Angiogenesis-dependent and independent phases of intimal hyperplasia.

BACKGROUND: Neointimal vascular smooth muscle cell (VSMC) proliferation is a primary cause of occlusive vascular disease, including atherosclerosis, restenosis after percutaneous interventions, and bypass graft stenosis. Angiogenesis is implicated in the progression of early atheromatous lesions in animal models, but its role in neointimal VSMC proliferation is undefined. Because percutaneous coronary interventions result in induction of periadventitial angiogenesis, we analyzed the role of this process in neointima formation. METHODS AND RESULTS: Local injury to the arterial wall in 2 different animal models induced periadventitial angiogenesis and neointima formation. Application of angiogenesis stimulators vascular endothelial growth factor (VEGF-A165) or a proline/arginine-rich peptide (PR39) to the adventitia of the injured artery induced a marked increase in neointimal thickening beyond that seen with injury alone in both in vivo models. Inhibition of either VEGF (with soluble VEGF receptor 1 [sFlt1]) or fibroblast growth factor (FGF) (with a dominant=negative form of FGF receptor 1 [FGF-R1DN]), respectively, signaling reduced adventitial thickening induced by VEGF and PR39 to the level seen with mechanical arterial injury alone. However, neither inhibitor was effective in preventing neointimal thickening after mechanical injury when administered in the absence of angiogenic growth factor. CONCLUSIONS: Our findings indicate that adventitial angiogenesis stimulates intimal thickening but does not initiate it

Low interleukin-2 concentration favors generation of early memory T cells over effector phenotypes during chimeric antigen receptor T-cell expansion

Errata: 10.1016/j.jcyt.2017.06.003Background. Adoptive T-cell therapy offers new options for cancer treatment. Clinical results suggest that T-cell persistence, depending on T-cell memory, improves efficacy. The use of interleukin (IL)-2 for in vitro T-cell expansion is not straightforward because it drives effector T-cell differentiation but does not promote the formation of T-cell memory. We have developed a cost-effective expansion protocol for chimeric antigen receptor (CAR) T cells with an early memory phenotype. Methods. Lymphocytes were transduced with third-generation lentiviral vectors and expanded using CD3/CD28 microbeads. The effects of altering the IL-2 supplementation (0-300 IU/mL) and length of expansion (10-20 days) on the phenotype of the T-cell products were analyzed. Results. High IL-2 levels led to a decrease in overall generation of early memory T cells by both decreasing central memory T cells and augmenting effectors. T memory stem cells (T-SCM, CD95(+)CD45RO(-)CD45RA(+)CD27(+)) were present variably during T-cell expansion. However, their presence was not IL-2 dependent but was linked to expansion kinetics. CD19-CART cells generated in these conditions displayed in vitro antileukemic activity. In summary, production of CART cells without any cytokine supplementation yielded the highest proportion of early memory T cells, provided a 10 fold cell expansion and the cells were functionally potent. Discussion. The number of early memory T cells in a T-cell preparation can be increased by simply reducing the amount of IL-2 and limiting the length of T-cell expansion, providing cells with potentially higher in vivo performance. These findings are significant for robust and cost:effective T-cell manufacturing.Peer reviewe

Lack of cardiac and high-fat diet induced metabolic phenotypes in two independent strains of Vegf-b knockout mice

Peer reviewe

Extracellular vesicles provide a capsid-free vector for oncolytic adenoviral DNA delivery

Extracellular vesicles (EVs) have been showcased as auspicious candidates for delivering therapeutic cargo, including oncolytic viruses for cancer treatment. Delivery of oncolytic viruses in EVs could provide considerable advantages, hiding the viruses from the immune system and providing alternative entry pathways into cancer cells. Here we describe the formation and viral cargo of EVs secreted by cancer cells infected with an oncolytic adenovirus (IEVs, infected cell-derived EVs) as a function of time after infection. IEVs were secreted already before the lytic release of virions and their structure resembled normally secreted EVs, suggesting that they were not just apoptotic fragments of infected cells. IEVs were able to carry the viral genome and induce infection in other cancer cells. As such, the role of EVs in the life cycle of adenoviruses may be an important part of a successful infection and may also be harnessed for cancer- and gene therapy.Peer reviewe

Long-Term Intravitreal Ranibizumab as a Potential Additional Risk Factor for Neurodegeneration in Parkinson's Disease: A Case Report.

