Paul Hengster

The cell-type specificity of mitochondrial dynamics

Recent advances in mitochondrial imaging have revealed that in many cells mitochondria can be highly dynamic. They can undergo fission/fusion processes modulated by various mitochondria-associated proteins and also by conformational transitions in the inner mitochondrial membrane. Moreover, precise mitochondrial distribution can be achieved by their movement along the cytoskeleton, recruiting various connector and motor proteins. Such movement is evident in various cell types ranging from yeast to mammalian cells and serves to direct mitochondria to cellular regions of high ATP demand or to transport mitochondria destined for elimination. Existing data also demonstrate that many aspects of mitochondrial dynamics, morphology, regulation and intracellular organization can be cell type-/tissue-specific. In many cells like neurons, pancreatic cells, HL-1 cells, etc., complex dynamics of mitochondria include fission, fusion, small oscillatory movements of mitochondria, larger movements like filament extension, retraction, fast branching in the mitochondrial network and rapid long-distance intracellular translocation of single mitochondria. Alternatively, mitochondria can be rather fixed in other cells and tissues like adult cardiomyocytes or skeletal muscles with a very regular organelle organization between myofibrils, providing the bioenergetic basis for contraction. Adult cardiac cells show no displacement of mitochondria with only very small-amplitude rapid vibrations, demonstrating remarkable, cell type-dependent differences in the dynamics and spatial arrangement of mitochondria. These variations and the cell-type specificity of mitochondrial dynamics could be related to specific cellular functions and demands, also indicating a significant role of integrations of mitochondria with other intracellular systems like the cytoskeleton, nucleus and endoplasmic reticulum (ER).

Valproic acid modulates islet cell insulin secretion: a possible mechanism of weight gain in epilepsy patients.

A significant weight gain in the course of treatment of epilepsy with valproic acid (VPA) was described in several clinical studies. We recently demonstrated that postprandial insulin levels are increased in patients with VPA therapy. This possible modulation of pancreatic insulin secretion by VPA could be due to the structure of VPA as a fatty acid derivative and thus to direct stimulation of pancreatic insulin secretion or competition with free fatty acids (FFA) for albumin binding. In order to investigate the effect of VPA on insulin secretion in pancreatic islet cells we performed in vitro experiments with islets from pancreases of multiorgan donors. After preparation, the incubation with valproate caused a time and dose-dependent increase of insulin concentration in the cell supernatant. This could also be demonstrated with the control drug, lorazepam, a benzodiazepine, but not with mirtazepin and phenytoin. It can be speculated that an increase in pancreatic insulin secretion under chronic VPA treatment enhances appetite and energy storage and is related to the observed weight gain.

Cytomegalovirus Infections After Treatment With Daclizumab, An Anti Il-2 Receptor Antibody, For Prevention Of Renal Allograft Rejection

Daclizumab is a newly developed humanized anti-IL-2 receptor monoclonal antibody. We describe the effect of adding daclizumab to conventional dual or triple cyclosporine A immunosuppressive therapy on the incidence and nature of cytomegalovirus (CMV) infections in patients receiving a first cadaveric renal graft. In the triple therapy study there was no evidence of any difference in CMV rate or course of disease between the two treatment arms, although in the dual therapy study a decrease in the incidence of CMV infection was observed in the patients treated with daclizumab. The onset of CMV disease was markedly delayed in the daclizumab groups in both studies. Daclizumab can effectively reduce the risk of acute rejection without causing a concomitant increase in opportunistic infections, and by decreasing the need for antirejection therapy may also have a beneficial effect on CMV infection rates.

Surgical treatment of bacillus Calmette Guérin lymphadenitis.

Abstract. Although its protective effect is contested and the risk of contracting tuberculosis is rather low nowadays, BCG vaccination is frequently performed. Changes of strain repeatedly led to an increased complication rate. In Austria between 1990 and 1991, of 3386 newborn babies (Strain Pasteur) 116 developed lymphadenitis 3 to 28 weeks after vaccination. The affected children received four types of treatment: nothing specific, isoniazid, or surgery with and without isoniazid. Surgical treatment was found to be necessary in 96 cases. Bacilli were successfully grown in culture in 46% of cases up to week 20 after vaccination; but later than 20 weeks no culture became positive. All cultured bacteria were isoniazid-sensitive. From our data we drew the following conclusions: Isoniazid therapy did not prove successful when inflamed lymph nodes exceeded a certain size. Suppurative lymphadenitis in lymph nodes exceeding 1.0 to 1.5 cm usually led to infiltration or even perforation of the skin. Surgery prevents these complications and significantly reduces healing time. Adjuvant isoniazid therapy cannot be recommended, except for generalized BCG tuberculosis.

