Effects of hyperglycemia on adipose-derived mesenchymal stromal cells: a study on their proangiogenic and immunomodulatory potential

Background. Hyperglycemia and poor glycemic control are two main features of diabetes mellitus. People affected by this disease are prone in developing serious secondary complications, which actually affect their life resulting in impaired life quality and even increased mortality. These are microvascular complications such as diabetic retinopathy (DR), diabetic nephropathy and diabetic neuropathy. The pathophysiological features of these complications are very similar and concern a progressive microvascular degeneration, which impairs the normal microvascular functions and vascular permeability. Hyperglycemia represents a connection point between all these secondary complications since high glucose has been identified as one of the causes mediating cell damage. Through years, clear evidences on hyperglycemia-mediated cell damage have been produced, leading to the elaboration of the “unifying hypothesis”. Here, glucose-mediated cell damage is explained as the result of a deleterious increase of oxidative stress in cells, which induces cell dysfunction and death. This mechanism has been demonstrated in several cell types from endothelial to neuronal cells, however, some aspects of hyperglycemia-mediated cell damages still need to be elucidated. The application of cell/stem cell-mediated therapies in diabetes and diabetic complications already gave some positive outcomes. In particular, mesenchymal stromal cells (MSC)-based interventions appeared to be safe and efficient. Thanks to their differentiating potential, proangiogenic features and immunomodulatory properties, pre-clinical studies of MSC application in DR gave very promising results. For instance, in animal models of DR, MSC applications often resulted in positive outcomes, delaying the progression of the disease, reducing vascular permeability and restoring functionality. However, the majority of studies used an allogenic approach proposing the application of MSC from healthy donors. Indeed, MSC from diabetic donors often revealed to be impaired by the disease, which caused reduction of their potency and even affecting their survival. In this context, specific investigations focused on possible effects of hyperglycemia on MSC are lacking. Aim. The main goal of this study was to evaluate whether adipose-derived MSC were impaired by hyperglycemia in their phenotype and functions through in vitro experimentations. Methods. Our study proposes a large-spectrum of investigations of possible hyperglycemia-mediated cell damage on adipose derived MSC (ASC) through in vitro experimentations. In particular, we assessed effects of hyperglycemia on (1) cellular phenotype, (2) ASC proangiogenic potential, (3) ASC pericyte-like function and (4) immunomodulatory potential. 1. ASC were cultured in normal glucose (1g/L, NG) and high glucose (4.5g/L, HG) culture media. These two counterparts were compared in terms of phenotype, cell growth, differentiation potential and cell surface markers. Oxidative stress and glucose uptake were also measured. Similarly, hyperglycemia-mediated effects were also assessed on microvascular retinal endothelial cells (HRMVEC). 2. The proangiogenic potential of ASC was evaluated establishing cocultures with endothelial cells (EC) and analyzing their supernatants for angiogenic growth factor. These supernatants were used as conditioned media to rescue HRMVEC angiogenic capacity in presence of HG. 3. Immunofluorescence was used to investigate ASC for pericyte-like marker when cocultured with HRMVEC. Anti-α-smooth muscle (SMA) and anti-NG-2 antibodies were tested. 4. ASC and CD4 T cell interactions were characterized through ASC and peripheral blood mononuclear cells (PBMC) direct and indirect (transwell) cocultures. Two distinct conditions were evaluated: (a) stimulated coculture, where PBMC were stimulated with CD3/CD28 beads and (b) unstimulated (or not-stimulated) cocultures, where resting PBMC were used. For both (a) and (b) CD4 T cell proliferation, quantification of CD4+CD25+ and regulatory T cell (Treg) fractions and analysis of coculture supernatants were performed. Results. Overall, we observed that hyperglycemia did not affect any of the evaluated aspects. Indeed, ASC demonstrated a strong refractory behavior towards HG exposure resulting only in a transient increase of intracellular oxidative stress. On the contrary, HRMVEC appeared sensitive to HG, which increased the level of oxidative stress and affected their angiogenic potential suggesting a possible connection between the two aspects. ASC proangiogenic potential was not affected by glucose and ASC sustained HRMVEC angiogenesis giving structural support and secreting proangiogenic growth factors. ASC conditioned media rescued angiogenic potential of HG HRMVEC. ASC pericyte-like function was detected independently of glucose, showing a strong α-SMA expression in ASC in contact with endothelial cells. Moreover, pericytes did not support the angiogenesis of EC, confirming their role in support homeostasis in the vascular department. Finally, we found that glucose did not affect the immunomodulatory potential of ASC. Actually, the major discriminant in the outcomes of the experiments was the presence of CD3/CD28-stimulated or not stimulated (resting) PBMC. In stimulated cocultures, ASC strongly inhibited CD4 proliferation via the IDO-kynurenine pathway, caused an increased CD25 expression on CD4 cells and did not induce Treg formation. In resting cocultures, IDO and kynurenine were also measured at low levels but ASC-inhibiting effect of CD4 was not detectable. Here, Treg highly proliferated in cocultures. Cytokine analysis confirmed Treg induction finding high concentrations of CCL-18 and TGF-β, two well-known factors involved in ASC-mediated Treg generation. Moreover, cocultures caused a long-lasting priming of PBMC, which secreted high concentration of IL-10 and CCL-18, denoting a switch towards an anti-inflammatory phenotype. Conclusions. In relation to DR and the eventual application of MSC-based cell therapy in diabetic patients our study provides the important evidence that hyperglycemia does not affect ASC in their basic characteristics, proangiogenic and immunomodulatory potential. Having demonstrated the strong immunomodulatory effect of ASC in vitro and because of recent publication on Treg involvement in the retina, we auspicate that in vivo experimentations may clarify and characterize this mechanism, which may represent a real novelty in DR treatment

