Development of a Laser Microdissection-Coupled Quantitative Shotgun Lipidomic Method to Uncover Spatial Heterogeneity.

Lipid metabolic disturbances are associated with several diseases, such as type 2 diabetes or malignancy. In the last two decades, high-performance mass spectrometry-based lipidomics has emerged as a valuable tool in various fields of biology. However, the evaluation of macroscopic tissue homogenates leaves often undiscovered the differences arising from micron-scale heterogeneity. Therefore, in this work, we developed a novel laser microdissection-coupled shotgun lipidomic platform, which combines quantitative and broad-range lipidome analysis with reasonable spatial resolution. The multistep approach involves the preparation of successive cryosections from tissue samples, cross-referencing of native and stained images, laser microdissection of regions of interest, in situ lipid extraction, and quantitative shotgun lipidomics. We used mouse liver and kidney as well as a 2D cell culture model to validate the novel workflow in terms of extraction efficiency, reproducibility, and linearity of quantification. We established that the limit of dissectible sample area corresponds to about ten cells while maintaining good lipidome coverage. We demonstrate the performance of the method in recognizing tissue heterogeneity on the example of a mouse hippocampus. By providing topological mapping of lipid metabolism, the novel platform might help to uncover region-specific lipidomic alterations in complex samples, including tumors.

Multifaceted Analyses of Isolated Mitochondria Establish the Anticancer Drug 2-Hydroxyoleic Acid as an Inhibitor of Substrate Oxidation and an Activator of Complex IV-Dependent State 3 Respiration.

The synthetic fatty acid 2-hydroxyoleic acid (2OHOA) has been extensively investigated as a cancer therapy mainly based on its regulation of membrane lipid composition and structure, activating various cell fate pathways. We discovered, additionally, that 2OHOA can uncouple oxidative phosphorylation, but this has never been demonstrated mechanistically. Here, we explored the effect of 2OHOA on mitochondria isolated by ultracentrifugation from U118MG glioblastoma cells. Mitochondria were analyzed by shotgun lipidomics, molecular dynamic simulations, spectrophotometric assays for determining respiratory complex activity, mass spectrometry for assessing beta oxidation and Seahorse technology for bioenergetic profiling. We showed that the main impact of 2OHOA on mitochondrial lipids is their hydroxylation, demonstrated by simulations to decrease co-enzyme Q diffusion in the liquid disordered membranes embedding respiratory complexes. This decreased co-enzyme Q diffusion can explain the inhibition of disjointly measured complexes I-III activity. However, it doesn't explain how 2OHOA increases complex IV and state 3 respiration in intact mitochondria. This increased respiration probably allows mitochondrial oxidative phosphorylation to maintain ATP production against the 2OHOA-mediated inhibition of glycolytic ATP production. This work correlates 2OHOA function with its modulation of mitochondrial lipid composition, reflecting both 2OHOA anticancer activity and adaptation to it by enhancement of state 3 respiration.

Endovascular Recanalization of Tandem Internal Carotid Occlusions Using the Balloon-assisted Tracking Technique.

PURPOSE: Tandem occlusive lesions are responsible for up to 20% of acute ischemic stroke cases and are associated with poor prognosis if complete recanalization cannot be achieved. Endovascular recanalization might be challenging due to difficulties in the safe passage of the occluded plaque at the origin of the internal carotid artery (ICA). The balloon-assisted tracking technique (BAT), where a partially deflated balloon is exposed out of the catheter tip to facilitate its passage through stenosed or spastic arterial segments was introduced by interventional cardiologists and the applicability of the technique has been recently proposed in the field of neurointervention as well. Here we describe our experience using the BAT technique in the endovascular recanalization of tandem occlusive lesions. METHODS: Procedures were performed from June 2013 to December 2020 in a single center. Baseline clinical and imaging data, procedural and follow-up details and clinical outcomes were retrospectively collected. RESULTS: In this study 107 patients, median age 66 years, median admission NIHSS 14 and median ASPECTS 8 were included. Successful recanalization of the ICA using the BAT technique was achieved in 100 (93%) and successful intracranial revascularization in 88 (82%) patients. There were no complications attributable to the BAT technique. Intraprocedural complications occurred in 9 (8%) patients. Emergent stenting was performed in 40 (37%) at the end of the procedure. Postprocedural adverse events (intracerebral hemorrhage [ICH], malignant infarction) occurred in 6 (5%) patients. Good clinical outcome at 3 months (modified Rankin scale [mRS] 0-2) was 54 (50%) and mortality 26 (24%). Delayed stent placement during follow-up occurred in 21 cases. CONCLUSION: Application of BAT technique in tandem occlusions appears feasible, safe, and efficient. Further evaluation of this technique is awaited.

