Radiofrequency Electromagnetic Fields Cause Non-Temperature-Induced Physical and Biological Effects in Cancer Cells.

Non-temperature-induced effects of radiofrequency electromagnetic fields (RF) have been controversial for decades. Here, we established measurement techniques to prove their existence by investigating energy deposition in tumor cells under RF exposure and upon adding amplitude modulation (AM) (AMRF). Using a preclinical device LabEHY-200 with a novel in vitro applicator, we analyzed the power deposition and system parameters for five human colorectal cancer cell lines and measured the apoptosis rates in vitro and tumor growth inhibition in vivo in comparison to water bath heating. We showed enhanced anticancer effects of RF and AMRF in vitro and in vivo and verified the non-temperature-induced origin of the effects. Furthermore, apoptotic enhancement by AM was correlated with cell membrane stiffness. Our findings not only provide a strategy to significantly enhance non-temperature-induced anticancer cell effects in vitro and in vivo but also provide a perspective for a potentially more effective tumor therapy.

Critical-size Defect Augmentation Using Sintered and Non-Sintered Bovine Bone Matrix - An Experimental Controlled Study in Minipigs.

PURPOSE: Xenogeneic bone substitute materials are often used for augmentation of larger bone defects. Purification methods for these materials vary, mainly in terms of temperature. The aim of this study was to determine in vivo how sintering affects quantitative and qualitative bone regeneration of 2 bovine augmentation materials. METHODS: A total of 56 critical size defects were set at the frontal bone of 14 domestic pigs (4 each) and filled randomly with either bovine, sintered hydroxyapatite (BO), bovine, non-sintered hydroxyapatite (BOS), local autologous bone (AB) or left empty. All defects were additionally covered with a collagen membrane. Specimens were harvested after 4 and 8 weeks and were evaluated histologically and histomorphometrically. RESULTS: Histologically new bone could be seen in every group. Significantly highest new bone formation was found in AB. No significant difference could be detected between BO and BOS. CONCLUSIONS: According to the results of this study, sintered bone substitute material remains histologically distinguishable but does not affect quantitative and qualitative bone regeneration.

Experimental Studies for Small Diameter Grafts in an In Vivo Sheep Model-Techniques and Pitfalls.

BACKGROUND: Scientific attempts to create the "ideal" small diameter vascular graft have been compared with the "search of the holy grail." Prosthetic material as expanded polytetrafluoroethylene or Dacron shows acceptable patency rates to large caliber vessels, while small diameter (< 6 mm) prosthetic conduits present unacceptably poor patency rates. Vascular tissue engineering represents a promising option to address this problem. MATERIAL AND METHODS: Thirty-two female Texel-sheep aged 6 months to 2 years underwent surgical common carotid artery (CCA) interposition using different tissue-engineered vascular substitutes. Explantation of the grafts was performed 12 (n = 12) and 36 (n = 20) weeks after surgery. Ultrasound was performed on postoperative day 1 and thereafter every 4 weeks to evaluate the graft patency. RESULTS: The average length of implanted substitutes was 10.3 ± 2.2 cm. Anesthesia and surgical procedure could be performed without major surgical complications in all cases.The grafts showed a systolic blood flow velocity (BFV) of 28.24 ± 13.5 cm/s, a diastolic BFV of 9.25 ± 4.53 cm/s, and a mean BFV of 17.85 ± 9.25 cm/s. Native vessels did not differ relevantly in hemodynamic measurements (systolic: 29.77 cm/s; diastolic: 7.99 cm/s ± 5.35; mean 15.87 ± 10.75). There was no incidence of neurologic complications or subsequent postoperative occlusion. Perioperative morbidity was low and implantation of conduits was generally well tolerated. CONCLUSION: This article aims to give a precise overview of in vivo experiments in sheep for the evaluation of small diameter vascular grafts performing CCA interposition, especially with regard to pitfalls and possible perioperative complications and to discuss advantages and disadvantages of this approach.