Mesenchymal stromal/stem cells as potential therapy in diabetic retinopathy

Diabetic retinopathy (DR) is a multifactorial microvascular disease induced by hyperglycemia and subsequent metabolic abnormalities. The resulting cell stress causes a sequela of events that ultimately can lead to severe vision impairment and blindness. The early stages are characterized by activation of glia and loss of pericytes, endothelial cells (EC) and neuronal cells. The integrity of the retinal microvasculature becomes affected, and, as a possible late response, macular edema may develop as a common reason for vision loss in patients with non proliferative DR. Moreover, the local ischemia can trigger vasoproliferation leading to vision-threating proliferative DR (PDR) in humans. Available treatment options include control of metabolic and hemodynamic factors. Timely intervention of advanced DR stages with laser photocoagulation, intraocular anti-vascular endothelial growth factor (VEGF) or glucocorticoid drugs can reduce vision loss. As the pathology involves cell loss of both the vascular and neuroglial compartments, cell replacement strategies by stem and progenitor cells have gained considerable interest in the past years. Compared to other disease entities, so far little is known about the efficacy and potential mode of action of cell therapy in treatment of DR. In preclinical models of DR different cell types have been applied ranging from embryonic or induced pluripotent stem cells, hematopoietic stem cells, and endothelial progenitor cells to mesenchymal stromal cells (MSC). The latter cell population can combine various modes of action (MoA), thus they are among the most intensely tested cell types in cell therapy. The aim of this review is to discuss the rationale for using MSC as potential cell therapy to treat DR. Accordingly, we will revise identified MoA of MSCs and speculate how these may support the repair of the damaged retina

Pro-angiogenic Activity Discriminates Human Adipose-Derived Stromal Cells From Retinal Pericytes: Considerations for Cell-Based Therapy of Diabetic Retinopathy

Diabetic retinopathy (DR) is a frequent diabetes-associated complication. Pericyte dropout can cause increased vascular permeability and contribute to vascular occlusion. Adipose-derived stromal cells (ASC) have been suggested to replace pericytes and restore microvascular support as potential therapy of DR. In models of DR, ASC not only generated a cytoprotective and reparative environment by the secretion of trophic factors but also engrafted and integrated into the retina in a pericyte-like fashion. The aim of this study was to compare the pro-angiogenic features of human ASC and human retinal microvascular pericytes (HRMVPC) in vitro. The proliferation and the expression of ASC and HRMVPC markers were compared. Adhesion to high glucose-conditioned endothelial extracellular matrix, mimicking the diabetic microenvironment, was measured. The angiogenesis-promoting features of both cell types and their conditioned media on human retinal endothelial cells (EC) were assessed. To identify a molecular basis for the observed differences, gene expression profiling was performed using whole-genome microarrays, and data were validated using PCR arrays and flow cytometry. Based on multiplex cytokine results, functional studies on selected growth factors were performed to assess their role in angiogenic support. Despite a distinct heterogeneity in ASC and HRMVPC cultures with an overlap of expressed markers, ASC differed functionally from HRMVPC. Most importantly, the pro-angiogenic activity was solely featured by ASC, whereas HRMVPC actively suppressed vascular network formation. HRMVPC, in contrast to ASC, showed impaired adhesion and proliferation on the high glucose-conditioned endothelial extracellular matrix. These data were supported by gene expression profiles with differentially expressed genes. The vessel-stabilizing factors were more highly expressed in HRMVPC, and the angiogenesis-promoting factors were more highly expressed in ASC. The vascular endothelial growth factor receptor-2 inhibition efficiently abolished the ASC angiogenic supportive capacities, whereas the addition of angiopoietin-1 and angiopoietin-2 did not alter these effects. Our results clearly show that ASC are pro-angiogenic, whereas HRMVPC are marked by anti-angiogenic/EC-stabilizing features. These data support ASC as pericyte replacement in DR but also suggest a careful risk-to-benefit analysis to take full advantage of the ASC therapeutic features

