Merlin, a "Magic" Linker between Extracellular Cues and Intracellular Signaling Pathways that Regulate Cell Motility, Proliferation, and Survival.

Genetic alterations of neurofibromatosis type 2 (NF2) gene lead to the development of schwannomas, meningiomas, and ependymomas. Mutations of NF2 gene were also found in thyroid cancer, mesothelioma, and melanoma, suggesting that it functions as a tumor suppressor in a wide spectrum of cells. The product of NF2 gene is merlin (moesin-ezrin-radixin-like protein), a member of the Band 4.1 superfamily proteins. Merlin shares significant sequence homology with the ERM (Ezrin-Radixin-Moesin) family proteins and serves as a linker between transmembrane proteins and the actin-cytoskeleton. Merlin is a multifunctional protein and involved in integrating and regulating the extracellular cues and intracellular signaling pathways that control cell fate, shape, proliferation, survival, and motility. Recent studies showed that merlin regulates the cell-cell and cell-matrix adhesions and functions of the cell surface adhesion/extracellular matrix receptors including CD44 and that merlin and CD44 antagonize each other's function and work upstream of the mammalian Hippo signaling pathway. Furthermore, merlin plays important roles in stabilizing the contact inhibition of proliferation and in regulating activities of several receptor tyrosine kinases. Accumulating data also suggested an emerging role of merlin as a negative regulator of growth and progression of several non-NF2 associated cancer types. Together, these recent advances have improved our basic understanding about merlin function, its regulation, and the major signaling pathways regulated by merlin and provided the foundation for future translation of these findings into the clinic for patients bearing the cancers in which merlin function and/or its downstream signaling pathways are impaired or altered.

Angiofil-mediated visualization of the vascular system by microcomputed tomography: a feasibility study.

Visualization of the vascular systems of organs or of small animals is important for an assessment of basic physiological conditions, especially in studies that involve genetically manipulated mice. For a detailed morphological analysis of the vascular tree, it is necessary to demonstrate the system in its entirety. In this study, we present a new lipophilic contrast agent, Angiofil, for performing postmortem microangiography by using microcomputed tomography. The new contrast agent was tested in 10 wild-type mice. Imaging of the vascular system revealed vessels down to the caliber of capillaries, and the digital three-dimensional data obtained from the scans allowed for virtual cutting, amplification, and scaling without destroying the sample. By use of computer software, parameters such as vessel length and caliber could be quantified and remapped by color coding onto the surface of the vascular system. The liquid Angiofil is easy to handle and highly radio-opaque. Because of its lipophilic abilities, it is retained intravascularly, hence it facilitates virtual vessel segmentation, and yields an enduring signal which is advantageous during repetitive investigations, or if samples need to be transported from the site of preparation to the place of actual analysis, respectively. These characteristics make Angiofil a promising novel contrast agent; when combined with microcomputed tomography, it has the potential to turn into a powerful method for rapid vascular phenotyping.

Pulmonary and systemic vascular dysfunction in young offspring of mothers with preeclampsia.

BACKGROUND: Adverse events in utero may predispose to cardiovascular disease in adulthood. The underlying mechanisms are unknown. During preeclampsia, vasculotoxic factors are released into the maternal circulation by the diseased placenta. We speculated that these factors pass the placental barrier and leave a defect in the circulation of the offspring that predisposes to a pathological response later in life. The hypoxia associated with high-altitude exposure is expected to facilitate the detection of this problem. METHODS AND RESULTS: We assessed pulmonary artery pressure (by Doppler echocardiography) and flow-mediated dilation of the brachial artery in 48 offspring of women with preeclampsia and 90 offspring of women with normal pregnancies born and permanently living at the same high-altitude location (3600 m). Pulmonary artery pressure was roughly 30% higher (mean+/-SD, 32.1+/-5.6 versus 25.3+/-4.7 mm Hg; P<0.001) and flow-mediated dilation was 30% smaller (6.3+/-1.2% versus 8.3+/-1.4%; P<0.0001) in offspring of mothers with preeclampsia than in control subjects. A strong inverse relationship existed between flow-mediated dilation and pulmonary artery pressure (r=-0.61, P<0.001). The vascular dysfunction was related to preeclampsia itself because siblings of offspring of mothers with preeclampsia who were born after a normal pregnancy had normal vascular function. Augmented oxidative stress may represent an underlying mechanism because thiobarbituric acid-reactive substances plasma concentration was increased in offspring of mothers with preeclampsia. CONCLUSIONS: Preeclampsia leaves a persistent defect in the systemic and the pulmonary circulation of the offspring. This defect predisposes to exaggerated hypoxic pulmonary hypertension already during childhood and may contribute to premature cardiovascular disease in the systemic circulation later in life.