In November 2012, a 72-year old patient was diagnosed with left eye wet age-related macular degeneration. The patient received three monthly intravitreal injections of ranibizumab, with complete resolution of retinal hemorrhage and edema and reinstatement of visual acuity. In May 2015, symptomatic relapse was detected. The patient was again treated with intravitreal ranibizumab, with overall six injections till the end of February 2016. In May 2016, the patient complained of left hand resting tremor, bradykinesia, and postural rigidity of head and trunk. A diagnosis of clinically established PD was made based on new criteria of the Movement Disorders Society. Single Photon Emission Computerized Tomography of the Dopamine Transporter with (123I) ioflupane documented a low Dopamine Transporter (DAT) uptake mostly in the right striatum. Due to the documented protective role of vascular endothelial growth factor (VEGF) on the dopaminergic neurons, intensive intravitreal injections of the anti-VEGF agent ranibizumab may have played as an additional risk factor accelerating the neurodegeneration process related to PD and the onset of the related clinical signs and symptoms

Angiopoietin-2 blocking antibodies reduce early atherosclerotic plaque development in mice

Objective: Angiopoietin-2 (Ang-2) blocking agents are currently undergoing clinical trials for use in cancer treatment. Ang-2 has also been associated with rupture-prone atherosclerotic plaques in humans, suggesting a role for Ang-2 in plaque stability. Despite the availability of Ang-2 blocking agents, their clinical use is still lacking. Our aim was to establish if Ang-2 has a role in atheroma development and in the transition of subclinical to clinically relevant atherosclerosis. We investigated the effect of antibody-mediated Ang-2 blockage on atherogenesis after in a mouse model of atherosclerosis. Methods: Hypercholesterolemic (low-density lipoprotein receptor(-/-) apolipoprotein B-100/100) mice were subjected to high-cholesterol diet for eight weeks, one group with and one group without Ang-2 blocking antibody treatment during weeks 4-8. To enhance plaque development, a peri-adventitial collar was placed around the carotid arteries at the start of antibody treatment. Aortic root, carotid arteries and brachiocephalic arteries were analyzed to evaluate the effect of Ang-2 blockage on atherosclerotic plaque size and stable plaque characteristics. Results: Anti-Ang-2 treatment reduced the size of fatty streaks in the brachiocephalic artery (-72%, p <0.05). In addition, antibody-mediated Ang-2 blockage reduced plasma triglycerides (-27%, p <0.05). In contrast, Ang-2 blockage did not have any effect on the size or composition (collagen content, macrophage percentage, adventitial microvessel density) of pre-existing plaques in the aortic root or collar-induced plaques in the carotid artery. Conclusions: Ang-2 blockage was beneficial as it decreased fatty streak formation and plasma triglyceride levels, but had no adverse effect on pre-existing atherosclerosis in hypercholesterolemic mice. (C) 2015 The Authors. Published by Elsevier Ireland Ltd.Peer reviewe

Intracerebral overexpression of miR-669c is protective in mouse ischemic stroke model by targeting MyD88 and inducing alternative microglial/macrophage activation

Background Ischemic stroke is a devastating disease without a cure. The available treatments for ischemic stroke, thrombolysis by tissue plasminogen activator, and thrombectomy are suitable only to a fraction of patients and thus novel therapeutic approaches are urgently needed. The neuroinflammatory responses elicited secondary to the ischemic attack further aggravate the stroke-induced neuronal damage. It has been demonstrated that these responses are regulated at the level of non-coding RNAs, especially miRNAs. Methods We utilized lentiviral vectors to overexpress miR-669c in BV2 microglial cells in order to modulate their polarization. To detect whether the modulation of microglial activation by miR-669c provides protection in a mouse model of transient focal ischemic stroke, miR-669c overexpression was driven by a lentiviral vector injected into the striatum prior to induction of ischemic stroke. Results Here, we demonstrate that miR-669c-3p, a member of chromosome 2 miRNA cluster (C2MC), is induced upon hypoxic and excitotoxic conditions in vitro and in two different in vivo models of stroke. Rather than directly regulating the neuronal survival in vitro, miR-669c is capable of attenuating the microglial proinflammatory activation in vitro and inducing the expression of microglial alternative activation markers arginase 1 (Arg1), chitinase-like 3 (Ym1), and peroxisome proliferator-activated receptor gamma (PPAR-gamma). Intracerebral overexpression of miR-669c significantly decreased the ischemia-induced cell death and ameliorated the stroke-induced neurological deficits both at 1 and 3 days post injury (dpi). Albeit miR-669c overexpression failed to alter the overall Iba1 protein immunoreactivity, it significantly elevated Arg1 levels in the ischemic brain and increased colocalization of Arg1 and Iba1. Moreover, miR-669c overexpression under cerebral ischemia influenced several morphological characteristics of Iba1 positive cells. We further demonstrate the myeloid differentiation primary response gene 88 (MyD88) transcript as a direct target for miR-669c-3p in vitro and show reduced levels of MyD88 in miR-669c overexpressing ischemic brains in vivo. Conclusions Collectively, our data provide the evidence that miR-669c-3p is protective in a mouse model of ischemic stroke through enhancement of the alternative microglial/macrophage activation and inhibition of MyD88 signaling. Our results accentuate the importance of controlling miRNA-regulated responses for the therapeutic benefit in conditions of stroke and neuroinflammation.Peer reviewe