Abstract A promising strategy to accelerate joint implant integration and reduce recovery time and failure rates is to deliver a combination of certain growth factors to the integration site. There ...is a need to control the quantity of growth factors delivered at different times during the healing process to maximize efficacy. Polyelectrolyte multilayer (PEM) films, built using the layer-by-layer (LbL) technique, are attractive for releasing controlled amounts of potent growth factors over a sustained period. Here, we present PEM films that sequester physiological amounts of osteogenic rhBMP-2 (recombinant human bone morphogenetic protein - 2) and angiogenic rhVEGF165 (recombinant human vascular endothelial growth factor) in different ratios in a degradable poly(β-amino ester)/polyanion/growth factor/polyanion LbL tetralayer repeat architecture where the biologic load scaled linearly with the number of tetralayers. No burst release of either growth factor was observed as the films degraded. The release of rhBMP-2 was sustained over a period of 2 weeks, while rhVEGF165 eluted from the film over the first 8 days. Both growth factors retained their efficacy, as quantified with relevant in vitro assays. rhBMP-2 initiated a dose dependent differentiation cascade in MC3T3-E1S4 pre-osteoblasts while rhVEGF165 upregulated HUVEC proliferation, and accelerated closure of a scratch in HUVEC cell cultures in a dose dependent manner. In vivo , the mineral density of ectopic bone formed de novo by rhBMP-2/rhVEGF165 PEM films was approximately 33% higher than when only rhBMP-2 was introduced, with a higher trabecular thickness, which would indicate a decrease in the risk of osteoporotic fracture. Bone formed throughout the scaffold when both growth factors were released, which suggests more complete remodeling due to an increased local vascular network. This study demonstrates a promising approach to delivering precise doses of multiple growth factors for a variety of implant applications where control over spatial and temporal release profile of the biologic is desired.
RATIONALE:The rapid induction and orchestration of new blood vessels are critical for tissue repair in response to injury, such as myocardial infarction, and for physiological angiogenic responses, ...such as embryonic development and exercise.
OBJECTIVE:We aimed to identify and characterize microRNAs (miR) that regulate pathological and physiological angiogenesis.
METHODS AND RESULTS:We show that miR-26a regulates pathological and physiological angiogenesis by targeting endothelial cell (EC) bone morphogenic protein/SMAD1 signaling in vitro and in vivo. MiR-26a expression is increased in a model of acute myocardial infarction in mice and in human subjects with acute coronary syndromes. Ectopic expression of miR-26a markedly induced EC cycle arrest and inhibited EC migration, sprouting angiogenesis, and network tube formation in matrigel, whereas blockade of miR-26a had the opposite effects. Mechanistic studies demonstrate that miR-26a inhibits the bone morphogenic protein/SMAD1 signaling pathway in ECs by binding to the SMAD1 3′-untranslated region, an effect that decreased expression of Id1 and increased p21 and p27. In zebrafish, miR-26a overexpression inhibited formation of the caudal vein plexus, a bone morphogenic protein-responsive process, an effect rescued by ectopic SMAD1 expression. In mice, miR-26a overexpression inhibited EC SMAD1 expression and exercise-induced angiogenesis. Furthermore, systemic intravenous administration of an miR-26a inhibitor, locked nucleic acid-anti–miR-26a, increased SMAD1 expression and rapidly induced robust angiogenesis within 2 days, an effect associated with reduced myocardial infarct size and improved heart function.
CONCLUSIONS:These findings establish miR-26a as a regulator of bone morphogenic protein/SMAD1-mediated EC angiogenic responses, and that manipulating miR-26a expression could provide a new target for rapid angiogenic therapy in ischemic disease states.