In-vitro and in-vivo investigations into the carbene-gold anticancer drug candidates NHC*-Au-SCSNMe2 and NHC*-Au-S-GLUC against advanced prostate cancer PC3.

The anticancer drug candidates 1,3-dibenzyl-4,5-diphenyl-imidazol-2-ylidene gold(I) dimethylamino dithiocarbamate and 2,3,4,6-tetra-O-acetyl-α-D-glucopyranosyl-1-thiolate derivative exhibited nanomolar in-vitro activity against prostate cancer cells advanced prostate cancer (PC3) and micromolar inhibition of mammalian thioredoxin reductase. Encouraging maximum tolerable dose experiments led to human prostate cancer subcutaneous xenograft experiments; 1,3-dibenzyl-4,5-diphenyl-imidazol-2-ylidene gold(I) dimethylamino dithiocarbamate and 2,3,4,6-tetra-O-acetyl-α-D-glucopyranosyl-1-thiolate derivative were applied twelve times at two doses in groups of n = 5 PC3 to tumor-bearing NMRI:nu/nu mice. 1,3-dibenzyl-4,5-diphenyl-imidazol-2-ylidene gold(I) dimethylamino dithiocarbamate and 2,3,4,6-tetra-O-acetyl-α-D-glucopyranosyl-1-thiolate derivative at the dose of 10 and 20 mg/kg showed good tolerability, while no significant body weight loss was seen in both groups. In particular, for the drug 1,3-dibenzyl-4,5-diphenyl-imidazol-2-ylidene gold(I) dimethylamino dithiocarbamate the tumor growth inhibition suggested to be dose dependent, reflected by the respective optimal T/C values of 0.45 at the dose of 10 mg/kg and of 0.31 at the dose of 20 mg/kg. By contrast, the 2,3,4,6-tetra-O-acetyl-α-D-glucopyranosyl-1-thiolate derivative treated groups showed no indication for dose-dependent antitumoral activity, as reflected by the optimal T/C values of 0.44 for the 10 mg/kg and for the 20 mg/kg treated mice. Immunohistochemical experiments involving Ki67 staining of tumor tissue showed that both compounds reduced PC3 cell proliferation against the difficult to treat advanced human prostate tumors derived from PC3.

A new device to prevent fascial retraction in the open abdomen - proof of concept in vivo.

BACKGROUND: An open abdomen is often necessary for survival of patients after peritonitis, compartment syndrome, or in damage control surgery. However, abdominal wall retraction relieves delays and complicates abdominal wall closure. The principle of the newly fascia preserving device (FPD) is the application of anteriorly directed traction on both fascial edges over an external support through a longitudinal beam to relieve increased abdominal pressure and prevent fascial retraction. METHODS: Twelve pigs were randomly divided into two groups. Both groups underwent midline laparotomy under general anesthesia. Group one was treated with the new device, group two served as controls. The tension for closing the abdominal fascia was measured immediately after laparotomy as well as at 24 and 48 h. Vital parameters and ventilation pressure were recorded. Post mortem, all fascial tissues were histologically examined. RESULTS: All pigs demonstrated increases in abdominal circumference. In both groups, forces for closing the abdomen increased over the observation period. Concerning the central closing force after 24 h we saw a significant lower force in the FPD group (14.4 ± 3 N) vs. control group (21.6 ± 5.7 N, p < 0.001). By testing the main effects using an ANOVA analysis we found a significant group related effect concerning closing force and abdominal circumference of the FDP-group vs. control group (p < 0.001; p < 0.001). The placement of the device on chest and pelvis did not influence vital parameters and ventilation pressure. Histologic exam detected no tissue damage. CONCLUSIONS: This trial shows the feasibility to prevent fascial retraction during the open abdomen by using the new device. Thus, it is expected that an earlier closure of the abdominal wall will be possible, and a higher rate of primary closure will be attained.