Heat therapy shows benefit in patients with type 2 diabetes mellitus: a systematic review and meta-analysis.

AIMS: Type-2 diabetes mellitus (T2DM) is a common health condition which prevalence increases with age. Besides lifestyle modifications, passive heating could be a promising intervention to improve glycemic control. This study aimed to assess the efficacy of passive heat therapy on glycemic and cardiovascular parameters, and body weight among patients with T2DM. METHODS: A systematic review and meta-analysis were reported according to PRISMA Statement. We conducted a systematic search in three databases (MEDLINE, Embase, CENTRAL) from inception to 19 August 2021. We included interventional studies reporting on T2DM patients treated with heat therapy. The main outcomes were the changes in pre-and post-treatment cardiometabolic parameters (fasting plasma glucose, glycated plasma hemoglobin, and triglyceride). For these continuous variables, weighted mean differences (WMD) with 95% confidence intervals (CIs) were calculated. Study protocol number: CRD42020221500. RESULTS: Five studies were included in the qualitative and quantitative synthesis, respectively. The results showed a not significant difference in the hemoglobin A1c [WMD -0.549%, 95% CI (-1.262, 0.164), p = 0.131], fasting glucose [WMD -0.290 mmol/l, 95% CI (-0.903, 0.324), p = 0.355]. Triglyceride [WMD 0.035 mmol/l, 95% CI (-0.130, 0.200), p = 0.677] levels were comparable regarding the pre-, and post intervention values. CONCLUSION: Passive heating can be beneficial for patients with T2DM since the slight improvement in certain cardiometabolic parameters support that. However, further randomized controlled trials with longer intervention and follow-up periods are needed to confirm the beneficial effect of passive heat therapy.

Male and Female Animals Respond Differently to High-Fat Diet and Regular Exercise Training in a Mouse Model of Hyperlipidemia.

Inappropriate nutrition and a sedentary lifestyle can lead to obesity, one of the most common risk factors for several chronic diseases. Although regular physical exercise is an efficient approach to improve cardiometabolic health, the exact cellular processes are still not fully understood. We aimed to analyze the morphological, gene expression, and lipidomic patterns in the liver and adipose tissues in response to regular exercise. Healthy (wild type on a normal diet) and hyperlipidemic, high-fat diet-fed (HFD-fed) apolipoprotein B-100 (APOB-100)-overexpressing mice were trained by treadmill running for 7 months. The serum concentrations of triglyceride and tumor necrosis factor α (TNFα), as well as the level of lipid accumulation in the liver, were significantly higher in HFD-fed APOB-100 males compared to females. However, regular exercise almost completely abolished lipid accumulation in the liver of hyperlipidemic animals. The expression level of the thermogenesis marker, uncoupling protein-1 (Ucp1), was significantly higher in the subcutaneous white adipose tissue of healthy females, as well as in the brown adipose tissue of HFD-fed APOB-100 females, compared to males. Lipidomic analyses revealed that hyperlipidemia essentially remodeled the lipidome of brown adipose tissue, affecting both the membrane and storage lipid fractions, which was partially restored by exercise in both sexes. Our results revealed more severe metabolic disturbances in HFD-fed APOB-100 males compared to females. However, exercise efficiently reduced the body weight, serum triglyceride levels, expression of pro-inflammatory factors, and hepatic lipid accumulation in our model.

Lipid and protein tumor markers for head and neck squamous cell carcinoma identified by imaging mass spectrometry.

