Novel multiparameter correlates of Coxiella burnetii infection and vaccination identified by longitudinal deep immune profiling.

Q-fever is a flu-like illness caused by Coxiella burnetii (Cb), a highly infectious intracellular bacterium. There is an unmet need for a safe and effective vaccine for Q-fever. Correlates of immune protection to Cb infection are limited. We proposed that analysis by longitudinal high dimensional immune (HDI) profiling using mass cytometry combined with other measures of vaccination and protection could be used to identify novel correlates of effective vaccination and control of Cb infection. Using a vaccine-challenge model in HLA-DR transgenic mice, we demonstrated significant alterations in circulating T-cell and innate immune populations that distinguished vaccinated from naïve mice within 10 days, and persisted until at least 35 days post-vaccination. Following challenge, vaccinated mice exhibited reduced bacterial burden and splenomegaly, along with distinct effector T-cell and monocyte profiles. Correlation of HDI data to serological and pathological measurements was performed. Our data indicate a Th1-biased response to Cb, consistent with previous reports, and identify Ly6C, CD73, and T-bet expression in T-cell, NK-cell, and monocytic populations as distinguishing features between vaccinated and naïve mice. This study refines the understanding of the integrated immune response to Cb vaccine and challenge, which can inform the assessment of candidate vaccines for Cb.

[Ocular sarcoidosis]. / Okuläre Sarkoidose.

Ocular manifestation of sarcoidosis occurs in up to 60% of patients with confirmed systemic sarcoidosis and represents one of the most common forms of noninfectious uveitis. In known pulmonary sarcoidosis, ocular involvement can occur in up to 80% of cases. Sarcoidosis can also present only in the eye, without a systemic manifestation (ocular sarcoidosis). Typically, ocular sarcoidosis shows bilateral granulomatous uveitis and can involve all parts of the eye. Apart from an acute anterior uveitis, chronic intermediate or posterior uveitis can be found. In order to prevent a severe reduction of visual acuity leading to blindness, early diagnosis and treatment is essential. For diagnosis, specific clinical signs involving the eye (bilateral granulomatous changes in all parts of the eye) and typical laboratory investigations (angiotensin-converting enzyme, ACE; lysozyme; soluble interleukin 2 receptor, sIL2R; chest X­ray; chest CT) have to be taken into account, since biopsy to prove noncaseating granulomas is not performed with changes restricted to the eye due to the high risk of vision loss. Ocular sarcoidosis mostly responds well to local or systemic steroid treatment. If the therapeutic effect is insufficient, immunosuppressive agents and biologics can be applied.

[Choroidal tumor with exsudative retinal detachment?]. / Aderhauttumor mit exsudativer Amotio retinae?

We present the case of a 41-year-old female patient with acute onset of nyctalopia, xanthopsia and impairment of visual field. The ophthalmologic examination revealed peripheral choroidal elevation with accompanying retinal detachment. Due to the the presence of a nanophthalmic eye without any evidence of inflammation or malignant growth, the diagnosis uveal effusion syndrome could be made.

The lazaroid tirilazad is a new inhibitor of direct and indirect UVA-induced lipid peroxidation in human dermal fibroblasts.

Lipid peroxidation caused by oxidative stress within the tissue leads to destruction and dysfunction of cellular membranes. Human dermal fibroblasts in the skin are subject to constant photooxidative stress caused mainly by deeply penetrating UVA irradiation. Therefore, the membrane damage caused by this photooxidative stress may be a major promoter of photoaging and photocarcinogenic processes initiated and promoted by long-term UVA exposure of the skin. The oxidative destruction is counterbalanced by a complex network of enzymatic and nonenzymatic antioxidants creating the skin's line of defence against UVA-induced reactive oxygen species. The lazaroid tirilazad represents a new synthetic group of antioxidants with structural molecular similarity to glucocorticosteroids. We investigated the antioxidative capacity of tirilazad by determining its effects on the levels of malondialdehyde (MDA), as a marker of lipid peroxidation, induced directly or indirectly by UVA in human dermal fibroblasts. In a time- and dose-dependent kinetic, we demonstrated that fibroblasts incubated with tirilazad are well protected against subsequent UVA irradiation and show no increase in MDA levels similar to the unirradiated controls. This was also observed when lipid peroxidation was caused chemically by incubation of human dermal fibroblasts with 200 micro M Fe(3+)-citrate and 1 m M ascorbyl phosphate as a model of indirect UVA-induced skin damage. Lysates of fibroblasts treated this way showed a tenfold increase in MDA levels, whereas preincubation with tirilazad resulted in a significantly lower increase in MDA levels. Furthermore, in a comparison with the well-established radical scavenger Trolox, an alpha-tocopherol analogue, tirilazad offered better protection to the membranes. Our results demonstrate for the first time that the lazaroid tirilazad is an effective inhibitor of direct and indirect UVA-induced increases in MDA as a marker of lipid peroxidation in human dermal fibroblasts.