Stereological assessment of the blood-air barrier and the surfactant system after mesenchymal stem cell pretreatment in a porcine non-heart-beating donor model for lung transplantation.

More frequent utilization of non-heart-beating donor (NHBD) organs for lung transplantation has the potential to relieve the shortage of donor organs. In particular with respect to uncontrolled NHBD, concerns exist regarding the risk of ischaemia/reperfusion (IR) injury-related graft damage or dysfunction. Due to their immunomodulating and tissue-remodelling properties, bone-marrow-derived mesenchymal stem cells (MSCs) have been suspected of playing a beneficial role regarding short- and long-term survival and function of the allograft. Thus, MSC administration might represent a promising pretreatment strategy for NHBD organs. To study the initial effects of warm ischaemia and MSC application, a large animal lung transplantation model was generated, and the structural organ composition of the transplanted lungs was analysed stereologically with particular respect to the blood-gas barrier and the surfactant system. In this study, porcine lungs (n = 5/group) were analysed. Group 1 was the sham-operated control group. In pigs of groups 2-4, cardiac arrest was induced, followed by a period of 3 h of ventilated ischaemia at room temperature. In groups 3 and 4, 50 × 106 MSCs were administered intravascularly via the pulmonary artery and endobronchially, respectively, during the last 10 min of ischaemia. The left lungs were transplanted, followed by a reperfusion period of 4 h. Then, lungs were perfusion-fixed and processed for light and electron microscopy. Samples were analysed stereologically for IR injury-related structural parameters, including volume densities and absolute volumes of parenchyma components, alveolar septum components, intra-alveolar oedema, and the intracellular and intra-alveolar surfactant pool. Additionally, the volume-weighted mean volume of lamellar bodies (lbs) and their profile size distribution were determined. Three hours of ventilated warm ischaemia was tolerated without eliciting histological or ultrastructural signs of IR injury, as revealed by qualitative and quantitative assessment. However, warm ischaemia influenced the surfactant system. The volume-weighted mean volume of lbs was reduced significantly (P = 0.024) in groups subjected to ischaemia (group medians of groups 2-4: 0.180-0.373 µm³) compared with the sham control group (median 0.814 µm³). This was due to a lower number of large lb profiles (size classes 5-15). In contrast, the intra-alveolar surfactant system was not altered significantly. No significant differences were encountered comparing ischaemia alone (group 2) or ischaemia plus application of MSCs (groups 3 and 4) in this short-term model.

Patency and in vivo compatibility of bacterial nanocellulose grafts as small-diameter vascular substitute.

OBJECTIVE: Despite the clinical success of large-diameter vascular grafts, synthetic grafts in small-diameter vessels are of limited use because of their poor patency rates. Previous experiments of our group provided evidence for good biocompatibility of bacterial nanocellulose (BNC) as a small-vessel graft in the carotid artery in sheep. However, the patency rate of our first-generation tubes after 3 months was only 50%. To advance our concept, we now used modified second-generation tubes with diminished wall thickness and a smoother inner surface to reduce the thrombogenic potential. The aim was to investigate mechanical characteristics of modified second-generation BNC tubes, to evaluate in vivo performance and biocompatibility, and to analyze patency rates. METHODS: We replaced the right carotid artery of 23 sheep with second-generation BNC tubes. Compared with our first-generation tubes, tubes were modified with different surface properties and diminished wall thickness (inner diameter, 4.0-5.0 mm; wall thickness, 1.0-2.5 mm; length, 100 mm) to generate a smoother inner surface with reduced thrombogenic potential and a more porous outer zone, allowing easier cell immigration. RESULTS: At the end of the investigational period, BNC tubes were explanted and grafts were processed for histopathologic analysis. Histologic analysis revealed no acute signs of foreign body reaction such as immigration of giant cells or other acute inflammatory reaction and therefore provided evidence for good biocompatibility of the second-generation tubes. However, all grafts of the sheep without antiplatelet therapy were occluded after 9 months, whereas grafts in sheep receiving dual platelet inhibition showed a patency rate of 67% (six of nine grafts). Further modified grafts revealed a patency rate of 80% (four of five grafts remained open). CONCLUSIONS: Patency rates of the second-generation tubes could be substantially improved compared with our first-generation tubes. However, poor patency rates of tissue-engineered blood vessels still limit their use in clinical studies. Further efforts in terms of in vitro and in vivo studies are essential to improve grafts of BNC.

