Immunological Compatibility Status of Placenta-Derived Stem Cells is Mediated by Scaffold 3D Structure

Placenta-derived amniotic epithelial cells (AECs), a great cell source for tissue engineering and stem cell therapy, are immunologically inert in their native state; however, immunological changes in these cells after culture and differentiation have challenged their applications. The aim of this study was to investigate the effect of 2D and 3D scaffolds on human lymphocyte antigens (HLA) expression by AECs. The effect of different preparation parameters including pre-freezing time and temperature was evaluated on 3D chitosan–gelatine scaffolds properties. Evaluation of MHC class I, HLA-DR and HLA-G expression in AECs after 7 d culture on 2D bed and 3D scaffold of chitosan–gelatine showed that culture of AECs on the 2D substrate up-regulated MHC class I and HLA-DR protein markers on AECs surface and down-regulated HLA-G protein. In contrast, 3D scaffold did not increase protein expression of MHC class I and HLA-DR. Moreover, HLA-G protein expression remained unchanged in 3D culture. These results confirm that 3D scaffold can remain AECs in their native immunological state and modification of physical properties of the scaffold is a key regulator of immunological markers at the gene and protein expression levels; a strategy which circumvents rejection challenge of amniotic stem cells to be translated into the clinic

Inhibition of Inflammation, Suppression of Matrix Metalloproteinases, Induction of Neurogenesis, and Antioxidant Property Make Bryostatin-1 a Therapeutic Choice for Multiple Sclerosis

Multiple sclerosis (MS) is a neurodegenerative disease characterized by inflammation and myelin damage. Pro-inflammatory cytokines, oxidative stress, high level of matrix metalloproteinases (MMPs) activity and blood–brain barrier (BBB) damage, immune-mediated destruction of myelin and neuron loss are involved in the pathogenesis of MS. The currently approved treatments for MS include injectable drugs (interferon-β and glatiramer acetate), oral drugs (fingolimod), and monoclonal antibodies (natalizumab). The mentioned therapeutic choices are mostly focused on the inhibition of inflammation. Therefore, the search for a multi-target therapeutic choice remains unchallenged. It seems that a drug with anti-inflammatory, oxidative stress inhibitory, reduction of MMPs activity, and neurogenesis stimulatory properties may be effective for treatment of MS. In this regard, Bryostatin-1 as a macrolide and marine natural product has been selected as a therapeutic choice. Studies indicate that Bryostatin-1 has anti-inflammatory and antioxidant properties and decreases MMPs level and BBB damage. Furthermore, Bryostatin-1 has a neuroprotective effect and promotes neurogenesis and differentiation of oligodendrocyte progenitor stem cells as a critical step for remyelination/myelogenesis. Based on these properties, we hypothesized here that Bryostatin-1 is an effective treatment in MS

Potential Therapeutic Features of Human Amniotic Mesenchymal Stem Cells in Multiple Sclerosis: Immunomodulation, Inflammation Suppression, Angiogenesis Promotion, Oxidative Stress Inhibition, Neurogenesis Induction, MMPs Regulation, and Remyelination Stimulation

Multiple sclerosis (MS) is an inflammatory and degenerative disorder of the central nervous system with unknown etiology. It is accompanied by demyelination of the nerves during immunological processes in the presence of oxidative stress, hypoxia, cerebral hypo-perfusion, and dysregulation in matrix metalloproteinases (MMPs). Human amniotic mesenchymal stem cells (hAMSCs) as pluripotent stem cells possess some conspicuous features which could be of therapeutic value in MS therapy. hAMSCs could mimic the cascade of signals and secrete factors needed for promoting formation of stable neovasculature and angiogenesis. hAMSCs also have immunomodulatory and immunosuppressive effects on inflammatory processes and reduce the activity of inflammatory cells, migration of microglia and inhibit recruitment of certain immune cells to injury sites. hAMSCs attenuate the oxidative stress supported by the increased level of antioxidant enzymes and the decreased level of lipid peroxidation products. Furthermore, hAMSCs enhance neuroprotection and neurogenesis in brain injuries by inhibition of inflammation and promotion of neurogenesis. hAMSCs could significantly increase the expression of neurotrophic factors, which prevents neurons from initiating programmed cell death and improves survival, development, and function of neurons. In addition, they induce differentiation of neural progenitor cells to neurons. hAMSCs could also inhibit MMPs dysregulation and consequently promote the survival of endothelial cells, angiogenesis and the stabilization of vascular networks. Considering the mentioned evidences, we hypothesized here that hAMSCs and their conditioned medium could be of therapeutic value in MS therapy due to their unique properties, including immunomodulation and inflammation suppression; angiogenesis promotion; oxidative stress inhibition; neurogenesis induction and neuroprotection; matrix metalloproteinases regulation; and remyelination stimulation

Intra-arterial tert-Butyl-Hydroperoxide Infusion Induces an Exacerbated Sensory Response in the Rat Hind Limb and is Associated with an Impaired Tissue Oxygen Uptake

The objective of this study was to investigate oxidative stress and oxygen extraction mechanisms in an animal model of continuous intra-arterial infusion of a free radical donor and in an in vitro model using isolated mitochondria. tert-Butyl-hydroperoxide (tert-BuOOH, 25 mM) was infused for 24 h in the left hind limb of rats to induce soft tissue damage (n = 8). After 7 days, we assessed local sensory response, tissue oxygen consumption, oxygen radicals, and antioxidant levels. In vitro mitochondrial function was measured after stimulation of isolated mitochondria of skeletal muscle cells with increasing doses of tert-BuOOH. tert-BuOOH infusion resulted in an increased skin temperature (p = 0.04), impaired function, and a significantly increased pain sensation (p = 0.03). Venous oxygen saturation levels (p = 0.01) and the antioxidant ceruloplasmin (p = 0.04) were increased. tert-BuOOH inhibited mitochondrial function in vitro. Induction of free radical formation in the rat hind limb results in an exacerbated sensory response and is associated with impaired oxygen extraction, which likely results from mitochondrial dysfunction caused by free radicals

