Shaping vessels and microenvironment: adipose stromal cells in retinal-related diseases

Diabetes veroorzaakt verschillende complicaties in het lichaam. In dit proefschrift hebben we ons gericht op diabetische retinopathie. Retinopathie beïnvloedt voornamelijk de microcirculatie in het netvlies. Vervolgens worden meer retinale cellulaire bestanddelen gevoelig voor de glucose-onevenwichtigheid. Een mogelijke oplossing is om de retinale micro-omgeving met stamcellen te implementeren. Stamcellen, met name adipose stromale cellen (ASC), zouden mogelijk kunnen worden geïntegreerd in de ogen om de door diabetes veroorzaakte schade te herstellen. ASC werken als ondersteunende cellen, met andere woorden microvaten beschadigd door glucose kunnen worden versterkt door injectie van autoloog ASC. Het doel van dit werk was om te begrijpen welke mechanismen betrokken zijn bij het ASC-potentieel om beschadigde micro-omgevingen te herstellen en daarom de behandeling van stamcellen te verbeteren. We ontdekten dat de notch-signalering betrokken is bij ASC-communicatie met microvaten. Microvessels spelen een fundamentele rol bij het geven van voedingsstoffen en bescherming van de delicate micro-omgeving van het netvlies. Uit deze studie concludeerden we dat toekomstig werk met notch-signalering de integratie van stamcellen in het netvlies zou kunnen verbeteren. Dit proefschrift bevat een review waarin de complexiteit van diabetische retinopathie verder wordt geanalyseerd. Ontsteking speelt ook een rol bij de ontwikkeling van de ziekte en moet worden overwogen voor mogelijke proeven bij de mens. In het tweede deel van het manuscript werd de temporele dynamiek van ASC en microvatenvorming en -assemblage bestudeerd in driedimensionale culturen in vitro. We stelden een stapsgewijze vorming voor van microvaatjes geleid door ASC inclusief extracellulaire matrixvorming. Toekomstig onderzoek naar de behandeling van stamcellen moet bestaan uit cellenmetabolisme en extracellulaire matrixbetrokkenheid tijdens de ziekteprogressie

The Pericytic Phenotype of Adipose Tissue-Derived Stromal Cells Is Promoted by NOTCH2

Long-term diabetes leads to macrovascular and microvascular complication. In diabetic retinopathy (DR), persistent hyperglycemia causes permanent loss of retinal pericytes and aberrant proliferation of microvascular endothelial cells (ECs). Adipose tissue-derived stromal cells (ASCs) may serve to functionally replace retinal pericytes and normalize retinal microvasculature during disease progression. We hypothesized that Notch signaling in ASC underlies regulation and stabilization of dysfunctional retinal microvascular networks such as in DR. ASC prominently and constitutively expressed NOTCH2. Genetic knockdown of NOTCH2 in ASC (SH-NOTCH2) disturbed the formation of vascular networks of human umbilical cord vein endothelial cells both on monolayers of ASC and in organotypical three-dimensional cocultures with ASC. On ASC SH-NOTCH2, cell surface platelet-derived growth factor receptor beta was downregulated which disrupted their migration toward the chemoattractant platelet-derived growth factor beta subunits (PDGF-BB) as well as to conditioned media from EC and bovine retinal EC. This chemoattractant is secreted by pro-angiogenic EC in newly formed microvascular networks to attract pericytes. Intravitreal injected ASC SH-NOTCH2 in oxygen-induced retinopathy mouse eyes did not engraft in the preexisting retinal microvasculature. However, the in vivo pro-angiogenic capacity of ASC SH-NOTCH2 did not differ from controls. In this respect, multifocal electroretinography displayed similar b-wave amplitudes in the avascular zones when either wild type ASC or SH-NOTCH2 ASC were injected. In conclusion, our results indicate that NOTCH2 is essential to support in vitro vasculogenesis via juxtacrine interactions. In contrast, ongoing in vivo angiogenesis is influenced by paracrine signaling of ASC, irrespective of Notch signaling

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

Supportive treatment for cast nephropathy in patients with multiple myeloma; a pilot study

Introduction: Cast nephropathy is a prevalent cause of acute kidney injury (AKI) in patients with myeloma. Objectives: The aim of this study is to define the outcome of a standardized supportive therapy for cast nephropathy. Patients and Methods: Retrospective analysis of the outcome of cast nephropathy in a University hospital for a period of five years. Data analysed; serum creatinine, estimated glomerular filtration rate (eGFR; mL/min/1.73 m2 BSA) and need for dialysis. Standardized therapy with the aim of preventing/removing tubular casts; fluid administration and mannitol to increase urine flow, sodium bicarbonate to alkalize the urine and low dose steroid to reduce peritubular inflammation. Statistical analysis: Student's t-test or the Mann-Whitney test according to data distribution. A two-tailed P value <0.05 was considered statistically significant. Survival curve was drawn according to Kaplan and Meier. Results: Twenty-seven cases were reviewed. Upon admission, mean serum creatinine was 7.1±4.9 mg/dL and mean eGFR 6±4 mL/min/1.73 m2 BSA; 30% of patients had oligo-anuria. Diagnosis of cast nephropathy was presumptive in 23 patients, and renal biopsy proven in four. Hemodialysis was required by 10 (37%) patients, two of whom continued dialysis after discharge. At discharge, serum creatinine was 3.7±2.5 mg/dL and eGFR 20±13 mL/min/1.73 m2 BSA (P=0.002), and after a median of 3.4 months, the values were 2.9±2.1 mg/dL and 35±32 mL/min/1.73 m2 BSA, respectively. Patient survival was 60% after 24 months. Conclusion: Administration of fluid, mannitol, sodium bicarbonate and low-dose steroid may improve the outcome of cast nephropathy. Despite the fact that the study has many limitations, its findings could be the base for prospective controlled trials on cast nephropathy and could be useful in those countries where the expensive extracorporeal treatments are not available