Head and neck squamous cell carcinoma (HNSCC) is the sixth most common cancer worldwide. To improve pre- and post-operative diagnosis and prognosis novel molecular markers are desirable. Here we used MALDI imaging mass spectrometry (IMS) and immunohistochemistry (IHC) to seek tumor specific expression of proteins and lipids in HNSCC samples. Among low molecular weight proteins visualized, S100A8 and S100A9 were found to be expressed in the regions of tumor tissue but not in the surrounding healthy stroma of a post-operative microdissected tissue. Marker potential of S100A8 and S100A9 was confirmed by immunohistochemistry of paraffin-embedded pathological samples. Imaging lipids showed a remarkable depletion of lysophosphatidylcholine species LPC[16:0], LPC[18:2] and, in parallel, accumulation of major glycerophospholipid species PE-P[36:4], PC[32:1], PC[34:1] in neoplastic areas. This was confirmed by shotgun lipidomics of dissected healthy and tumor tissue sections. A combination of the negative (LPC[16:0]) and positive (PC[32:1], PC[34:1]) markers was also applicable to uncover tumorous character of a pre-operative biopsy. Furthermore, marker potential of lysophospholipids was supported by elevated expression levels of the lysophospholipid degrading enzyme lysophospholipase A1 (LYPLA1) in the tumor regions of paraffin-embedded HNSCC samples. Finally, experimental evidence of 3D cell spheroid tests showed that LPC[16:0] facilitates HNSCC invasion, implying that HNSCC progression in vivo may be dependent on lysophospholipid supply. Altogether, a series of novel proteins and lipid species were identified by IMS and IHC screening, which may serve as potential molecular markers for tumor diagnosis, prognosis, and may pave the way to better understand HNSCC pathophyisiology.

Modulation of Plasma Membrane Composition and Microdomain Organization Impairs Heat Shock Protein Expression in B16-F10 Mouse Melanoma Cells.

The heat shock response (HSR) regulates induction of stress/heat shock proteins (HSPs) to preserve proteostasis during cellular stress. Earlier, our group established that the plasma membrane (PM) acts as a sensor and regulator of HSR through changes in its microdomain organization. PM microdomains such as lipid rafts, dynamic nanoscale assemblies enriched in cholesterol and sphingomyelin, and caveolae, cholesterol-rich PM invaginations, constitute clustering platforms for proteins functional in signaling cascades. Here, we aimed to compare the effect of cyclodextrin (MßCD)- and nystatin-induced cholesterol modulations on stress-activated expression of the representative HSPs, HSP70, and HSP25 in mouse B16-F10 melanoma cells. Depletion of cholesterol levels with MßCD impaired the heat-inducibility of both HSP70 and HSP25. Sequestration of cholesterol with nystatin impaired the heat-inducibility of HSP25 but not of HSP70. Imaging fluorescent correlation spectroscopy marked a modulated lateral diffusion constant of fluorescently labelled cholesterol in PM during cholesterol deprived conditions. Lipidomics analysis upon MßCD treatment revealed, next to cholesterol reductions, decreased lysophosphatidylcholine and phosphatidic acid levels. These data not only highlight the involvement of PM integrity in HSR but also suggest that altered dynamics of specific cholesterol pools could represent a mechanism to fine tune HSP expression.

Sustained maternal smoking-associated changes in the physico-chemical properties of fetal RBC membranes might serve as early markers for vascular comorbidities.

Maternal smoking-induced congenital heart and microvascular defects are closely associated with the impaired functioning of the in-utero feto-placental circulation system. Current groundbreaking facts revealed intimate crosstalk between circulating red blood cells (RBCs) and the vascular endothelium. Thus, RBCs have become the protagonists under varied pathological and adverse pro-oxidative cellular stress conditions. We isolated and screened fetal RBCs from the arterial cord blood of neonates, born to non-smoking (RBC-NS) and smoking mothers (RBC-S), assuming that parameters of fetal RBCs are blueprints of conditions experienced in-utero. Using atomic force microscopy and mass spectrometry-based shotgun lipidomics in the RBC-S population we revealed induced membrane stiffness, loss in intrinsic plastic activities and several abnormalities in their membrane-lipid composition, that could consequently result in perturbed hemodynamic flow movements. Altogether, these features are indicative of the outcome of neonatal microvascular complications and suggest unavailability for the potential rescue mechanism in cases of vascular endothelium impairment due to altered membrane integrity and rheological properties.

Dihydropyridine Derivatives Modulate Heat Shock Responses and have a Neuroprotective Effect in a Transgenic Mouse Model of Alzheimer's Disease.

