.SchlussfolgerungenDie vorgestellte Technik ermöglicht es, spektrale Unterscheidungen in Bildern der Autofluoreszenz des Augenhintergrunds vorzunehmen. Neben der des Lipofuszins wurde bei Patienten mit nichtexsudativer AMD eine kurzwelligere Fluoreszenz beobachtet.AbstractIntroductionAlterations in the intrinsic ocular fundus fluorescence are under discussion particularly in connection with age-related macular degeneration. However, further knowledge of the chemical substrate of fluorescence is necessary.MethodOcular fundus autofluorescence was observed using a fundus camera. The fluorescence emission was recorded using a 3-chip CCD camera. For comparison, the fluorescence spectra of lipofuscin, A2-E, FAD, NADH2 and AGE’s were recorded by the Jena ophthalmo-spectrometer.ResultsIn contrast to the homogeneous intrinsic fluorescence of a normal fundus, the fluorescence images of patients suffering from AMD showed remarkable local differences. The detected fluorescence spectra showed remarkable overlaps.DiscussionWe introduced a new technique enabling the detection of spectral differences in images of ocular fundus autofluorescence. Besides the fluorescence of lipofuscin, we found a green one in patients suffering from nonexudative AMD.

Comparison of Time-Resolved Autofluorescence in the Eye-Ground of Healthy Subjects and Patients Suffering from Age-Related Maculer Degeneration

Age-related macular degeneration (AMD) is the most frequent cause for blindness for person older than 65 years in western countries. Besides the subjective pain, it is also an economic problem in the ageing population. As the pathomechanism is unknown, no curative treatment is possible. International research for discovering of early age-related alterations at the fundus is directed on in vivo measurements of autofluorescence. One way is the measurement of fluorescence spectra. Unfortunately, any selective excitation of fluorophores is excluded by the absorption edge of the ocular media at 400 nm. Furthermore, the shape of fluorescence spectra is influenced by the spectral absorption of layers in front of the emitting fluorophore. Weakly emitting fluorophores are covered by intensive emitting substances. The most serious limitation in fluorescence measurements of the living human fundus is the maximal permissible exposure. For that reason, fluorescence spectra of the fundus can not be detected with a high spatial resolution. The detection of dynamic fluorescence provides substance-specific lifetimes Ti, amplitudes Ai, and information about the relative contribution Qi of components in fluorescence images. As these parameters are calculated for each image point, diagrams of Ti vs. Tj, Ai vs. Aj, and Qi vs. Qj can be drawn, in which specific clusters appear for healthy subjects or AMD - patients. The projection of lifetime - clusters onto corresponding axis represents histogram of the considered lifetime. The slope in the correlation between Ai and Aj can also be used as a discriminating mark. Considering image lines as intersection through characteristic anatomical structures (optic disc, macula) profiles of Ti, Ai, or Qi show changes of these parameters (e.g. depigmentation) as function of location, which might be specific for AMD.

In vivo autofluorescence lifetime imaging at the fundus of the human eye

Changes in cellular metabolism are considered first signs of fundus diseases, e.g. of age-related macular degeneration. Changes in the metabolism can potentially be detected by measuring the autofluorescence of the fundus. The fundus contains a wide variety of fluorophores in different binding and quenching states. The fluorescence signals cannot be clearly discriminated by commonly used steady state imaging techniques, even when these are combined with spectral resolution and excitation wavelength multiplexing. A considerable improvement is obtained by fluorescence lifetime imaging (FLIM). FLIM not only adds an additional discrimination parameter to distinguish different fluorophores but also resolves different quenching states of the same fluorophore. Due to its high sensitivity and high time resolution, its capability to resolve multi-exponential decay functions, and its easy combination with fast scanning we use multi-dimensional time-correlated single photon counting for fundus imaging. By analyzing the spectral properties of the expected fluorophores in the fundus, we show that improved discrimination of fluorophores is obtained by FLIM in combination with selected excitation wavelength and emission wavelength. As demonstrated in lifetime histograms of 40° fundus images, several fluorophores are excited at 446 nm, but predominantly lipofuscin at 468 nm excitation. Simultaneous detection of lifetime images in two emission ranges 500 nm to 560 nm and 560 nm to 700 nm improves further the discrimination of fluorophores.