Role of LTBP4 in alveolarization, angiogenesis, and fibrosis in lungs.

Deficiency of the extracellular matrix protein latent transforming growth factor-ß (TGF-ß)-binding protein-4 (LTBP4) results in lack of intact elastic fibers, which leads to disturbed pulmonary development and lack of normal alveolarization in humans and mice. Formation of alveoli and alveolar septation in pulmonary development requires the concerted interaction of extracellular matrix proteins, growth factors such as TGF-ß, fibroblasts, and myofibroblasts to promote elastogenesis as well as vascular formation in the alveolar septae. To investigate the role of LTBP4 in this context, lungs of LTBP4-deficient (Ltbp4-/-) mice were analyzed in close detail. We elucidate the role of LTBP4 in pulmonary alveolarization and show that three different, interacting mechanisms might contribute to alveolar septation defects in Ltbp4-/- lungs: 1) absence of an intact elastic fiber network, 2) reduced angiogenesis, and 3) upregulation of TGF-ß activity resulting in profibrotic processes in the lung.

Ltbp4 regulates Pdgfrß expression via TGFß-dependent modulation of Nrf2 transcription factor function.

Latent transforming growth factor beta binding protein 4 (LTBP4) belongs to the fibrillin/LTBP family of proteins and plays an important role as a structural component of extracellular matrix (ECM) and local regulator of TGFß signaling. We have previously reported that Ltbp4S knock out mice (Ltbp4S-/-) develop centrilobular emphysema reminiscent of late stage COPD, which could be partially rescued by inactivating the antioxidant protein Sestrin 2 (Sesn2). More recent studies showed that Sesn2 knock out mice upregulate Pdgfrß-controlled alveolar maintenance programs that protect against cigarette smoke induced pulmonary emphysema. Based on this, we hypothesized that the emphysema of Ltbp4S-/- mice is primarily caused by defective Pdgfrß signaling. Here we show that LTBP4 induces Pdgfrß signaling by inhibiting the antioxidant Nrf2/Keap1 pathway in a TGFß-dependent manner. Overall, our data identified Ltbp4 as a major player in lung remodeling and injury repair.

Morphological characterization and immunohistochemical detection of the proinflammatory cytokines IL-1ß, IL-17A, and TNF-α in lung lesions associated with contagious bovine pleuropneumonia.

Contagious bovine pleuropneumonia (CBPP), a severe respiratory disease, is characterized by massive inflammation of the lung especially during the acute clinical stage of infection. Tissue samples from cattle, experimentally infected with Mycoplasma mycoides subsp. mycoides Afadé, were subjected to histopathological and immunohistochemical examination in order to provide insight into innate immune pathways that shape inflammatory host responses. Lung lesions were characterized by vasculitis, necrosis, and increased presence of macrophages and neutrophils, relative to uninfected animals. The presence of three cytokines associated with innate inflammatory immune responses, namely, IL-1ß, IL-17A, and TNF-α, were qualitatively investigated in situ. Higher cytokine levels were detected in lung tissue samples from CBPP-affected cattle compared to samples derived from an uninfected control group. We therefore conclude that the cytokines TNF-α and IL-1ß, which are prevalent in the acute phase of infections, play a role in the inflammatory response seen in the lung tissue in CBPP. IL-17A gets released by activated macrophages and attracts granulocytes that modulate the acute phase of the CBPP lesions.