Heat shock proteins (Hsps) have chaperone activity and play a pivotal role in the homeostasis of proteins by preventing misfolding, by clearing aggregated and damaged proteins from cells, and by maintaining proteins in an active state. Alzheimer's disease (AD) is thought to be caused by amyloid-ß peptide that triggers tau hyperphosphorylation, which is neurotoxic. Although proteostasis capacity declines with age and facilitates the manifestation of neurodegenerative diseases such as AD, the upregulation of chaperones improves prognosis. Our research goal is to identify potent Hsp co-inducers that enhance protein homeostasis for the treatment of AD, especially 1,4-dihydropyridine derivatives optimized for their ability to modulate cellular stress responses. Based on favorable toxicological data and Hsp co-inducing activity, LA1011 was selected for the in vivo analysis of its neuroprotective effect in the APPxPS1 mouse model of AD. Here, we report that 6 months of LA1011 administration effectively improved the spatial learning and memory functions in wild type mice and eliminated neurodegeneration in double mutant mice. Furthermore, Hsp co-inducer therapy preserves the number of neurons, increases dendritic spine density, and reduces tau pathology and amyloid plaque formation in transgenic AD mice. In conclusion, the Hsp co-inducer LA1011 is neuroprotective and therefore is a potential pharmaceutical candidate for the therapy of neurodegenerative diseases, particularly AD.

Reduced expression of CDP-DAG synthase changes lipid composition and leads to male sterility in Drosophila.

Drosophila spermatogenesis is an ideal system to study the effects of changes in lipid composition, because spermatid elongation and individualization requires extensive membrane biosynthesis and remodelling. The bulk of transcriptional activity is completed with the entry of cysts into meiotic division, which makes post-meiotic stages of spermatogenesis very sensitive to even a small reduction in gene products. In this study, we describe the effect of changes in lipid composition during spermatogenesis using a hypomorphic male sterile allele of the Drosophila CDP-DAG synthase (CdsA) gene. We find that the CdsA mutant shows defects in spermatid individualization and enlargement of mitochondria and the axonemal sheath of the spermatids. Furthermore, we could genetically rescue the male sterile phenotype by overexpressing Phosphatidylinositol synthase (dPIS) in a CdsA mutant background. The results of lipidomic and genetic analyses of the CdsA mutant highlight the importance of correct lipid composition during sperm development and show that phosphatidic acid levels are crucial in late stages of spermatogenesis.

Lipidomic analysis reveals a radiosensitizing role of gamma-linolenic acid in glioma cells.

Previous studies have demonstrated that gamma-linolenic acid (GLA) is effective against glioma cells under both in vitro and in vivo conditions. In the present study we determined how GLA alone or in combination with irradiation alters the fatty acid (FA) and lipid profiles, the lipid droplet (LD) content, the lipid biosynthetic gene expression and the apoptosis of glioma cells. In GLA-treated cells direct correlations were found between the levels of various FAs and the expression of the corresponding FA biosynthetic genes. The total levels of saturated and monosaturated FAs decreased in concert with the down-regulation of FASN and SCD1 gene expression. Similarly, decreased FADS1 gene expression was paralleled by lowered arachidonic acid (20:4 n-6) and eicosapentaenoic acid (20:5 n-3) contents, while the down-regulation of FADS2 expression was accompanied by a diminished docosahexaenoic acid (22:6 n-3) content. Detailed mass spectrometric analyses revealed that individual treatments gave rise to distinct lipidomic fingerprints. Following uptake, GLA was subjected to elongation, resulting in dihomo-gamma-linolenic acid (20:3 n-6, DGLA), which was used for the synthesis of the LD constituent triacylglycerols and cholesteryl esters. Accordingly, an increased number of LDs were observed in response to GLA administration after irradiation. GLA increased the radioresponsiveness of U87 MG cells, as demonstrated by an increase in the number of apoptotic cells determined by FACS analysis. In conclusion, treatment with GLA increased the apoptosis of irradiated glioma cells, and GLA might therefore increase the therapeutic efficacy of irradiation in the treatment of gliomas.

Membrane lipid therapy: Modulation of the cell membrane composition and structure as a molecular base for drug discovery and new disease treatment.