Dkk-3 expression in the tumor endothelium: a novel prognostic marker of pancreatic adenocarcinomas.

Dkk‐3 is proposed to be a new specific marker for tumor endothelial cells. Here we analyzed the clinical relevance of Dkk‐3 expression in pancreas adenocarcinomas and determined its role on endothelial cell growth in vitro. Microvessel density in tumor samples was immunohistochemically determined using Dkk‐3 and CD31 as endothelial cell markers, respectively. Based on the median microvessel density as a cut‐off point, patients were categorized into high and low microvessel density groups and a correlation with survival and clinical parameters was assessed. Moreover, the role of Dkk‐3 expression on chemosensitivity of endothelial cells was analyzed. In contrast to CD31 staining, Dkk‐3‐positive vessels were found only in tumor tissue and Dkk‐3 microvessel density significantly correlated negative with tumor grading. In survival analysis the median survival time was 7 months for patients with Dkk‐3 low, and 15 months for Dkk‐3 high microvessel density (P = 0.0013). Subset analysis of patients receiving gemcitabine therapy showed that overall survival was significantly decreased in Dkk‐3 low tumors than in high tumors (P = 0.009). In Cox regression Dkk‐3 emerged as a significant independent parameter (P = 0.024). Dkk‐3 overexpression in endothelial cells resulted in significantly enhanced growth inhibition after 5‐fluorouracil or gemcitabine treatment compared to control endothelial cells and cancer cell lines. Dkk‐3 low microvessel density was associated with tumor progression and worse clinical outcome. Overexpression of Dkk‐3 enhanced endothelial cell growth inhibition to chemotherapeutic drugs. Therefore, we suggest that Dkk‐3 high microvessel density may help to select patients who may benefit from chemotherapy. (Cancer Sci 2009)

A cluster of rotavirus enteritis in adult transplant recipients

Diarrhea following solid organ transplantation is a common side effect of some immunosuppressive agents but can also be caused by many pathogens. An outbreak of rotavirus (RV) enteritis presenting with severe diarrhea in four solid organ recipients was analyzed. The first case was diagnosed in a 6‐month‐old liver recipient who was prehospitalized on a pediatric ward. Within 1 month, three adult patients (two liver, one renal recipient) presented with enteritis. During diarrhea a significant rise in tacrolimus levels was observed. One patient developed toxic megacolon with ulcerative colitis. Infections were self‐limiting but led to secondary infectious complications and prolonged hospitalization. This is the first reported outbreak of RV enteritis in a multiorgan transplant unit involving adult patients. Although no fingerprinting or subtyping of the virus was performed we assume the child was the primary source. In transplant recipients presenting with diarrhea RV infection should be considered.

Peroxisome Proliferator-Activated Receptor-α Activation Inhibits Langerhans Cell Function

Epidermal Langerhans cells (LC) play a pivotal role in initiating and maintaining primary immune responses in the skin. In the present study, we asked whether peroxisome proliferator-activated receptor-α (PPARα) activation modulates LC function. Our results show that PPARα is expressed in immature LC and is down-regulated in mature LC suggesting that an early decrease of PPARα expression in LC may allow them to mature after contact with an Ag. We further show that pharmacologic PPARα activation inhibits LC maturation, migratory capacity, cytokine expression, and the ability to drive T cell proliferation. Moreover, PPARα activation inhibits NF-κB but not stress-activated protein kinase/JNK, p38MAPK, and ERK1/2. In conclusion, PPARα activation by endogenous ligands may provide a molecular signal that allows LC to remain in an immature state within the epidermis for extended periods of time despite minor environmental stimuli.

Protective effects of various preservation solutions on cultured endothelial cells.