Pro-angiogenic Activity Discriminates Human Adipose-Derived Stromal Cells From Retinal Pericytes: Considerations for Cell-Based Therapy of Diabetic Retinopathy

Diabetic retinopathy (DR) is a frequent diabetes-associated complication. Pericyte dropout can cause increased vascular permeability and contribute to vascular occlusion. Adipose-derived stromal cells (ASC) have been suggested to replace pericytes and restore microvascular support as potential therapy of DR. In models of DR, ASC not only generated a cytoprotective and reparative environment by the secretion of trophic factors but also engrafted and integrated into the retina in a pericyte-like fashion. The aim of this study was to compare the pro-angiogenic features of human ASC and human retinal microvascular pericytes (HRMVPC) in vitro. The proliferation and the expression of ASC and HRMVPC markers were compared. Adhesion to high glucose-conditioned endothelial extracellular matrix, mimicking the diabetic microenvironment, was measured. The angiogenesis-promoting features of both cell types and their conditioned media on human retinal endothelial cells (EC) were assessed. To identify a molecular basis for the observed differences, gene expression profiling was performed using whole-genome microarrays, and data were validated using PCR arrays and flow cytometry. Based on multiplex cytokine results, functional studies on selected growth factors were performed to assess their role in angiogenic support. Despite a distinct heterogeneity in ASC and HRMVPC cultures with an overlap of expressed markers, ASC differed functionally from HRMVPC. Most importantly, the pro-angiogenic activity was solely featured by ASC, whereas HRMVPC actively suppressed vascular network formation. HRMVPC, in contrast to ASC, showed impaired adhesion and proliferation on the high glucose-conditioned endothelial extracellular matrix. These data were supported by gene expression profiles with differentially expressed genes. The vessel-stabilizing factors were more highly expressed in HRMVPC, and the angiogenesis-promoting factors were more highly expressed in ASC. The vascular endothelial growth factor receptor-2 inhibition efficiently abolished the ASC angiogenic supportive capacities, whereas the addition of angiopoietin-1 and angiopoietin-2 did not alter these effects. Our results clearly show that ASC are pro-angiogenic, whereas HRMVPC are marked by anti-angiogenic/EC-stabilizing features. These data support ASC as pericyte replacement in DR but also suggest a careful risk-to-benefit analysis to take full advantage of the ASC therapeutic features

Differential cross section measurements for the production of a W boson in association with jets in proton–proton collisions at √s = 7 TeV

Measurements are reported of differential cross sections for the production of a W boson, which decays into a muon and a neutrino, in association with jets, as a function of several variables, including the transverse momenta (pT) and pseudorapidities of the four leading jets, the scalar sum of jet transverse momenta (HT), and the difference in azimuthal angle between the directions of each jet and the muon. The data sample of pp collisions at a centre-of-mass energy of 7 TeV was collected with the CMS detector at the LHC and corresponds to an integrated luminosity of 5.0 fb[superscript −1]. The measured cross sections are compared to predictions from Monte Carlo generators, MadGraph + pythia and sherpa, and to next-to-leading-order calculations from BlackHat + sherpa. The differential cross sections are found to be in agreement with the predictions, apart from the pT distributions of the leading jets at high pT values, the distributions of the HT at high-HT and low jet multiplicity, and the distribution of the difference in azimuthal angle between the leading jet and the muon at low values.United States. Dept. of EnergyNational Science Foundation (U.S.)Alfred P. Sloan Foundatio

Optimasi Portofolio Resiko Menggunakan Model Markowitz MVO Dikaitkan dengan Keterbatasan Manusia dalam Memprediksi Masa Depan dalam Perspektif Al-Qur`an