Diabetes mellitus and vascular lesions.

Cardiovascular complications represent by far the most severe manifestations of diabetes mellitus. Treatment aimed at stopping progression of vascular lesions may fall short if initiated when the disease becomes clinically evident. Therefore, identification of the earliest vascular disfunctions may offer the best opportunity to interfere with pathogenic mechanisms and avoid progression of diabetic vasculopathy. In this report, we present a few mechanisms that alter hemodynamic and metabolic homeostasis in the course of diabetes mellitus. Endothelial function with special emphasis on nitric oxide and oxidative stress, advanced glycation end products, and the renin angiotensin system are briefly discussed. New pharmacological agents that may favorably influence these parameters are presently undergoing clinical trials. However, tight control of plasma glucose and cardiovascular risk factors represent the cornerstone of the treatment in diabetes to slow progression of vascular disease.

Neuronal nitric oxide synthase does not contribute to the modulation of pulmonary vascular tone in fetal lambs with congenital diaphragmatic hernia (nNOS in CDH lambs).

AIM: The aim of this study was to determine the presence of the neuronal nitric oxide synthase (nNOS) in near full-term lambs with congenital diaphragmatic hernia (CDH) and its role in the modulation of pulmonary vascular basal tone. METHODS: We surgically created diaphragmatic hernia on the 85th day of gestation. On the 135th, catheters were used to measure pulmonary pressure and blood flow. We tested the effects of 7-nitroindazole (7-NINA), a specific nNOS antagonist and of N-nitro-L-arginine (L-NNA), a nonspecific nitric oxide synthase antagonist. In vitro, we tested the effects of the same drugs on isolated pulmonary vessels. The presence of nNOS protein in the lungs was detected by Western blot analysis. RESULTS: Neither 7-NINA nor L-NNA modified pulmonary vascular basal tone in vivo. After L-NNA injection, acetylcholine (ACh) did not decrease significantly pulmonary vascular resistance (PVR). In vitro, L-NNA increased the cholinergic contractile-response elicited by electric field stimulation (EFS) of vascular rings from lambs with diaphragmatic hernia. CONCLUSION: We conclude that nNOS protein is present in the lungs and pulmonary artery of near full-term lamb fetuses with diaphragmatic hernia, but that it does not contribute to the reduction of pulmonary vascular tone at birth

Postmortem pump-driven reperfusion of the vascular system of porcine lungs: towards a new model for surgical training.

PURPOSE: The objective of this experiment is to establish a continuous postmortem circulation in the vascular system of porcine lungs and to evaluate the pulmonary distribution of the perfusate. This research is performed in the bigger scope of a revascularization project of Thiel embalmed specimens. This technique enables teaching anatomy, practicing surgical procedures and doing research under lifelike circumstances. METHODS: After cannulation of the pulmonary trunk and the left atrium, the vascular system was flushed with paraffinum perliquidum (PP) through a heart-lung machine. A continuous circulation was then established using red PP, during which perfusion parameters were measured. The distribution of contrast-containing PP in the pulmonary circulation was visualized on computed tomography. Finally, the amount of leak from the vascular system was calculated. RESULTS: A reperfusion of the vascular system was initiated for 37 min. The flow rate ranged between 80 and 130 ml/min throughout the experiment with acceptable perfusion pressures (range: 37-78 mm Hg). Computed tomography imaging and 3D reconstruction revealed a diffuse vascular distribution of PP and a decreasing vascularization ratio in cranial direction. A self-limiting leak (i.e. 66.8% of the circulating volume) towards the tracheobronchial tree due to vessel rupture was also measured. CONCLUSIONS: PP enables circulation in an isolated porcine lung model with an acceptable pressure-flow relationship resulting in an excellent recruitment of the vascular system. Despite these promising results, rupture of vessel walls may cause leaks. Further exploration of the perfusion capacities of PP in other organs is necessary. Eventually, this could lead to the development of reperfused Thiel embalmed human bodies, which have several applications.