Prenatal exposure to the phytoestrogen daidzein resulted in persistent changes in ovarian surface epithelial cell height, folliculogenesis, and estrus phase length in adult Sprague-Dawley rat offspring.

Daidzein (DZ), an isoflavone with the potential to interfere with estrogen signaling, is found in soy products, which have gained popularity due to purported beneficial effects on the cardiovascular and skeletal systems and potential antineoplastic properties. However, the ingestion of phytoestrogens has been associated with impaired reproductive function in many species. The aim of this study was to determine the long-term effects on the ovaries of rat offspring exposed to DZ or ethinyl estradiol (EE) during prenatal development. Gravid rats were administered either vehicle or 5 or 60 mg DZ/kg body weight/d or 0.002 mg 17-α EE /kg body weight/d on gestational days 6-21. Ovarian-related endpoints were investigated during adulthood in female offspring. The mean cell height of the ovarian surface epithelium was significantly reduced in all treated groups. Alterations in folliculogenesis included increased follicular atresia, a reduction in secondary and tertiary follicle numbers, and cyst formation. An elevated prevalence of a slightly prolonged estrus phase was also observed. The morphological changes to the ovarian surface epithelium are consistent with an antiproliferative effect, while ovarian folliculogenesis was adversely affected. The effects of the high dose DZ were similar to those observed with 17-α EE.

Class IA Phosphatidylinositol 3-Kinase Isoform p110α Mediates Vascular Remodeling.

OBJECTIVE: Neointima formation after vascular injury remains a significant problem in clinical cardiology, and current preventive strategies are suboptimal. Phosphatidylinositol 3'-kinase is a central downstream mediator of growth factor signaling, but the role of phosphatidylinositol 3'-kinase isoforms in vascular remodeling remains elusive. We sought to systematically characterize the precise role of catalytic class IA phosphatidylinositol 3'-kinase isoforms (p110α, p110ß, p110δ), which signal downstream of receptor tyrosine kinases, for vascular remodeling in vivo. APPROACH AND RESULTS: Western blot analyses revealed that all 3 isoforms are abundantly expressed in smooth muscle cells. To analyze their significance for receptor tyrosine kinases-dependent cellular responses, we used targeted gene knockdown and isoform-specific small molecule inhibitors of p110α (PIK-75), p110ß (TGX-221), and p110δ (IC-87114), respectively. We identified p110α to be crucial for receptor tyrosine kinases signaling, thus affecting proliferation, migration, and survival of rat, murine, and human smooth muscle cells, whereas p110ß and p110δ activities were dispensable. Surprisingly, p110δ exerted noncatalytic functions in smooth muscle cell proliferation, but had no effect on migration. Based on these results, we generated a mouse model of smooth muscle cell-specific p110α deficiency (sm-p110α(-/-)). Targeted deletion of p110α in sm-p110α(-/-) mice blunted growth factor-induced cellular responses and abolished neointima formation after balloon injury of the carotid artery in mice. In contrast, p110δ deficiency did not affect vascular remodeling in vivo. CONCLUSIONS: Receptor tyrosine kinases-induced phosphatidylinositol 3'-kinase signaling via the p110α isoform plays a central role for vascular remodeling in vivo. Thus, p110α represents a selective target for the prevention of neointima formation after vascular injury, whereas p110ß and p110δ expression and activity do not play a significant role.

Controlled lung reperfusion to reduce pulmonary ischaemia/reperfusion injury after cardiopulmonary bypass in a porcine model.