Nowadays we understand cell membranes not as a simple double lipid layer but as a collection of complex and dynamic protein-lipid structures and microdomains that serve as functional platforms for interacting signaling lipids and proteins. Membrane lipids and lipid structures participate directly as messengers or regulators of signal transduction. In addition, protein-lipid interactions participate in the localization of signaling protein partners to specific membrane microdomains. Thus, lipid alterations change cell signaling that are associated with a variety of diseases including cancer, obesity, neurodegenerative disorders, cardiovascular pathologies, etc. This article reviews the newly emerging field of membrane lipid therapy which involves the pharmacological regulation of membrane lipid composition and structure for the treatment of diseases. Membrane lipid therapy proposes the use of new molecules specifically designed to modify membrane lipid structures and microdomains as pharmaceutical disease-modifying agents by reversing the malfunction or altering the expression of disease-specific protein or lipid signal cascades. Here, we provide an in-depth analysis of this emerging field, especially its molecular bases and its relevance to the development of innovative therapeutic approaches.

Membrane fluidity matters: hyperthermia from the aspects of lipids and membranes.

Hyperthermia is a promising treatment modality for cancer in combination both with radio- and chemotherapy. In spite of its great therapeutic potential, the underlying molecular mechanisms still remain to be clarified. Due to lipid imbalances and 'membrane defects' most of the tumour cells possess elevated membrane fluidity. However, further increasing membrane fluidity to sensitise to chemo- or radiotherapy could have some other effects. In fact, hyperfluidisation of cell membrane induced by membrane fluidiser initiates a stress response as the heat shock protein response, which may modulate positively or negatively apoptotic cell death. Overviewing some recent findings based on a technology allowing direct imaging of lipid rafts in live cells and lipidomics, novel aspects of the intimate relationship between the 'membrane stress' of tumour cells and the cellular heat shock response will be highlighted. Our findings lend support to both the importance of membrane remodelling and the release of lipid signals initiating stress protein response, which can operate in tandem to control the extent of the ultimate cellular thermosensitivity. Overall, we suggest that the fluidity variable of membranes should be used as an independent factor for predicting the efficacy of combinational cancer therapies.

Hydroximic acid derivatives: pleiotropic HSP co-inducers restoring homeostasis and robustness.

According to the "membrane sensor" hypothesis, the membrane's physical properties and microdomain organization play an initiating role in the heat shock response. Clinical conditions such as cancer, diabetes and neurodegenerative diseases are all coupled with specific changes in the physical state and lipid composition of cellular membranes and characterized by altered heat shock protein levels in cells suggesting that these "membrane defects" can cause suboptimal hsp-gene expression. Such observations provide a new rationale for the introduction of novel, heat shock protein modulating drug candidates. Intercalating compounds can be used to alter membrane properties and by doing so normalize dysregulated expression of heat shock proteins, resulting in a beneficial therapeutic effect for reversing the pathological impact of disease. The membrane (and lipid) interacting hydroximic acid (HA) derivatives discussed in this review physiologically restore the heat shock protein stress response, creating a new class of "membrane-lipid therapy" pharmaceuticals. The diseases that HA derivatives potentially target are diverse and include, among others, insulin resistance and diabetes, neuropathy, atrial fibrillation, and amyotrophic lateral sclerosis. At a molecular level HA derivatives are broad spectrum, multi-target compounds as they fluidize yet stabilize membranes and remodel their lipid rafts while otherwise acting as PARP inhibitors. The HA derivatives have the potential to ameliorate disparate conditions, whether of acute or chronic nature. Many of these diseases presently are either untreatable or inadequately treated with currently available pharmaceuticals. Ultimately, the HA derivatives promise to play a major role in future pharmacotherapy.

Nutritional lipid supply can control the heat shock response of B16 melanoma cells in culture.