Maintenance therapy with thalidomide improves survival in patients with multiple myeloma.

Newer chemotherapeutic protocols as well as high-dose chemotherapy have increased the response rate in myeloma. However, these treatments are not curative. Effective maintenance strategies are now required to prolong the duration of response. We conducted a randomized trial of maintenance treatment with thalidomide and pamidronate. Two months after high-dose therapy, 597 patients younger than age 65 years were randomly assigned to receive no maintenance (arm A), pamidronate (arm B), or pamidronate plus thalidomide (arm C). A complete or very good partial response was achieved by 55% of patients in arm A, 57% in arm B, and 67% in arm C (P = .03). The 3-year postrandomization probability of event-free survival was 36% in arm A, 37% in arm B, and 52% in arm C (P < .009). The 4-year postdiagnosis probability of survival was 77% in arm A, 74% in arm B, and 87% in arm C (P < .04). The proportion of patients who had skeletal events was 24% in arm A, 21% in arm B, and 18% in arm C (P = .4). Thalidomide is an effective maintenance therapy in patients with multiple myeloma. Maintenance treatment with pamidronate does not decrease the incidence of bone events.

Alkylpurine-DNA-N-glycosylase confers resistance to temozolomide in xenograft models of glioblastoma multiforme and is associated with poor survival in patients.

Glioblastoma multiforme (GBM) is the most common and lethal of all gliomas. The current standard of care includes surgery followed by concomitant radiation and chemotherapy with the DNA alkylating agent temozolomide (TMZ). O⁶-methylguanine-DNA methyltransferase (MGMT) repairs the most cytotoxic of lesions generated by TMZ, O⁶-methylguanine. Methylation of the MGMT promoter in GBM correlates with increased therapeutic sensitivity to alkylating agent therapy. However, several aspects of TMZ sensitivity are not explained by MGMT promoter methylation. Here, we investigated our hypothesis that the base excision repair enzyme alkylpurine-DNA-N-glycosylase (APNG), which repairs the cytotoxic lesions N³-methyladenine and N⁷-methylguanine, may contribute to TMZ resistance. Silencing of APNG in established and primary TMZ-resistant GBM cell lines endogenously expressing MGMT and APNG attenuated repair of TMZ-induced DNA damage and enhanced apoptosis. Reintroducing expression of APNG in TMZ-sensitive GBM lines conferred resistance to TMZ in vitro and in orthotopic xenograft mouse models. In addition, resistance was enhanced with coexpression of MGMT. Evaluation of APNG protein levels in several clinical datasets demonstrated that in patients, high nuclear APNG expression correlated with poorer overall survival compared with patients lacking APNG expression. Loss of APNG expression in a subset of patients was also associated with increased APNG promoter methylation. Collectively, our data demonstrate that APNG contributes to TMZ resistance in GBM and may be useful in the diagnosis and treatment of the disease.