OBJECTIVES: Ischaemia/reperfusion (I/R) injury of the lungs contributes to pulmonary dysfunction after cardiac surgery with cardiopulmonary bypass (CPB), leading to increased morbidity and mortality of patients. This study investigated the value of controlled lung reperfusion strategies on lung ischaemia-reperfusion injury in a porcine CPB model. METHODS: Pigs were subjected to routine CPB for 120 min with 60 min of blood cardioplegic cardiac arrest (CCA). Following CCA, the uncontrolled reperfusion (UR, n = 6) group was conventionally weaned from CPB. Two groups underwent controlled lung reperfusion strategies (CR group: controlled reperfusion conditions, n = 6; MR group: controlled reperfusion conditions and modified reperfusate, n = 6) via the pulmonary artery before CPB weaning. Sham-operated pigs (n = 7) served as controls. Animals were followed up until 4 h after CPB. Pulmonary function, haemodynamics, markers of inflammation, endothelial injury and oxidative stress as well as morphological lung alterations were analysed. RESULTS: CPB (UR group) induced deterioration of pulmonary function (lung mechanics, oxygenation index and lung oedema). Also, controlled lung reperfusion groups (CR and MR) presented with pulmonary dysfunction after CPB. However, compared with UR, controlled lung reperfusion strategies (CR and MR) improved lung mechanics and reduced markers of oxidative stress, but without alteration of haemodynamics, oxygenation, inflammation, endothelial injury and lung morphology. Both controlled reperfusion groups were similar without relevant differences. CONCLUSION: Controlled lung reperfusion strategies attenuated a decrease in lung mechanics and an increase in oxidative stress, indicating an influence on CPB-related pulmonary injury. However, they failed to avoid completely CPB-related lung injury, implying the need for additional strategies given the multifactorial pathophysiology of postoperative pulmonary dysfunction.

BID-dependent release of mitochondrial SMAC dampens XIAP-mediated immunity against Shigella.

The X-linked inhibitor of apoptosis protein (XIAP) is a potent caspase inhibitor, best known for its anti-apoptotic function in cancer. During apoptosis, XIAP is antagonized by SMAC, which is released from the mitochondria upon caspase-mediated activation of BID. Recent studies suggest that XIAP is involved in immune signaling. Here, we explore XIAP as an important mediator of an immune response against the enteroinvasive bacterium Shigella flexneri, both in vitro and in vivo. Our data demonstrate for the first time that Shigella evades the XIAP-mediated immune response by inducing the BID-dependent release of SMAC from the mitochondria. Unlike apoptotic stimuli, Shigella activates the calpain-dependent cleavage of BID to trigger the release of SMAC, which antagonizes the inflammatory action of XIAP without inducing apoptosis. Our results demonstrate how the cellular death machinery can be subverted by an invasive pathogen to ensure bacterial colonization.

In vivo application of tissue-engineered blood vessels of bacterial cellulose as small arterial substitutes: proof of concept?

BACKGROUND: Tissue-engineered blood vessels (TEBVs) represent an innovative approach for overcoming reconstructive problems associated with vascular diseases by providing small-caliber vascular grafts. This study aimed to evaluate a novel biomaterial of bacterially synthesized cellulose (BC) as a potential scaffold for small-diameter TEBV. METHODS: Small-diameter blood vessels with a supramolecular fiber network structure consisting of tubular hydrogels from biodesigned cellulose were created using Gluconacetobacter strains and Matrix reservoir technology. BC tubes (length: 100 mm, inner diameter: 4.0-5.0 mm) were applied to replace the carotid arteries of 10 sheep for a period of 3 mo to gain further insights into (a) functional (in vivo) performance, (b) ability of providing a scaffold for the neoformation of a vascular wall and (c) their proinflammatory potential, and the (d) technical feasibility of the procedure. RESULTS: Preoperative analysis revealed a bursting strength of the grafts of approximately 800 mm Hg and suture retention strength of 4-5 N. Postexplantation analysis showed a patency rate of 50% (n = 5) and physiological performance of the patent grafts at 4, 8, and 12 wk postoperatively, compared with native arteries. Histologic analysis revealed a neoformation of a vascular wall-like structure along the BC scaffold consisting of immigrated vascular smooth muscle cells and a homogeneous endothelialization of the inner graft surface without signs of prothrombogenic or inflammatory potential. Scanning electron microscopy revealed a confluent luminal endothelial cell layer and the immigration of vascular smooth muscle cells into the BC matrix. CONCLUSIONS: BC grafts provide a scaffold for the neoformation of a three-layered vascular wall exhibit attractive properties for their use in future TEBV programs for cardiovascular surgery.