The use of mesenchymal stromal cells (MSCs) for treatment of bacterial infections, including systemic processes like sepsis, is an evolving field of investigation. This study was designed to ...investigate the potential use of MSCs, harvested from compact bone, and their interactions with the innate immune system, during polymicrobial sepsis induced by cecal ligation and puncture (CLP). We also wanted to elucidate the role of endogenous heme oxygenase (HO)‐1 in MSCs during a systemic bacterial infection. MSCs harvested from the bones of HO‐1 deficient (−/−) and wild‐type (+/+) mice improved the survival of HO‐1−/− and HO‐1+/+ recipient mice when administered after the onset of polymicrobial sepsis induced by CLP, compared with the administration of fibroblast control cells. The MSCs, originating from compact bone in mice, enhanced the ability of neutrophils to phagocytize bacteria in vitro and in vivo and to promote bacterial clearance in the peritoneum and blood after CLP. Moreover, after depleting neutrophils in recipient mice, the beneficial effects of MSCs were entirely lost, demonstrating the importance of neutrophils for this MSC response. MSCs also decreased multiple organ injury in susceptible HO‐1−/− mice, when administered after the onset of sepsis. Taken together, these data demonstrate that the beneficial effects of treatment with MSCs after the onset of polymicrobial sepsis is not dependent on endogenous HO‐1 expression, and that neutrophils are crucial for this therapeutic response. STEM CELLS2013;31:397–407
Current clinical products delivering the osteogenic growth factor bone morphogenetic protein 2 (BMP-2) for bone regeneration have been plagued by safety concerns due to a high incidence of off-target ...effects resulting from bolus release and supraphysiological doses. Layer-by-layer (LbL) film deposition offers the opportunity to coat bone defect-relevant substrates with thin films containing proteins and other therapeutics; however, control of release kinetics is often hampered by interlayer diffusion of drugs throughout the film during assembly, which causes burst drug release. In this work, we present the design of different laponite clay diffusional barrier layer architectures in self-assembled LbL films to modulate the release kinetics of BMP-2 from the surface of a biodegradable implant. Release kinetics were tuned by incorporating laponite in different film arrangements and with varying deposition techniques to achieve release of BMP-2 over 2 days, 4 days, 14 days, and 30 days. Delivery of a low dose (0.5 μg) of BMP-2 over 2 days and 30 days using these LbL film architectures was then compared in an in vivo rat critical size calvarial defect model to determine the effect of BMP-2 release kinetics on bone regeneration. After 6 weeks, sustained release of BMP-2 over 30 days induced 3.7 times higher bone volume and 7.4 times higher bone mineral density as compared with 2-day release of BMP-2, which did not induce more bone growth than the uncoated scaffold control. These findings represent a crucial step in the understanding of how BMP-2 release kinetics influence treatment efficacy and underscore the necessity to optimize protein delivery methods in clinical formulations for bone regeneration. This work could be applied to the delivery of other therapeutic proteins for which careful tuning of the release rate is a key optimization parameter.
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•Clinical osteogenic protein formulations exhibit burst release of excessive doses.•Assembly of protein-containing films via layer-by-layer deposition enables controlled release.•Incorporation of diffusional barrier layers enables release over 2, 4, 14, or 30 days.•An extended-release formulation enhances bone healing over a rapid release formulation.
The 2021 WHO Classification of Tumors of the Heart Maleszewski, Joseph J.; Basso, Cristina; Bois, Melanie C. ...
Journal of thoracic oncology,
April 2022, 2022-04-00, 20220401, Letnik:
17, Številka:
4
Journal Article
A variety of infectious agents, including varicella zoster virus (VZV), have been hypothesized to play a role in the pathogenesis of giant cell arteritis (GCA). The detectability of the virus in ...patients with GCA is debatable. To further investigate an association between GCA and VZV infection, 10 years of GCA cases were evaluated for VZV by immunohistochemistry (IHC).
All temporal artery biopsies and ascending aortic resections positive for GCA from 2007 to 2017 at Brigham and Women's Hospital were immunostained using a VZV antibody cocktail (SG1-1, SG1-SG4, NCP-1, and IE-62).
Forty-one temporal artery biopsies and 47 ascending aortic resections positive for GCA were identified, all of which were found to be negative for VZV by IHC. Twelve temporal artery biopsies in this cohort were previously analyzed by unbiased metagenomics sequencing and were negative for VZV DNA.
These results argue against a clinically relevant association between VZV infection and GCA, and support neither routine testing for VZV nor treatment with antiviral drugs.
Somatic mutations that activate phosphoinositide 3-kinase (PI3K) have been identified in the p110-α catalytic subunit (encoded by PIK3CA). They are most frequently observed in two hotspots: the ...helical domain (E545K and E542K) and the kinase domain (H1047R). Although the p110-α mutants are transforming in vitro, their oncogenic potential has not been assessed in genetically engineered mouse models. Furthermore, clinical trials with PI3K inhibitors have recently been initiated, and it is unknown if their efficacy will be restricted to specific, genetically defined malignancies. In this study, we engineered a mouse model of lung adenocarcinomas initiated and maintained by expression of p110-α H1047R. Treatment of these tumors with NVP-BEZ235, a dual pan-PI3K and mammalian target of rapamycin (mTOR) inhibitor in clinical development, led to marked tumor regression as shown by positron emission tomography-computed tomography, magnetic resonance imaging and microscopic examination. In contrast, mouse lung cancers driven by mutant Kras did not substantially respond to single-agent NVP-BEZ235. However, when NVP-BEZ235 was combined with a mitogen-activated protein kinase kinase (MEK) inhibitor, ARRY-142886, there was marked synergy in shrinking these Kras-mutant cancers. These in vivo studies suggest that inhibitors of the PI3K-mTOR pathway may be active in cancers with PIK3CA mutations and, when combined with MEK inhibitors, may effectively treat KRAS mutated lung cancers.