Role of intra-operative duplex ultrasound in living renal donor transplantation

Objective: The treatment of choice in end-stage renal disease is¦transplantation.¦Hemodynamic disturbances can evoke graft loss, while early ultrasound¦identification of vascular problems improves outcome. The primary endpoint¦of this study was to identify differences in post-op complications with and¦without systematic, intraoperative Doppler ultrasound use. A secondary aim¦was to find a predictive resistance index limit which would show where surgical¦reintervention was necessary.¦Methods: Between Jan 2000 and Dec 2010, 108 renal transplants were¦performed from living donors at our institution. In group 1 (n = 67),¦intra-operative duplex ultrasound and intra-parenchymatous resistance index¦measurements assessed patients, while in group 2 (n = 41), no ultrasound was¦performed.¦Results. There were no inter-group differences in the overall post-op¦complication rate or in benefit to graft or patient survival with Doppler use,¦however, significantly more vascular complications (10% vs. 0%, p = 0·02) and¦more acute rejections (37% vs. 10%) occurred in group 2 than in group 1,¦respectively. When the resistance index was <0·5 intra-operatively, immediate¦surgical revision was undertaken to raise the index >0·6.¦Results: There were no inter-group differences in the overall post-op¦complication rate or in benefit to graft or patient survival with Doppler use,¦however, significantly more vascular complications (10% vs. 0%, p = 0·02) and¦more acute rejections (37% vs. 10%) occurred in group 2 than in group 1,¦respectively. When the resistance index was <0·5 intra-operatively, immediate¦surgical revision was undertaken to raise the index >0·6.¦Conclusion: This is the first report demonstrating benefits of systematic¦intraoperative Doppler ultrasound on post-operative complications in renal¦transplantation from living donors. Our results support surgical revision with a¦resistance index <0·5.

Survival impact of lung transplantation for COPD.

Chronic obstructive pulmonary disease (COPD) is the primary indication for lung transplantation (LTx), but survival benefit is still under debate. We analysed the survival impact of LTx in COPD with a new approach, using the BODE (body mass index, airway obstruction, dyspnoea, exercise capacity) index. We retrospectively reviewed 54 consecutive lung transplants performed for COPD. The pre-transplant BODE score was calculated for each patient and a predicted survival was derived from the survival functions of the original BODE index validation cohort. Predicted and observed post-transplant survival was then compared. In the subgroups with a BODE score >or=7 and <7, a majority of patients (66% and 69%, respectively) lived for longer after LTx than predicted by their individual BODE index. The median survival was significantly improved in the entire cohort and in the subgroup with a BODE score >or=7. 4 yrs after LTx a survival benefit was only apparent in patients with a pre-transplant BODE score of >or=7. In conclusion, while a majority of COPD patients had an individual survival benefit from LTx regardless of their pre-transplant BODE score, a global survival benefit was seen only in patients with more severe disease. This supports the use of the BODE index as a selection criteria for LTx candidates.

Life-history evolution and the polyphenic regulation of somatic maintenance and survival.

Here we discuss life-history evolution from the perspective of adaptive phenotypic plasticity, with a focus on polyphenisms for somatic maintenance and survival. Polyphenisms are adaptive discrete alternative phenotypes that develop in response to changes in the environment. We suggest that dauer larval diapause and its associated adult phenotypes in the nematode (Caenorhabditis elegans), reproductive dormancy in the fruit fly (Drosophila melanogaster) and other insects, and the worker castes of the honey bee (Apis mellifera) are examples of what may be viewed as the polyphenic regulation of somatic maintenance and survival. In these and other cases, the same genotype can--depending upon its environment--express either of two alternative sets of life-history phenotypes that differ markedly with respect to somatic maintenance, survival ability, and thus life span. This plastic modulation of somatic maintenance and survival has traditionally been underappreciated by researchers working on aging and life history. We review the current evidence for such adaptive life-history switches and their molecular regulation and suggest that they are caused by temporally and/or spatially varying, stressful environments that impose diversifying selection, thereby favoring the evolution of plasticity of somatic maintenance and survival under strong regulatory control. By considering somatic maintenance and survivorship from the perspective of adaptive life-history switches, we may gain novel insights into the mechanisms and evolution of aging.

Supraclavicular flap in head and neck reconstruction: experience in 50 consecutive patients.