Latent transforming growth factor ß-binding protein 4 is downregulated in esophageal cancer via promoter methylation.

Latent transforming growth factor ß-binding protein 4 (LTBP4) is an extracellular matrix molecule that is a member of important connective tissue networks and is needed for the correct folding and the secretion of TGF-ß1. LTBP4 is downregulated in carcinomas of various tissues. Here we show that LTBP4 is also downregulated in adenocarcinomas and squamous cell carcinomas of the esophagus in vitro and in vivo. Re-expression of LTBP4 in esophageal cancer cell lines reduced cell migration ability, whereas cell viability and cell proliferation remained unchanged. Hypermethylation of the promoter regions of the two main human LTBP4 transcriptional forms, LTBP4L and LTBP4S, was found to be involved in LTBP4 silencing. Detailed investigations of the methylation patterns of the promoter regions of LTBP4L and LTBP4S identified GATA1, SP1, E2F4 and SMAD3 as potential transcription factors involved in LTBP4 expression. In in vitro transcription factor activity studies we discovered E2F4 as novel powerful regulator for LTBP4S expression.

Rosuvastatin reloading before cardiac surgery with cardiopulmonary bypass.

BACKGROUND/PURPOSE: Recent evidence suggests that statin-mediated cardioprotection after chronic statin therapy decreases over time and can be reactivated by preprocedural high-dose statin reloading therapy. We tested in a porcine cardiopulmonary bypass (CPB) model whether statin-related cardioprotection is further enhanced by a preoperative rosuvastatin reloading therapy. METHODS: Control (n = 6), rosuvastatin-pretreated (n = 6; 20 mg/day for 7 days p.o.) and rosuvastatin-reloaded (n = 6; p.o. treatment plus 0.10 mg/kg/h i.v. during surgery) pigs (Deutsche Landrasse) were subjected to CPB for 2 h with 1 h of cardioplegic cardiac arrest. Systemic hemodynamics, cardiac index (CI), coronary blood flow (CBF) and left ventricular (LV) function [pressure-volume area (PVA), preload recruitable stroke work (PRSW)] were determined before and 4 h after CPB. Myocardial expression (PCR) and protein content (Western blot) of endothelial NO synthase (eNOS) and phosphatase and tensin homolog deleted on chromosome ten (PTEN) were measured, and right coronary relaxation was assessed postmortem. All data are given as mean ± SD. RESULTS: Preoperative plasma LDL, HDL and cholesterol did not differ between treatment groups. Compared to control, oral treatment improved post-CPB CI, CBF, first derivative of maximal LV-pressure (LVdp/dt) and PVA (p < 0.05). Significant enhancement was achieved with perioperative reloading therapy (CI: 5.2 ± 1.0 vs. 3.9 ± 1.5 l/min/m(2); CBF: 76 ± 32 vs. 43 ± 8 ml/min; LVdp/dt: 1,980 ± 333 vs. 1,249 ± 461 mm Hg/s; PVA: 6,954 ± 941 vs. 3,252 ± 1,822 mm Hg·ml; p < 0.05) with improved in vitro NO-dependent coronary relaxation (102 ± 10 vs. 79 ± 14%; p = 0.003). Irrespective of recapture therapy statin pretreatment augmented myocardial eNOS and PTEN (p < 0.05), but failed to increase cardiac eNOS or PTEN expression after CPB. CONCLUSIONS: Periprocedural statin reloading therapy enhances myocardial and coronary function after cardiac surgery with CPB and may therefore provide a valuable therapeutic approach for the reduction of myocardial ischemia-reperfusion injury.

Severe SMA mice show organ impairment that cannot be rescued by therapy with the HDACi JNJ-26481585.