Power Flow Management by Active Nodes: A Case Study in Real Operating Conditions

The role of distributor system operators is experiencing a gradual but relevant change to include enhanced ancillary and energy dispatch services needed to manage the increased power provided by intermittent distributed generations in medium voltage networks. In this context, the paper proposes the insertion, in strategic points of the network, of specific power electronic systems, denoted as active nodes, which permit the remote controllability of the active and reactive power flow. Such capabilities, as a further benefit, enable the distributor system operators to provide ancillary network services without requiring any procurement with distributed generation systems owners. In particular, the paper highlights the benefits of active nodes, demonstrating their capabilities in reducing the inverse power flow issues from medium to high voltage lines focusing on a network cluster including renewable energy resources. As a further novelty, this study has accounted for a real cluster operated by the Italian distributor system operator Areti. A specific simulation model of the electrical lines has been implemented in DigSilent PowerFactory (DIgSILENT GmbH–Germany) software using real operating data obtained during a 1-year measurement campaign. A detailed cost-benefit analysis has been provided, accounting for different load flow scenarios. The results have demonstrated that the inclusion of active nodes can significantly reduce the drawbacks related to the reverse power flow

Casting light on the European anchovy: from biology to conservation and industry

This manuscript explores the role of European anchovies (Engraulis encrasicolus) in the central Mediterranean Region, shedding light on their ecological significance, conservation challenges, and sustainable utilization. The European anchovy is one of Europe’s most important fish resources in the Mediterranean basin, and it is considered a keystone species, playing a pivotal role in both ecological and socio-economic dimensions. However, in recent decades, European anchovy, together with Sardina pilchardus (commonly known as European sardine), has suffered a population decline for several reasons. Consequently, it is necessary to improve the management of anchovy fisheries by understanding the reproductive modes and characteristics, the influence of currents on the passive transport of eggs and larvae, the feeding habits, the environmental adaptability (e.g., salinity), and the distribution of ecotypes along the Italian coasts. Such information is fundamental for the management of fisheries, especially artisanal ones, and to control frauds, especially in protected, geographically referred, and traditional high-quality commercial products. Various aspects, ranging from their population dynamics to their industrial processing and the ecological implications of these activities were delved, highlighting the knowledge about anchovy populations and ecotypes and its importance in maintaining ecosystem balance and sustaining human communities. The ecological interaction of anchovies within the food web, as essential data in the conservation actions and management of these resources was emphasized. In addition, the metabolic and stomach contents diversity among anchovy populations and ecotypes was discussed, enhancing our understanding of their adaptability to varying environmental conditions. The manuscript then explores the traditional and industrial processing of anchovies, encompassing aspects ranging from fishing techniques (i.e., methods of capture) to their industrial significance, sustainability concerns, issues of fraud, and the establishment of geographical traceability. Finally, the opportunities for sustainable and biotechnological utilization of anchovy discards were also further explored, demonstrating the potential for waste reduction and resource optimization

Molecular and Biomechanical Clues From Cardiac Tissue Decellularized Extracellular Matrix Drive Stromal Cell Plasticity

Decellularized-organ-derived extracellular matrix (dECM) has been used for many years in tissue engineering and regenerative medicine. The manufacturing of hydrogels from dECM allows to make use of the pro-regenerative properties of the ECM and, simultaneously, to shape the material in any necessary way. The objective of the present project was to investigate differences between cardiovascular tissues (left ventricle, mitral valve, and aorta) with respect to generating dECM hydrogels and their interaction with cells in 2D and 3D. The left ventricle, mitral valve, and aorta of porcine hearts were decellularized using a series of detergent treatments (SDS, Triton-X 100 and deoxycholate). Mass spectrometry-based proteomics yielded the ECM proteins composition of the dECM. The dECM was digested with pepsin and resuspended in PBS (pH 7.4). Upon warming to 37°C, the suspension turns into a gel. Hydrogel stiffness was determined for samples with a dECM concentration of 20 mg/mL. Adipose tissue-derived stromal cells (ASC) and a combination of ASC with human pulmonary microvascular endothelial cells (HPMVEC) were cultured, respectively, on and in hydrogels to analyze cellular plasticity in 2D and vascular network formation in 3D. Differentiation of ASC was induced with 10 ng/mL of TGF-β1 and SM22α used as differentiation marker. 3D vascular network formation was evaluated with confocal microscopy after immunofluorescent staining of PECAM-1. In dECM, the most abundant protein was collagen VI for the left ventricle and mitral valve and elastin for the aorta. The stiffness of the hydrogel derived from the aorta (6,998 ± 895 Pa) was significantly higher than those derived from the left ventricle (3,384 ± 698 Pa) and the mitral valve (3,233 ± 323 Pa) (One-way ANOVA, p = 0.0008). Aorta-derived dECM hydrogel drove non-induced (without TGF-β1) differentiation, while hydrogels derived from the left ventricle and mitral valve inhibited TGF-β1-induced differentiation. All hydrogels supported vascular network formation within 7 days of culture, but ventricular dECM hydrogel demonstrated more robust vascular networks, with thicker and longer vascular structures. All the three main cardiovascular tissues, myocardium, valves, and large arteries, could be used to fabricate hydrogels from dECM, and these showed an origin-dependent influence on ASC differentiation and vascular network formation