The in vitro culture of cells offers an extremely valuable method for probing biochemical questions and many commonly-used protocols are available. For mammalian cells a source of lipid is usually provided in the serum component. In this study we examined the question as to whether the nature of the lipid could become limiting at high cell densities and, therefore, prospectively influence the metabolism and physiology of the cells themselves. When B16 mouse melanoma cells were cultured, we noted a marked decrease in the proportions of n-3 and n-6 polyunsaturated fatty acids (PUFAs) with increasing cell density. This was despite considerable quantities of these PUFAs still remaining in the culture medium and seemed to reflect the preferential uptake of unesterified PUFA rather than other lipid classes from the media. The reduction in B16 total PUFA was reflected in changes in about 70% of the molecular species of membrane phosphoglycerides which were analysed by mass spectrometry. The importance of this finding lies in the need for n-3 and n-6 PUFA in mammalian cells (which cannot synthesize their own). Although the cholesterol content of cells was unchanged the amount of cholesterol enrichment in membrane rafts (as assessed by fluorescence) was severely decreased, simultaneous with a reduced heat shock response following exposure to 42°C. These data emphasize the pivotal role of nutrient supply (in this case for PUFAs) in modifying responses to stress and highlight the need for the careful control of culture conditions when assessing cellular responses in vitro.

[Endoscopic submucosal dissection in our practice -- new possibilities in the endoscopic treatment of neoplastic changes in the alimentary canal]. / Endoszkópos submucosus dissectio saját gyakorlatunkban -- új lehetoség a tápcsatorna korai neoplasztikus elváltozásainak endoszkópos kezelésében.

UNLABELLED: Endoscopic submucosal dissection is a promising new endoscopic technique for en block resection of early, superficial, flat, and laterally spreading gastrointestinal neoplasms. The primary aims of this prospective study were to summarize the first Hungarian experience with endoscopic submucosal dissection, and to compare the experimental in vivo procedures in pigs with the human application of this technique in two academic centers. METHODS: The ex vivo Erlangen pig model was used until initial competence was achieved. Thereafter 15 endoscopic submucosal dissection procedures were performed in up to 5 sessions in living pigs under general anesthesia. After submucosal injection with saline Fujinone ball tip, a Flush knife was used with a transparent hood for circumferential incision and submucosal dissection. The following variables were analyzed: specimen size, complete and en bloc resection rate, total duration of the procedure, and complications. Furthermore, between 2009 and 2012, 14 endoscopic submucosal dissections were carried out in humans; 1 duodenal, 3 gastric and 10 colorectal interventions were performed due to large laterally spreading intramucosal neoplasms. The specimen size, en bloc resection rate, total duration of the procedure and complications were next analyzed. RESULTS: 87% complete resection rate, 29.4 ± 19.5 cm2 specimen size and 136.2 ± 26.8 min of procedure duration were achieved in the live pig models. Nontransmural damages of the muscular layer occurred in 3/15 (20%) cases which were successfully closed with hemoclips, except in one pig that died due to fatal esophageal perforation. All bleedings were easily managed with hot biopsy forceps coagulation and hemoclips without hemodynamic instability. In humans, endoscopic submucosal dissection was accomplished with an 11/14 (79%) en block resection rate, with average resected specimen size of 11.56 ± 4.9 cm2 and with 157 ± 55 min of operating time. Two cases of bleeding and 2 cases of perforation occurred (14%), all of which were managed endoscopically. After 1.5 year of average follow up, local recurrence occurred in one case only. In addition, due to the 3 incomplete resections and unsuccessful endoscopic submucosal dissection, surgical intervention was required in 4 cases altogether. CONCLUSIONS: This study demonstrates the first Hungarian experience with endoscopic submucosal dissection. Training in live pig models could help endoscopists to overcome the learning curve and minimize the risk of complications before starting the procedure in humans. Reduction in the resection time and low risk of complications, especially bleeding, could be achieved by the application of a flush knife.

Membrane-lipid therapy in operation: the HSP co-inducer BGP-15 activates stress signal transduction pathways by remodeling plasma membrane rafts.