Celotno besedilo
Dostopno za:
DOBA, IJS, IZUM, KILJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
Objective We compared the histologic findings in explanted CorMatrix (9 patients) and autologous pericardium (9 patients) used for valvuloplasty of the aortic (7 patients) and/or mitral (11 patients) ...valve in patients with congenital heart defects. Methods We used standard tissue stains and immunohistochemistry to identify the inflammatory cell type. Results CorMatrix was associated with an intense inflammatory response in the surrounding native tissue, extending into CorMatrix in 8 of 9 cases, continuing to the longest follow-up point (9 months). The typical response included macrophages and giant cells in contact with the material, surrounded by lymphocytes, macrophages, plasma cells, and eosinophils. The thickness of the residual CorMatrix material was 280 to 300 μm, similar to the nominal thickness at implantation and unrelated to the implantation duration. Only at the longest follow-up interval was any significant resorption of CorMatrix material evident. A neointima had formed on the surface of CorMatrix, increasing in thickness with the period in situ. Mild cellular infiltration of CorMatrix was noted in all cases; however, in no case, did it appear that CorMatrix was being remodeled into tissue resembling a 3-layered native valve. In contrast, a near absence of any inflammatory reaction was seen and no eosinophilia associated with autologous pericardium was present, irrespective of the duration in situ. Furthermore, we observed more tissue infiltration, remodeling, vascularization, and neointima formation with autologous pericardium. Conclusions Although CorMatrix used for valve repair induced an intense inflammatory response, little or no remodeling to form tissue resembling a 3-layered native valve was seen at ≤9 months after implantation.
Many studies have been performed in severe COVID-19 on immune cells in the circulation and on cells obtained by bronchoalveolar lavage. Most studies have tended to provide relative information rather ...than a quantitative view, and it is a combination of approaches by various groups that is helping the field build a picture of the mechanisms that drive severe lung disease. Approaches employed to date have not revealed information on lung parenchymal T cell subsets in severe COVID-19. Therefore, we sought to examine early and late T cell subset alterations in the lungs and draining lymph nodes in severe COVID-19 using a rapid autopsy protocol and quantitative imaging approaches. Here, we have established that cytotoxic CD4+ T cells (CD4 + CTLs) increase in the lungs, draining lymph nodes and blood as COVID-19 progresses. CD4 + CTLs are prominently expanded in the lung parenchyma in severe COVID-19. In contrast CD8+ T cells are not prominent, exhibit increased PD-1 expression, and no obvious increase is seen in the number of Granzyme B+ CD8+ T cells in the lung parenchyma in severe COVID-19. Based on quantitative evidence for re-activation in the lung milieu, CD4 + CTLs may be as likely to drive viral clearance as CD8+ T cells and may also be contributors to lung inflammation and eventually to fibrosis in severe COVID-19.
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•CD4 + CTLs were the only T cell subset that were significantly expanded in the lungs in severe COVID-19.•The majority of CD8+ T cells in the lungs in severe COVID-19 express PD-1.•Re-activated GZMB+ CD8 + CTLs and GZMB+ CD4 + CTLs were found in equivalent proportions in the lungs.•CD4 + CTLs were prominent in the circulation, were SARS-CoV-2 specific and this expansion was linked to disease severity.
Severe viral pneumonia caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is characterized by a hyperinflammatory state typified by elevated circulating pro-inflammatory ...cytokines, frequently leading to potentially lethal vascular complications including thromboembolism, disseminated intracellular coagulopathy and vasculitis. Though endothelial infection and subsequent endothelial damage have been described in patients with fatal COVID-19, the mechanism by which this occurs remains elusive, particularly given that, under naïve conditions, pulmonary endothelial cells demonstrate minimal cell surface expression of the SARS-CoV-2 binding receptor ACE2. Herein we describe SARS-CoV-2 infection of the pulmonary endothelium in postmortem lung samples from individuals who died of COVID-19, demonstrating both heterogeneous ACE2 expression and endothelial damage. In primary endothelial cell cultures, we show that SARS-CoV-2 infection is dependent on the induction of ACE2 protein expression and that this process is facilitated by type 1 interferon-alpha (IFNα) or -beta(β)—two of the main anti-viral cytokines induced in severe SARS-CoV-2 infection—but not significantly by other cytokines (including interleukin 6 and interferon γ/λ). Our findings suggest that the stereotypical anti-viral interferon response may paradoxically facilitate the propagation of COVID-19 from the respiratory epithelium to the vasculature, raising concerns regarding the use of exogenous IFNα/β in the treatment of patients with COVID-19.
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