The supraclavicular flap (SCF) is a fasciocutaneous flap used to cover head, oral, and neck region defects after tumor resection. Its main vascular supply is the supraclavicular artery and accompanying veins and it can be harvested as a vascularised pedicled flap. The SCF serves as an excellent outer skin cover as well as a good inner mucosal lining after oral cavity and head-neck tumor resections. The flap has a wide arc of rotation and matches the skin colour and texture of the face and neck. Between March 2006 and March 2011, the pedicled supraclavicular flap was used for reconstruction in 50 consecutive patients after head and neck tumor resections and certain benign conditions in a tertiary university hospital setting. The flaps were tunnelized under the neck skin to cover the external cervicofacial defects or passed medial to the mandible to give an inner epithelial lining after the oral cavity and oropharyngeal tumor excision. Forty-four of the 50 patients had 100% flap survival with excellent wound healing. All the flaps were harvested in less than 1 h. There were four cases of distal tip desquamation and two patients had complete flap necrosis. Distal flap desquamation was observed in SCFs used for resurfacing the external skin defects after oral cavity tumor ablation and needed only conservative treatment measures. Total flap failure was encountered in two patients who had failed in previous chemoradiotherapy for squamous cell cancer of the floor of mouth and tonsil, respectively, and the SCF was used in mucosal defect closure after tumor ablation. The benefits of a pedicled fasciocutaneous supraclavicular flap are clear; it is thin, reliable, easy, and quick to harvest. In head, face and neck reconstructions, it is a good alternative to free fasciocutaneous flaps, regional pedicled myocutaneous flaps, and the deltopectoral flap.

Frequent accesses to totally implanted vascular ports in pediatric oncology patients are associated with higher infection rates.

PURPOSE: Totally implanted vascular (TIVA) ports are used in children for repeated blood samples or intravenous treatments. We have recently published a prospective evaluation of surgical incidents and early complications associated with these devices. This work is the final part of the same study, assessing late complications over a follow-up of 2 yrs. METHODS: From January 2006 to January 2008, children older than 1 yr of age with a diagnosis of solid or blood cell malignancy were included. Insertion technique and care of the device were standardized. Every manipulation was prospectively recorded by specialized nurses. Obstruction was documented clinically. When bacteremia was suspected, routine central and peripheral blood cultures were drawn. RESULTS: Forty-five consecutive patients were enrolled in the study. Mean age at the time of the procedure was 8.5 yrs. There was no catheter-related infection within the first 4 weeks post-surgery. No device had to be removed because of infection or obstruction during follow-up. Frequent accesses to the port (=3 per day over a 10-day period) were associated with an 8-fold risk of infection. CONCLUSION: Insertion and use of TIVA devices were frequently associated with complications. No device had to be removed because of infection or obstruction over the follow-up period, although no prophylactic antibiotic agent was used. Restrictive use of antibiotics may prevent opportunistic infection. Frequent access to the device was significantly associated with line infection (odds ratio=8.43). No risk factor was identified for obstruction which occurred at a rate of 5.3 per 10,000 accesses.

Contribution of (sub)domains of Staphylococcus aureus fibronectin-binding protein to the proinflammatory and procoagulant response of human vascular endothelial cells.

The Staphylococcus aureus fibronectin (Fn) -binding protein A (FnBPA) is involved in bacterium-endothelium interactions which is one of the crucial events leading to infective endocarditis (IE). We previously showed that the sole expression of S. aureus FnBPA was sufficient to confer to non-invasive Lactococcus lactis bacteria the capacity to invade human endothelial cells (ECs) and to launch the typical endothelial proinflammatory and procoagulant responses that characterize IE. In the present study we further questioned whether these bacterium-EC interactions could be reproduced by single or combined FnBPA sub-domains (A, B, C or D) using a large library of truncated FnBPA constructs expressed in L. lactis. Significant invasion of cultured ECs was found for L. lactis expressing the FnBPA subdomains CD (aa 604-877) or A4(+16) (aa 432-559). Moreover, this correlates with the capacity of these fragments to elicit in vitro a marked increase in EC surface expression of both ICAM-1 and VCAM-1 and secretion of the CXCL8 chemokine and finally to induce a tissue factor-dependent endothelial coagulation response. We thus conclude that (sub)domains of the staphylococcal FnBPA molecule that express Fn-binding modules, alone or in combination, are sufficient to evoke an endothelial proinflammatory as well as a procoagulant response and thus account for IE severity.