Mitochondria-Targeted Antioxidants SkQ1 and MitoTEMPO Failed to Exert a Long-Term Beneficial Effect in Murine Polymicrobial Sepsis

Mitochondrial-derived reactive oxygen species have been deemed an important contributor in sepsis pathogenesis. We investigated whether two mitochondria-targeted antioxidants (mtAOX; SkQ1 and MitoTEMPO) improved long-term outcome, lessened inflammation, and improved organ homeostasis in polymicrobial murine sepsis. 3-month-old female CD-1 mice (n = 90) underwent cecal ligation and puncture (CLP) and received SkQ1 (5 nmol/kg), MitoTEMPO (50 nmol/kg), or vehicle 5 times post-CLP. Separately, 52 SkQ1-treated CLP mice were sacrificed at 24 h and 48 h for additional endpoints. Neither MitoTEMPO nor SkQ1 exerted any protracted survival benefit. Conversely, SkQ1 exacerbated 28-day mortality by 29%. CLP induced release of 10 circulating cytokines, increased urea, ALT, and LDH, and decreased glucose but irrespectively of treatment. Similar occurred for CLP-induced lymphopenia/neutrophilia and the NO blood release. At 48 h post-CLP, dying mice had approximately 100-fold more CFUs in the spleen than survivors, but this was not SkQ1 related. At 48 h, macrophage and granulocyte counts increased in the peritoneal lavage but irrespectively of SkQ1. Similarly, hepatic mitophagy was not altered by SkQ1 at 24 h. The absence of survival benefit of mtAOX may be due to the extended treatment and/or a relatively moderate-risk-of-death CLP cohort. Long-term effect of mtAOX in abdominal sepsis appears different to sepsis/inflammation models arising from other body compartments

Mitochondria-Targeted Antioxidants SkQ1 and MitoTEMPO Failed to Exert a Long-Term Beneficial Effect in Murine Polymicrobial Sepsis

Mitochondrial-derived reactive oxygen species have been deemed an important contributor in sepsis pathogenesis. We investigated whether two mitochondria-targeted antioxidants (mtAOX; SkQ1 and MitoTEMPO) improved long-term outcome, lessened inflammation, and improved organ homeostasis in polymicrobial murine sepsis. 3-month-old female CD-1 mice (n=90) underwent cecal ligation and puncture (CLP) and received SkQ1 (5 nmol/kg), MitoTEMPO (50 nmol/kg), or vehicle 5 times post-CLP. Separately, 52 SkQ1-treated CLP mice were sacrificed at 24 h and 48 h for additional endpoints. Neither MitoTEMPO nor SkQ1 exerted any protracted survival benefit. Conversely, SkQ1 exacerbated 28-day mortality by 29%. CLP induced release of 10 circulating cytokines, increased urea, ALT, and LDH, and decreased glucose but irrespectively of treatment. Similar occurred for CLP-induced lymphopenia/neutrophilia and the NO blood release. At 48 h post-CLP, dying mice had approximately 100-fold more CFUs in the spleen than survivors, but this was not SkQ1 related. At 48 h, macrophage and granulocyte counts increased in the peritoneal lavage but irrespectively of SkQ1. Similarly, hepatic mitophagy was not altered by SkQ1 at 24 h. The absence of survival benefit of mtAOX may be due to the extended treatment and/or a relatively moderate-risk-of-death CLP cohort. Long-term effect of mtAOX in abdominal sepsis appears different to sepsis/inflammation models arising from other body compartments

Minimum Quality Threshold in Pre-Clinical Sepsis Studies (MQTiPSS): an international expert consensus initiative for improvement of animal modeling in sepsis

Purpose: Pre-clinical animal studies precede the majority of clinical trials. While the clinical sepsis definitions and recommended treatments are regularly updated, a systematic review of pre-clinical models of sepsis has not been done and clear modeling guidelines are lacking. To address this deficit, a Wiggers-Bernard Conference on pre-clinical sepsis modeling was held in Vienna in May, 2017. The conference goal was to identify limitations of pre-clinical sepsis models and to propose a set of guidelines, defined as the “Minimum Quality Threshold in Pre-Clinical Sepsis Studies” (MQTiPSS), to enhance translational value of these models. Methods: 31 experts from 13 countries participated and were divided into 6 thematic Working Groups (WG): (1) Study Design, (2) Humane modeling, (3) Infection types, (4) Organ failure/dysfunction, (5) Fluid resuscitation and (6) Antimicrobial therapy endpoints. As basis for the MQTiPSS discussions, the participants conducted a literature review of the 260 most highly cited scientific articles on sepsis models (2002–2013). Results: Overall, the participants reached consensus on 29 points; 20 at “recommendation” (R) and 9 at “consideration” (C) strength. This Executive Summary provides a synopsis of the MQTiPSS consensus (Tables 1, 2 and 3). Conclusions: We believe that these recommendations and considerations will serve to bring a level of standardization to pre-clinical models of sepsis and ultimately improve translation of pre-clinical findings. These guideline points are proposed as “best practices” that should be implemented for animal sepsis models. In order to encourage its wide dissemination, this article is freely accessible in Shock, Infection and Intensive Care Medicine Experimental