Effects of non-vascularized adipose tissue transplantation on its genetic profile.

Subcutaneous adipose tissue (SAT) is recognized as a highly active metabolic and inflammatory tissue. Interestingly, adipose tissue transplantation is widely performed in plastic surgery via lipofilling, yet little is known about the gene alteration of adipocytes after transplantation. We performed an RNA-expression analysis of fat transplants before and after fat transplantation.In C57BL/6 N mice SAT was autologously transplanted. Samples of SAT were analysed before transplantation, 7, and 15 days after transplantation and gene expression profiles were measured.Analysis revealed that lipid metabolism-related genes were downregulated while inflammatory and extracellular matrix related genes were up-regulated 7 and 15 days after transplantation. When comparing gene expression profile 7 days after transplantation to 15 days after transplantation developmental pathways showed most changes.

Peptidyl-Prolyl-cis/trans-Isomerases Mip and PpiB of Legionella pneumophila Contribute to Surface Translocation, Growth at Suboptimal Temperature, and Infection.

The gammaproteobacterium Legionella pneumophila is the causative agent of Legionnaires' disease, an atypical pneumonia that manifests itself with severe lung damage. L. pneumophila, a common inhabitant of freshwater environments, replicates in free-living amoebae and persists in biofilms in natural and man-made water systems. Its environmental versatility is reflected in its ability to survive and grow within a broad temperature range as well as its capability to colonize and infect a wide range of hosts, including protozoa and humans. Peptidyl-prolyl-cis/trans-isomerases (PPIases) are multifunctional proteins that are mainly involved in protein folding and secretion in bacteria. In L. pneumophila the surface-associated PPIase Mip was shown to facilitate the establishment of the intracellular infection cycle in its early stages. The cytoplasmic PpiB was shown to promote cold tolerance. Here, we set out to analyze the interrelationship of these two relevant PPIases in the context of environmental fitness and infection. We demonstrate that the PPIases Mip and PpiB are important for surfactant-dependent sliding motility and adaptation to suboptimal temperatures, features that contribute to the environmental fitness of L. pneumophila Furthermore, they contribute to infection of the natural host Acanthamoeba castellanii as well as human macrophages and human explanted lung tissue. These effects were additive in the case of sliding motility or synergistic in the case of temperature tolerance and infection, as assessed by the behavior of the double mutant. Accordingly, we propose that Mip and PpiB are virulence modulators of L. pneumophila with compensatory action and pleiotropic effects.

High resolution XUV Fourier transform holography on a table top.

Today, coherent imaging techniques provide the highest resolution in the extreme ultraviolet (XUV) and X-ray regions. Fourier transform holography (FTH) is particularly unique, providing robust and straightforward image reconstruction at the same time. Here, we combine two important advances: First, our experiment is based on a table-top light source which is compact, scalable and highly accessible. Second, we demonstrate the highest resolution ever achieved with FTH at any light source (34 nm) by utilizing a high photon flux source and cutting-edge nanofabrication technology. The performance, versatility and reliability of our approach allows imaging of complex wavelength-scale structures, including wave guiding effects within these structures, and resolving embedded nanoscale features, which are invisible for electron microscopes. Our work represents an important step towards real-world applications and a broad use of XUV imaging in many areas of science and technology. Even nanoscale studies of ultra-fast dynamics are within reach.

High speed and high resolution table-top nanoscale imaging.

We present a table-top coherent diffractive imaging (CDI) experiment based on high-order harmonics generated at 18 nm by a high average power femtosecond fiber laser system. The high photon flux, narrow spectral bandwidth, and high degree of spatial coherence allow for ultrahigh subwavelength resolution imaging at a high numerical aperture. Our experiments demonstrate a half-pitch resolution of 15 nm, close to the actual Abbe limit of 12 nm, which is the highest resolution achieved from any table-top extreme ultraviolet (XUV) or x-ray microscope. In addition, sub-30 nm resolution was achieved with only 3 s of integration time, bringing live diffractive imaging and three-dimensional tomography on the nanoscale one step closer to reality. The current resolution is solely limited by the wavelength and the detector size. Thus, table-top nanoscopes with only a few-nanometer resolutions are in reach and will find applications in many areas of science and technology.