Spinal muscular atrophy (SMA) is the leading genetic cause of early childhood death worldwide and no therapy is available today. Many drugs, especially histone deacetylase inhibitors (HDACi), increase SMN levels. As all HDACi tested so far only mildly ameliorate the SMA phenotype or are unsuitable for use in humans, there is still need to identify more potent drugs. Here, we assessed the therapeutic power of the pan-HDACi JNJ-26481585 for SMA, which is currently used in various clinical cancer trials. When administered for 64 h at 100 nM, JNJ-26481585 upregulated SMN levels in SMA fibroblast cell lines, including those from non-responders to valproic acid. Oral treatment of Taiwanese SMA mice and control littermates starting at P0 showed no overt extension of lifespan, despite mild improvements in motor abilities and weight progression. Many treated and untreated animals showed a very rapid decline or unexpected sudden death. We performed exploratory autopsy and histological assessment at different disease stages and found consistent abnormalities in the intestine, heart and lung and skeletal muscle vasculature of SMA animals, which were not prevented by JNJ-26481585 treatment. Interestingly, some of these features may be only indirectly caused by α-motoneuron function loss but may be major life-limiting factors in the course of disease. A better understanding of - primary or secondary - non-neuromuscular organ involvement in SMA patients may improve standard of care and may lead to reassessment of how to investigate SMA patients clinically.

Evaluation of the use of lower body perfusion at 28°C in aortic arch surgery.

OBJECTIVES: Although hypothermic circulatory arrest (HCA) and selective cerebral perfusion (SCP) are widely used for cerebral protection during aortic arch surgery, these strategies offer no protection for mesenteric ischaemia during prolonged circulatory arrest. This study explored mesenteric haemodynamics, metabolism, oxidative stress and inflammatory response levels during isolated SCP and combined cerebral and lower body perfusion (CLBP) in pigs. METHODS: Fourteen pigs (35-45 kg) were cooled on CPB to 28°C. After 10 min of HCA, they were randomized to 60 min of isolated SCP (n = 7) and CLBP (n = 7) at low-flow pump rates: 10 ml/kg/min (SCP) and 20 ml/kg/min (LBP). Microspheres were injected at baseline, 5 and 60 min of SCP/CLBP and 5 and 60 min off CPB, to calculate mesenteric regional blood flow (RBF). Lactate levels and Oxy-DNA expression [fluorescence activated cell sorting (FACS)] in the portal venous blood were determined at the same time points. Semi-quantitative assessment of inflammatory cytokines was performed using real-time polymerase chain reaction (PCR) and immunhistochemical analyses. RESULTS: At baseline mesenteric, RBF was 61 ± 31 ml/min/100 g in the jejunum and 78 ± 43 ml/min/100 g in the colon. Whereas SCP provided a residual mesenteric RBF of 5%, CLBP offered 47% of the baseline jejunal (34 ± 10 ml/min/100 g) and 68% of the colonic RBF (52 ± 34 ml/min/100 g; P = 0.001). Lactate levels were significantly higher in then SCP group (15 ± 2 vs. 11 ± 3 mmol/l; P = 0.01). Oxy-DNA increased, reaching 137% of baseline (SCP) and 129% (CLBP) at 60 min SCP/CLBP, but recovered promptly during reperfusion. Real-time PCR revealed a massive increase in early cytokine expression vs. baseline, showing significant higher interleukin (IL) -6 (29 vs.2; P = 0.027) and COX-relative expression (7 vs. 3, P = 0.016) in the SCP group. Immunhistochemical analysis confirmed a higher immunological activity in the SCP group, showing more intensive signal for tumour necrosis factor-α, IL-6 and p38 when compared with the CLBP group. CONCLUSIONS: Low-flow CLBP provides a diminished but considerable mesenteric RBF, leading to lower lactate and oxidative stress levels and a diminished local inflammatory response reaction than isolated SCP.