Vascular endothelium represents the first target in organ preservation and plays an important role in reperfusion injury. Bovine aortic endothelial cells were cultivated and the most commonly used preservation solutions, such as University of Wisconsin HTK (Brettschneiders histidine-tryptophane-ketoglutarate), and Euro-Collins solutions were tested on the endothelial monolayer. In addition, one group of cultivated cells was preserved with cold saline solution, and endothelial monolayers grown in culture medium were used as controls. The quality of preservation was assessed after 24, 48, and 72 hours of cold storage. Reperfusion was simulated and its effects were observed by reincubation in culture medium at 37 degrees C for 6 hours. The total number of cells, cell viability (determined using trypan blue exclusion), and morphologic alterations were determined. Prostacyclin release was evaluated as a biochemical marker. University of Wisconsin solution maintains more than 99% cell viability after rewarming after both 24 and 48 hours of cold storage. After 72 hours, 86.7% of cells were still viable. Preservation with HTK and Euro-Collins solution allowed cell survival for only 24 hours (96.7%, HTK; 49.9%, Euro-Collins), with no viable cells seen after 48 hours. The cold saline-preserved sample showed 57.8% viable cells after 24 hours and 29.7% after 48 hours. No viable cells were detectable after 72 hours. Light microscopy revealed several patterns of both structural damage and intracellular change (nucleus and cytoplasm) in the endothelial monolayer after preservation with HTK, Euro-Collins solution, and cold saline solution. No morphologic alterations were seen in the University of Wisconsin solution group for as long as 72 hours.(ABSTRACT TRUNCATED AT 250 WORDS)

Irradiation Causes Biphasic Neutrophilic Granulocyte Phagocytic Function

Background and Purpose:The anti-inflammatory effect of low-dose radiotherapy is clinically well described. Nevertheless, until now neither the optimal dose nor the background of tissue reactions have been defined. The current study examines the influence of low radiation doses on neutrophilic granulocyte function, which could be helpful in finding the optimal dose for either stimulation or suppression of anti-inflammatory activity.Material and Methods:Lymphoprep® density gradient-purified neutrophilic granulocytes of three voluntary, healthy donors were used for all experiments. Granulocytes were incubated 48 h in RPMI 1640 and irradiated with single doses of 0.5, 1.0, 1.5, 2.0, 2.5, 3.0, 3.5, 4.0, 6.0, and 12 Gy using a 137Cs IBL 437L irradiator. Their function was assessed by measuring granulocytic release of reactive oxygen species (ROS) with luminol-enhanced chemiluminescence after stimulation with phorbol myristate acetate (PMA).Results:Relative changes of ROS release (ROS release before stimulation was set to 100%) increased after stimulation with PMA (mean ± standard deviation [SD]): 0 Gy: 147.6% ± 60%; 0.5 Gy: 153.6% ± 70%; 1.0 Gy: 164.9% ± 63%; 1.5 Gy: 177.8% ± 66%; 2.0 Gy: 162.5% ± 57%; 2.5 Gy: 156.2% ± 60%; 3.0 Gy: 159.2% ± 60%; 3.5 Gy: 126.9% ± 55%; 4.0 Gy: 137.9% ± 71%; 6.0 Gy: 148.3% ± 65%; 12.0 Gy: 156.1% ± 52%. The relative ROS release showed a significant increase at 1.5 Gy (p < 0.001) after PMA stimulation and a significant decrease of ROS release at 3.5 Gy (p < 0.005) and less markedly at 4.0 Gy (p < 0.05). 6.0 and 12.0 Gy showed a significant (p < 0.05) increase again.Conclusion:This ex vivo in vitro study on native human neutrophilic granulocytes shows an increase at 1.5 Gy and a significant decrease of granulocyte function at 3.5 and 4.0 Gy, as it was described for different other phenomena in low-dose radiotherapy. These results may provide a further explanation for the local anti-inflammatory effect of low-dose ionizing irradiationHintergrund und Ziel:Der antiinflammatorische Effekt niedrigdosierter Strahlentherapie ist klinischerseits bereits gut beschrieben. Dennoch konnte bis heute weder die optimale Dosierung noch die Grundlage der Gewebereaktion definiert werden. Die vorliegende Studie untersucht den Einfluss niedriger Strahlendosen auf die Funktion neutrophiler Granulozyten, um so Hinweise auf die optimale Dosis zur Stimulation bzw. Unterdrückung antiinflammatorischer Aktivität zu erhalten.Material und Methodik:Native humane neutrophile Granulozyten wurden von drei gesunden Probanden mittels Dichtegradienten gewonnen. Die Granulozyten wurden 48 h in RPMI 1640 inkubiert und mittels eines 137Cs-IBL-437L-Bestrahlungsgeräts mit 0,5, 1, 1,5, 2, 2,5, 3, 3,5, 4, 6 und 12 Gy bestrahlt. Ihre Funktion wurde anhand der granulozytären Freisetzung von reaktiven Sauerstoffradikalen (ROS [„reactive oxygen species“]) mittels luminolverstärkter Chemilumineszenz nach Stimulation mit Phorbol-Myristat-Acetat (PMA) gemessen.Ergebnisse:Die relativen Änderungen der ROS-Freisetzung (ROS-Freisetzung vor Stimulation wurde gleich 100% gesetzt) nach Stimulation mit PMA betrugen (Mittelwert ± Standardabweichung [SD]): 0 Gy: 147,6% ± 60%; 0,5 Gy: 153,6% ± 70%; 1 Gy: 164,9% ± 63%; 1,5 Gy: 177,8% ± 66%; 2 Gy: 162,5% ± 57%; 2,5 Gy: 156,2% ± 60%; 3 Gy: 159,2% ± 60%; 3,5 Gy: 126,9% ± 55%; 4 Gy: 137,9% ± 71%; 6 Gy: 148,3% ± 65%; 12 Gy: 156,1% ± 52%. Die relative ROS-Freisetzung zeigte einen signifikanten Anstieg bei 1,5 Gy (p < 0,001) nach PMA-Stimulation, allerdings einen deutlich signifikanten Abfall der ROS-Freisetzung bei 3,5 Gy (p < 0,005) und etwas weniger deutlich bei 4 Gy (p < 0,05). 6 und 12 Gy zeigten wieder einen signifikanten Anstieg.Schlussfolgerung:Diese Ex-vivo-in-vitro-Untersuchung an nativen humanen neutrophilen Granulozyten zeigte einen Anstieg der Granulozytenfunktion bei 1,5 Gy sowie einen signifikanten Abfall bei 3,5 und 4,0 Gy, wie er schon für verschiedene Phänomene niedrigdosierter Strahlentherapie beschrieben worden ist. Die Ergebnisse können einen weiteren Erklärungsansatz für den lokalen entzündungshemmenden Effekt niedrigdosierter Strahlentherapie bieten.