Image formation properties and inverse imaging problem in aperture based scanning near field optical microscopy.

Aperture based scanning near field optical microscopes are important instruments to study light at the nanoscale and to understand the optical functionality of photonic nanostructures. In general, a detected image is affected by both the transverse electric and magnetic field components of light. The discrimination of the individual field components is challenging as these four field components are contained within two signals in the case of a polarization resolved measurement. Here, we develop a methodology to solve the inverse imaging problem and to retrieve the vectorial field components from polarization and phase resolved measurements. Our methodology relies on the discussion of the image formation process in aperture based scanning near field optical microscopes. On this basis, we are also able to explain how the relative contributions of the electric and magnetic field components within detected images depend on the chosen probe. We can therefore also describe the influence of geometrical and material parameters of individual probes within the image formation process. This allows probes to be designed that are primarily sensitive either to the electric or magnetic field components of light.

Isolation of Staphylococcus sciuri from horse skin infection.

Staphylococcus sciuri is known as an opportunistic pathogen colonizing domesticated animals and has also been associated with wound infections in humans. Particularly over the last decade, oxacillin (methicillin) resistant strains had been emerged, which now increase the medical relevance of this species. This report describes the identification of an oxacillin-resistant S. sciuri isolate from a wound infection of a horse. We determined the absence of coagulase and hyaluronidase activity and analysed the antibiotic resistance profile.

Enhancing 18F-FDG-PET/CT analysis in lung cancer patients. Is CT-CT image fusion helpful in predicting pleural involvement? A pilot study.

AIM: To retrospectively evaluate the feasibility and value of CT-CT image fusion to assess the shift of peripheral lung cancers with/-out chest wall infiltration, comparing computed tomography acquisitions in shallow-breathing (SB-CT) and deep-inspiration breath-hold (DIBH-CT) in patients undergoing FDG-PET/CT for lung cancer staging. METHODS: Image fusion of SB-CT and DIBH-CT was performed with a multimodal workstation used for nuclear medicine fusion imaging. The distance of intrathoracic landmarks and the positional shift of tumours were measured using semi-transparent overlay of both CT series. Statistical analyses were adjusted for confounders of tumour infiltration. Cutoff levels were calculated for prediction of no-/infiltration. RESULTS: Lateral pleural recessus and diaphragm showed the largest respiratory excursions. Infiltrating lung cancers showed more limited respiratory shifts than non-infiltrating tumours. A large respiratory tumour-motility accurately predicted non-infiltration. However, the tumour shifts were limited and variable, limiting the accuracy of prediction. CONCLUSION: This pilot fusion study proved feasible and allowed a simple analysis of the respiratory shifts of peripheral lung tumours using CT-CT image fusion in a PET/CT setting. The calculated cutoffs were useful in predicting the exclusion of chest wall infiltration but did not accurately predict tumour infiltration. This method can provide additional qualitative information in patients with lung cancers with contact to the chest wall but unclear CT evidence of infiltration undergoing PET/CT without the need of additional investigations. Considering the small sample size investigated, further studies are necessary to verify the obtained results.

[VATS lobectomy--a standard procedure in the therapy for stage I non-small cell lung cancer?]. / VATS-Lobektomie--Ein Standardverfahren in der Therapie des nicht kleinzelligen Lungenkarzinoms im Stadium I?

Even though VATS lobectomy has been practised since 1991 in stage I of non-small cell lung cancer (NSCLC), it was not being considered equivalent to conventional lobectomy due to considerable doubts in terms of safety and oncological permissibility. This study describes our experience and an evaluation of the systematic establishment of lobectomy by means of video-assisted thoracic surgery (VATS) as standard treatment of NSCLC in stage I, which serves as an alternative to conventional surgery. For this purpose, 42 NSCLC patients in stage I, undergoing a conventional lobectomy in 2010 (group I), were retrospectively compared to 30 patients in the same tumour stage (group II) who were treated in 2011 using VATS lobectomy. The comparison of these two groups was drawn regarding operation time, number of resected lymph nodes, required analgesics, duration of drainage, rate of postoperative complications and length of hospital stay. Although VATS lobectomy requires a longer operation time of approximately 30 minutes, it shows significant advantages in reference to postoperative need of analgesics, duration of drainage and complications after surgery. Furthermore, the amount of resected lymph nodes was comparable in both groups. Therefore, VATS lobectomy constitutes an essential extension for the operative management in a lung cancer centre. Our results show that this new method is not only of equal, but of superior value compared to conventional lobectomy. Our experience and recent data in the literature illustrate that VATS lobectomy will play a decisive role in therapy for NSCLC in stage I, potentially even in stages II and IIIA.