Aging and pathophysiological conditions are linked to membrane changes which modulate membrane-controlled molecular switches, causing dysregulated heat shock protein (HSP) expression. HSP co-inducer hydroxylamines such as BGP-15 provide advanced therapeutic candidates for many diseases since they preferentially affect stressed cells and are unlikely have major side effects. In the present study in vitro molecular dynamic simulation, experiments with lipid monolayers and in vivo ultrasensitive fluorescence microscopy showed that BGP-15 alters the organization of cholesterol-rich membrane domains. Imaging of nanoscopic long-lived platforms using the raft marker glycosylphosphatidylinositol-anchored monomeric green fluorescent protein diffusing in the live Chinese hamster ovary (CHO) cell plasma membrane demonstrated that BGP-15 prevents the transient structural disintegration of rafts induced by fever-type heat stress. Moreover, BGP-15 was able to remodel cholesterol-enriched lipid platforms reminiscent of those observed earlier following non-lethal heat priming or membrane stress, and were shown to be obligate for the generation and transmission of stress signals. BGP-15 activation of HSP expression in B16-F10 mouse melanoma cells involves the Rac1 signaling cascade in accordance with the previous observation that cholesterol affects the targeting of Rac1 to membranes. Finally, in a human embryonic kidney cell line we demonstrate that BGP-15 is able to inhibit the rapid heat shock factor 1 (HSF1) acetylation monitored during the early phase of heat stress, thereby promoting a prolonged duration of HSF1 binding to heat shock elements. Taken together, our results indicate that BGP-15 has the potential to become a new class of pharmaceuticals for use in 'membrane-lipid therapy' to combat many various protein-misfolding diseases associated with aging.

Heat stress causes spatially-distinct membrane re-modelling in K562 leukemia cells.

Cellular membranes respond rapidly to various environmental perturbations. Previously we showed that modulations in membrane fluidity achieved by heat stress (HS) resulted in pronounced membrane organization alterations which could be intimately linked to the expression and cellular distribution of heat shock proteins. Here we examine heat-induced membrane changes using several visualisation methods. With Laurdan two-photon microscopy we demonstrate that, in contrast to the enhanced formation of ordered domains in surface membranes, the molecular disorder is significantly elevated within the internal membranes of cells preexposed to mild HS. These results were compared with those obtained by anisotropy, fluorescence lifetime and electron paramagnetic resonance measurements. All probes detected membrane changes upon HS. However, the structurally different probes revealed substantially distinct alterations in membrane heterogeneity. These data call attention to the careful interpretation of results obtained with only a single label. Subtle changes in membrane microstructure in the decision-making of thermal cell killing could have potential application in cancer therapy.

[Computerized modelling of operation (CMO) before thoracoplasty]. / Thoraco-myoplastica mutéteink számítógépes modellezése és tervezése.

INTRODUCTION: Thoraco-myoplasty (TMP) has proven to be the only successful method of treatment for severe cases of chronic pyothorax (CPT). Great loss of muscle tissue and permanent bone-structure defects with severe functional damage can be caused by this method. The aim of the applied CMO was to prevent additional loss of muscle function. Preoperative evaluation of spiral CT scan 3-dimensional imaging provided a significant aid in decreasing adverse effects of the TMP. MATERIALS AND METHOD: Between 1990 and 2010, TMP was applied in 85 patients, whose CPT came from several different origins. CLINICAL DATA: average age: 62.7; mortality: 4.8%. Tissue and function preserving TMP was achieved following open treatment after thoracic fenestration in 76 cases, and drainage with continuous suction was performed in 9 patients prior to this. In our department TMP with computerized modeling was introduced as the definitive treatment of CPT in 2006, since then it has been applied in 8 patients. RESULTS: The number and length of the ribs to be resected can be anticipated applying spiral CT imaging. Furthermore, accurate volume measurement of the empyema cavity and rotatable muscles (pectoral major and latissimus dorsal) faciliatates elimination of the primary defect during surgery. CONCLUSION: Therefore, we believe that CMO can provide significant aid for surgeons to reduce the negative effects caused by muscle trauma and the structural changes in the thoracic wall.

[Vacuum assisted closure (VAC) of pyothorax with bronchial fistula following radiotherapy, pneumonectomy and thoraco-myoplasty]. / Irradiatiót és pneumonectomiát követo hörgosipolyos empyema thoracis nyitott, vákuumtámogatott (VAC) kezelése thoraco-myoplastica után.

Pulmonary fibrosis and impeding gangrene as complications of radiotherapy and chemotherapy developed in a patient who previously underwent right upper lobectomy. Following completion pneumonectomy, bronchopleural fistula and consecutive thoracic empyema formed. This case presentation is to demonstrate the vicious circle of severe complications following oncological treatment. The role of vacuum assisted closure and other management options are discussed which resulted in full recovery of the patient after three years finally.