Risk portfolio on modern finance has become increasingly technical, requiring the use of sophisticated mathematical tools in both research and practice. Since companies cannot insure themselves completely against risk, as human incompetence in predicting the future precisely that written in Al-Quran surah Luqman verse 34, they have to manage it to yield an optimal portfolio. The objective here is to minimize the variance among all portfolios, or alternatively, to maximize expected return among all portfolios that has at least a certain expected return. Furthermore, this study focuses on optimizing risk portfolio so called Markowitz MVO (Mean-Variance Optimization). Some theoretical frameworks for analysis are arithmetic mean, geometric mean, variance, covariance, linear programming, and quadratic programming. Moreover, finding a minimum variance portfolio produces a convex quadratic programming, that is minimizing the objective function ðð¥with constraintsð ð 𥠥 ðandð´ð¥ = ð. The outcome of this research is the solution of optimal risk portofolio in some investments that could be finished smoothly using MATLAB R2007b software together with its graphic analysis

Penilaian Kinerja Keuangan Koperasi di Kabupaten Pelalawan

This paper describe development and financial performance of cooperative in District Pelalawan among 2007 - 2008. Studies on primary and secondary cooperative in 12 sub-districts. Method in this stady use performance measuring of productivity, efficiency, growth, liquidity, and solvability of cooperative. Productivity of cooperative in Pelalawan was highly but efficiency still low. Profit and income were highly, even liquidity of cooperative very high, and solvability was good

Juxtaposing BTE and ATE – on the role of the European insurance industry in funding civil litigation

One of the ways in which legal services are financed, and indeed shaped, is through private insurance arrangement. Two contrasting types of legal expenses insurance contracts (LEI) seem to dominate in Europe: before the event (BTE) and after the event (ATE) legal expenses insurance. Notwithstanding institutional differences between different legal systems, BTE and ATE insurance arrangements may be instrumental if government policy is geared towards strengthening a market-oriented system of financing access to justice for individuals and business. At the same time, emphasizing the role of a private industry as a keeper of the gates to justice raises issues of accountability and transparency, not readily reconcilable with demands of competition. Moreover, multiple actors (clients, lawyers, courts, insurers) are involved, causing behavioural dynamics which are not easily predicted or influenced. Against this background, this paper looks into BTE and ATE arrangements by analysing the particularities of BTE and ATE arrangements currently available in some European jurisdictions and by painting a picture of their respective markets and legal contexts. This allows for some reflection on the performance of BTE and ATE providers as both financiers and keepers. Two issues emerge from the analysis that are worthy of some further reflection. Firstly, there is the problematic long-term sustainability of some ATE products. Secondly, the challenges faced by policymakers that would like to nudge consumers into voluntarily taking out BTE LEI

Search for stop and higgsino production using diphoton Higgs boson decays

Results are presented of a search for a "natural" supersymmetry scenario with gauge mediated symmetry breaking. It is assumed that only the supersymmetric partners of the top-quark (stop) and the Higgs boson (higgsino) are accessible. Events are examined in which there are two photons forming a Higgs boson candidate, and at least two b-quark jets. In 19.7 inverse femtobarns of proton-proton collision data at sqrt(s) = 8 TeV, recorded in the CMS experiment, no evidence of a signal is found and lower limits at the 95% confidence level are set, excluding the stop mass below 360 to 410 GeV, depending on the higgsino mass

Impacts of the Tropical Pacific/Indian Oceans on the Seasonal Cycle of the West African Monsoon

The current consensus is that drought has developed in the Sahel during the second half of the twentieth century as a result of remote effects of oceanic anomalies amplified by local land–atmosphere interactions. This paper focuses on the impacts of oceanic anomalies upon West African climate and specifically aims to identify those from SST anomalies in the Pacific/Indian Oceans during spring and summer seasons, when they were significant. Idealized sensitivity experiments are performed with four atmospheric general circulation models (AGCMs). The prescribed SST patterns used in the AGCMs are based on the leading mode of covariability between SST anomalies over the Pacific/Indian Oceans and summer rainfall over West Africa. The results show that such oceanic anomalies in the Pacific/Indian Ocean lead to a northward shift of an anomalous dry belt from the Gulf of Guinea to the Sahel as the season advances. In the Sahel, the magnitude of rainfall anomalies is comparable to that obtained by other authors using SST anomalies confined to the proximity of the Atlantic Ocean. The mechanism connecting the Pacific/Indian SST anomalies with West African rainfall has a strong seasonal cycle. In spring (May and June), anomalous subsidence develops over both the Maritime Continent and the equatorial Atlantic in response to the enhanced equatorial heating. Precipitation increases over continental West Africa in association with stronger zonal convergence of moisture. In addition, precipitation decreases over the Gulf of Guinea. During the monsoon peak (July and August), the SST anomalies move westward over the equatorial Pacific and the two regions where subsidence occurred earlier in the seasons merge over West Africa. The monsoon weakens and rainfall decreases over the Sahel, especially in August.Peer reviewe