[Physical lipolysis]. / Physikalische Lipolyse.

The number of non-invasive procedures is steadily growing. Lipolysis with focused ultrasound, cryo-lipolysis, laser-lipolysis, radiofrequency-lipolysis, HIFU-lipolysis, classic liposuction, vibration-assisted liposuction and water beam-assisted Liposuction are newly established procedures to treat diet- and sport resistant fat deposits. Non-invasive lipolysis with focused ultrasound in combination with radiofrequency as well as cryo-lipolysis are the most established procedures. The methods are described in detail and evaluated.

[Pathogenesis, diagnosis and therapy of Legionella infections]. / Pathogenese, Diagnostik und Therapie der Legionella-Infektion.

Legionella species are ubiquitous in aquatic environments. About 50 years ago they entered the engineered (technical) environment, i.e. warm water systems with zones of stagnation. Since that time they represent a hygienic problem. After transmission to humans via aerosols legionellae might cause Legionella pneumonia (legionnaires' disease) or influenza-like respiratory infections (Pontiac fever). Epidemiological data suggest that Legionella strains might differ substantially in their virulence properties. Although the molecular basis is not understood L. pneumophila serogroup 1 especially MAb 3/1-positive strains cause the majority of infections. The main virulence feature is the ability to multiply intracellularly. After uptake into macrophages legionellae multiply in a specialized vacuole and finally lyse their host cells. Several bacterial factors like surface components, secretion systems and iron uptake systems are involved in this process. Since the clinical picture of Legionella pneumonia does not allow differentiation from pneumoniae caused by other pathogens, microbiological diagnostic methods are needed to establish the diagnosis. Cultivation of legionellae from clinical specimens, detection of antigens and DNA in patients' samples and detection of antibodies in serum samples are suitable methods. However, none of the diagnostic tests presently available offers the desired quality with respect to sensitivity and specificity. Therefore, the standard technique is to use several diagnostic tests in parallel. Advantages and disadvantages of the diagnostic procedures are discussed. Therapeutic options for Legionella infections are newer macrolides like azithromycin and chinolones (ciprofloxacin, levofloxacin and moxifloxacin).

Microbial diversity of marine sponges.

The recent application of molecular microbial ecology tools to sponge-microbe associations has revealed a glimpse into the biodiversity of these microbial communities, that is considered just 'the tip of the iceberg'. This chapter provides an overview over these new findings with regard to identity, diversity and distribution patterns of sponge-associated microbial consortia. The sponges Aplysina aerophoba (Verongida), Rhopaloeides odorabile (Dicytoceratida) and Theonella swinhoei (Lithistida) were chosen as model systems for this review because they have been subject to both, cultivation-dependent and cultivation-independent approaches. A discussion of the microbial assemblages of Halichondriapanicea is presented in the accompanying chapter by Imhoff and Stöhr. Considering that a large fraction of sponge-associated microbes is not yet amenable to cultivation, an emphasis has been placed on the techniques centering around the 16S rRNA gene. A section has been included that covers the potential of sponge microbial communities for drug discovery. Finally, a 'sponge-microbe interaction model' is presented that summarizes our current understanding of the processes that might have shaped the community structure of the microbial assemblages within sponges.

Differential expression and activity status of MMP-1, MMP-2 and MMP-9 in tumor and stromal cells of squamous cell carcinomas of the lung.