[pH values in chronic wounds. Evaluation during modern wound therapy]. / pH-Wert des Milieus chronischer Wunden. Untersuchungen im Rahmen einer modernen Wundtherapie.

The pH value of chronic wounds influences directly or indirectly many factors important to wound healing. We measured 247 pH values in 39 patients with chronic wounds of varying origins, detecting values from 5.45 to 8.65. Over a period of 12 months, individual values varied up to 1.73 units. Knowledge of pH values helps insure an adequate chronic wound therapy. For example, many enzymatic wound products have pH values outside the above values. Our results also show that the pH value is a dynamic factor which should be monitored regularly. Our studies offer additional insight into the specific features of chronic wounds and allow a more individualized therapy.

L-type calcium channels: binding domains for dihydropyridines and benzothiazepines are located in close proximity to each other.

We investigated the binding of a fluorescent diltiazem analogue (3R,4S)-cis-1-[2-[[3-[[3-[4,4-difluoro-3a,4-dihydro-5,7-dimethyl-4-bo ra-3a,4a-diaza-s-indacen-3-yl]propionyl]amino]propyl]amin o]ethy]-1,3,4,5-tetrahydro-3-hydroxy-4-(4-methoxyphenyl)-6-(triflu oromethyl)-2H-1-benzazepin-2-one (DMBODIPY-BAZ) to L-type Ca2+ channels in the presence of different 1,4-dihydropyridines (DHPs) by using fluorescence resonance energy transfer (FRET) [Brauns, T., Cai, Z.-W., Kimball, S. D., Kang, H.-C., Haugland, R. P., Berger, W., Berjukov, S., Hering, S., Glossmann, H., & Striessnig, J. (1995) Biochemistry 34, 3461]. When channels are occupied with DMBODIPY-BAZ, a rapid fluorescence change occurred upon addition of different DHPs. The direction of this intensity modulation was found to be only dependent on the chemical composition of the dihydropyridine employed. DHPs containing a nitro group decreased, whereas others (e.g., isradipine) enhanced the fluorescence signal. In addition, all DHPs markedly decreased the association rate constant for DMBODIPY-BAZ without affecting equilibrium binding. Both observations together are best explained by a steric model where the DHP binding site is located in close proximity to the accession pathway of DMBODIPY-BAZ.

Benzothiazepine binding domain of purified L-type calcium channels: direct labeling using a novel fluorescent diltiazem analogue.

We have synthesized a series of N-propylamino-substituted benzazepinones (NPSBs) as specific probes for the benzothiazepinone (BTZ) binding domain of muscle L-type calcium channels (LTCCs). NPSBs were identified which possess high affinity for the channel after purification. We synthesized a fluorescent NPSB, DMBODIPY-BAZ, as the first benz(othi)azepinone derivative known to reversibly label partially purified LTCCs. DMBODIPY-BAZ binds to the partially purified channel with high affinity (Kd = 25 nM, Bmax = 580 pmol/mg of protein). Fluorescence resonance energy transfer (FRET) occurred between tryptophan residues of the channel protein and the DMBODIPY fluorophore upon specific drug binding. FRET was exploited to allow highly time-resolved detection of specific drug binding kinetics. We found that the dissociation half-life (t1/2) of DMBODIPY-BAZ decreased with the concentration of an unlabeled competitor, which indicates ligand-induced accelerated dissociation. In contrast, t1/2 was concentration-dependently increased by the dihydropyridine (DHP) (+)-isradipine. These kinetic properties of DMBODIPY-BAZ indicate that a high-affinity BTZ binding domain also exists on purified LTCCs. NPSBs represent novel tools to provide further insight into the molecular pharmacology of the BTZ binding domain on LTCCs.