HIGH FREQUENCY OF HTLV-III ANTIBODIES AMONG HETEROSEXUAL INTRAVENOUS DRUG ABUSERS IN THE AUSTRIAN TYROL

34 intravenous drug abuse prisoners (5 females 2 of whom were prostitutes and 29 heterosexual males; aged 20-34) have been tested for antibodies to human T-lymphotropic virus type III (HTLV-III) both in Innsbruck (by enzyme-linked immunosorbent assay (ELISA) and immunoperoxidase straining. No one had had a recent infection at the time of the study. All these drug abusers had used intravenous opioids for 1-12 years and 18 had also taken cocaine in the 2-12 years before the study. However they had been free of drugs since their imprisonment 1 week to 18 months ealier. Immunological studies were done by fluorescence-activated cell sorting with antibodies Leu 3a Leu 2a. Neopterin a marker of the activation status of the cellular immune system was measured in urine and serum by high performance liquid chromatography and radioimmunoassay respectively. Raised neopterin levels have been found in AIDS patients and in risk groups for AIDS. Serum antibodies to HTLV-III were found in 15 (44%) of the drug addicts 14 seropositive individuals being heterosexual males and 1 a female prostitute. 15/29 (52%) had reversed helper-suppressor lymphocyte ratios. Neopterin levels were raised in 16/33 (48%) urine and 15/34 (44%) serum samples. The HTLV-III seropositive and seronegative groups were similar in respect of previous hepatitis infections hepatitis A and B virus antigen-antibody status duration and type of drug abuse and detoxification time. HTLV-III seropositive drug abusers were more likely to have raised neopterin levels and reversed T-helper suppressor ratios (0.9 or less). The significance of the raised neopterin levels in 4 subjects without HTLV-III antibodies is not clear. A prospective study is needed to show whether HTLV-III antibodies develop primarily among individuals with increased neopterin levels or independently of neopterin levels. The frequency of HTLV-III antibodies in drug abusers in the Austrian Tyrol is high compared with figures from the United States Britain and West Germany especially since the drug abusers we studied although in prison at the time do not live in an area of high risk for AIDS. The high frequency of HTLV-III in drug abusers has implications for the spread of the virus into the community at large since the heterosexual group especially after rehabilitation will not be so limited in their social contacts as homosexuals are. Futhermore some drug addicts are non-registered prostitutes. (full text)