Matrix metalloproteinases (MMPs) play a key role in cancer progression. Interstitial collagenase (MMP-1) and type IV collagenases (MMP-2, MMP-9) are involved in the initial breakdown of collagen and basement membrane components during tumor growth and invasion. Besides tumor cells, fibroblasts are especially involved in MMP production. The aim of this study was to quantify MMP-1, MMP-2 and MMP-9 within tumor cells and tumor-surrounding fibroblasts compared to normal lung epithelial cells to gain an insight into the function of these MMPs in squamous cell carcinomas of the lung. The expression and activity of MMP-1, MMP-2 and MMP-9 were analyzed in 30 squamous cell carcinomas and in normal lung tissue from the same patients by immunohistology and gelatin zymography. The majority of tumor cells were positive for MMP-1 (mean +/- SD: 67.3 +/- 26.7%) and MMP-9 (64.7 +/- 22.8%), whereas a significantly lower percentage of normal bronchoepithelial cells (47.3 +/- 25.4 and 40.3 +/- 24.2%, respectively; p < 0.01) and fibroblasts located in the tumor-surrounding tissue (39.7 +/- 14.3 and 38.1 +/- 24.1%, respectively; p < 0.01) expressed these MMPs. Only a few tumor cells showed any immunoreactivity for MMP-2 (4.4 +/- 6.7%), whereas a higher percentage of fibroblasts tested positive for this enzyme (8.6 +/- 13.1%; p < 0.01). Using gelatin zymography, we could demonstrate that MMP-2 is activated in the tumor only, not in normal lung tissue. The coordinated expression of MMP-1, MMP-2 and MMP-9 in tumor cells and/or their induction in tumor-surrounding fibroblasts and further activation in the tumor tissue may be involved in the high invasive and metastatic potential of squamous cell carcinomas of the lung. Comparing the results from immunohistology and zymography can give indications for distribution and activity of proteinases, especially certain MMPs such as MMP-2.

[Intraoperative electrophysiological monitoring of the recurrent laryngeal nerve in thyroid gland surgery--a prospective study]. / Anwendung des Neuromonitoring des N. laryngeus recurrens in der Schilddrüsenchirurgie - eine prospektive Studie.

INTRODUCTION: In the last few years the use of intraoperative electrophysiological monitoring of the recurrent laryngeal nerve (RLN) in thyroid gland surgery has become more and more important. PATIENTS AND METHOD: In a prospective study 223 nerves at risk in 116 patients were monitored with the Neurosign(R)100 (Fa. Magstim Ltd., UK). We used intramuscular needle electrodes inserted into the vocal muscle through the conic ligament. Practicability, complications, acceptance and predictive value of the method were documented. Recurrent nerve palsy rate and complications were compared with a control group operated upon without monitoring. RESULTS: The intraoperative delay using this method was on average 8.9 minutes. There were problems with monitoring equipment avoiding use in 6.4 %. In 2 cases (1.7 %) an accidental lesion of endotracheal tube cuff was found related to malpositioning of the needle and in 7.7 % a hematoma of the vocal cords was observed. 73.3 % of the surgeons accepted the method to identify and control the nerve integrity. False-positive and false-negative signals may occur. In cases of a final real stimulus response a regular vocal cord motility was found in 95 %. If a nerve conduction block was noted an immobility of ipsilateral vocal cord was diagnosed postoperatively in 50 %. There was no decrease in transient recurrent palsy rate using monitoring (10.7 % vs. 9.6 % without monitoring) but in permanent paralysis (1.8 % vs. 3.0 %). CONCLUSIONS: It may be concluded that intraoperative electrophysiological monitoring of the RLN is a simple and accepted method with low complications reducing the incidence of permanent RLN palsy rate. We found the monitoring especially useful for operations of recurrent goiter and carcinomas of the thyroid gland as well as for learning thyroid gland surgery.

Graves' disease is associated with an altered CXCR3 and CCR5 expression in thyroid-derived compared to peripheral blood lymphocytes.

The mechanisms by which T cells accumulate in the thyroid and support the autoimmune process in patients with Graves' disease (GD) are poorly understood. Chemokines and their receptors may be involved in this process. We have analysed the expression of CXCR3 and CCR5 as Th1-specific chemokine receptors, CCR3 as a marker for Th2 cells, CXCR4 (expressed on unprimed, naive T cells) and CCR2 (known to be involved in autoimmunity) on peripheral blood (PBL) and thyroid-derived lymphocytes (TL) using flow cytometry. Chemokine receptor expression on PBL of GD patients (n = 16) did not differ from that of normal controls (n = 10). In GD, CXCR3+ (67.3 +/- 4.0% versus 45.7 +/- 2.1%) and CCR5+ T cells (42.5 +/- 3.4% versus 18.8 +/- 2.1%) showed a significant enrichment in the TL compared to PBL. The positive cells were contributed mainly by the CD4+CD45R0+ subset. TL are mostly primed CD45R0+ T cells, but surprisingly, they had significantly higher levels of CXCR4+ cells among TL (96.2 +/- 1.0%) compared to PBL (66.8 +/- 4.2%). However, CXCR4 has been induced during in vitro isolation of TL. There was no correlation between chemokine receptors and the level of TSH-receptor and thyroid peroxidase autoantibodies. CCR3+ and CCR2+ cells remained unchanged in TL compared to PBL. We could confirm the results using RT PCR and immunohistology. In summary, TL showed a different chemokine receptor pattern compared to PBL from the same patient. This indicates a role for CXCR3 and CCR5 in the recruitment of T cells to the thyroid in GD.

The Lly protein is essential for p-hydroxyphenylpyruvate dioxygenase activity in Legionella pneumophila.

The lly locus confers fluorescence, haemolysis, brown pigmentation and an increased resistance to light in Legionella pneumophila. In this study, we correlated the pigment production of two lly-positive L. pneumophila isolates and a recombinant lly-positive Escherichia coli strain with the presence of homogentisic acid (HGA) in the culture supernatant. The detection of HGA by high performance liquid chromatography and the data analysis of the deduced amino acid sequence of the lly gene indicate that the lly locus codes for a p-hydroxyphenylpyruvate dioxygenase (HPPD). This enzyme catalyses the transformation of p-hydroxyphenylpyruvate into HGA, which subsequently oxidises and polymerises into a melanin-like pigment. One open reading frame (ORF 1) in the lly region exhibited homologies with genes of Synechocystis sp., Petroselium crispum and Streptomyces mycarofaciens that code for methyltransferases. By screening a genomic library of L. pneumophila (serogroup 1) strain Corby with a monoclonal antibody against the legiolysin (lly), we identified two recombinant E. coli clones that did not produce the brown pigment and showed no haemolysis and fluorescence. DNA sequencing revealed that both clones contained 874 nucleotides of the N-terminal part of the lly gene. The recombinant strains expressed truncated legiolysin proteins of 39.5 and 35.7 kDa and did not produce HGA. Considering the highly conserved structure of legiolysin-like HPPD genes from other organisms, we suggest that the C-terminus of the legiolysin may be essential for the enzymatic activity that conferred pigmentation via HGA polymerisation, haemolysis and fluorescence.

GRO-alpha in normal and pathological thyroid tissues and its regulation in thyroid-derived cells.

Thyroid glands affected by Graves' disease (GD) show striking leukocytic infiltration, mainly by T-cells. The mechanisms by which the various leukocytes are maintained in the thyroid are unknown. Growth-regulated oncogene-alpha (GRO-alpha) in interaction with its receptor CXCR2 is a chemoattractant for both T-cells and neutrophils and may be one of the chemokines involved in the cell maintenance. GRO-alpha and CD18 mRNA as a marker of leukocytic infiltration were quantified in thyroid tissue using competitive RT-PCR. We found very high GRO-alpha mRNA levels in all thyroid tissues. In GD patients (n=16), the GRO-alpha mRNA did not correlate with the CD18 mRNA level or thyroid peroxidase and TSH-receptor antibodies in patients' sera. In thyroid autonomy (n=10), the GRO-alpha mRNA levels were significantly lower in autonomous single adenomas compared with the corresponding normal tissue. In order to define the cellular source of GRO-alpha mRNA and protein, we examined various thyroid-derived cells. Thyrocytes, thyroid-derived leukocytes and fibroblasts showed basal GRO-alpha mRNA and protein expression, which was remarkably upregulated by different stimuli in vitro. The expression of GRO-alpha by thyroid carcinoma cell lines confirms that thyrocytes may actually produce GRO-alpha. As shown by flow cytometry and immunohistology, CD68+ monocytes/macrophages are the only cell population strongly expressing CXCR2 in the thyroid.