Long-term engraftment and maturation of autologous iPSC-derived cardiomyocytes in two rhesus macaques.

Cellular therapies with cardiomyocytes produced from induced pluripotent stem cells (iPSC-CMs) offer a potential route to cardiac regeneration as a treatment for chronic ischemic heart disease. Here, we report successful long-term engraftment and in vivo maturation of autologous iPSC-CMs in two rhesus macaques with small, subclinical chronic myocardial infarctions, all without immunosuppression. Longitudinal positron emission tomography imaging using the sodium/iodide symporter (NIS) reporter gene revealed stable grafts for over 6 and 12 months, with no teratoma formation. Histological analyses suggested capability of the transplanted iPSC-CMs to mature and integrate with endogenous myocardium, with no sign of immune cell infiltration or rejection. By contrast, allogeneic iPSC-CMs were rejected within 8 weeks of transplantation. This study provides the longest-term safety and maturation data to date in any large animal model, addresses concerns regarding neoantigen immunoreactivity of autologous iPSC therapies, and suggests that autologous iPSC-CMs would similarly engraft and mature in human hearts.

Asthmatic patients with high serum amyloid A have proinflammatory HDL: Implications for augmented systemic and airway inflammation.

RATIONALE: Serum amyloid A (SAA) is bound to high-density lipoproteins (HDL) in blood. Although SAA is increased in the blood of patients with asthma, it is not known whether this modifies asthma severity. OBJECTIVE: We sought to define the clinical characteristics of patients with asthma who have high SAA levels and assess whether HDL from SAA-high patients with asthma is proinflammatory. METHODS: SAA levels in serum from subjects with and without asthma were quantified by ELISA. HDLs isolated from subjects with asthma and high SAA levels were used to stimulate human monocytes and were intravenously administered to BALB/c mice. RESULTS: An SAA level greater than or equal to 108.8 µg/mL was defined as the threshold to identify 11% of an asthmatic cohort (n = 146) as being SAA-high. SAA-high patients with asthma were characterized by increased serum C-reactive protein, IL-6, and TNF-α; older age; and an increased prevalence of obesity and severe asthma. HDL isolated from SAA-high patients with asthma (SAA-high HDL) had an increased content of SAA as compared with HDL from SAA-low patients with asthma and induced the secretion of IL-6, IL-1ß, and TNF-α from human monocytes via a formyl peptide receptor 2/ATP/P2X purinoceptor 7 axis. Intravenous administration to mice of SAA-high HDL, but not normal HDL, induced systemic inflammation and amplified allergen-induced neutrophilic airway inflammation and goblet cell metaplasia. CONCLUSIONS: SAA-high patients with asthma are characterized by systemic inflammation, older age, and an increased prevalence of obesity and severe asthma. HDL from SAA-high patients with asthma is proinflammatory and, when intravenously administered to mice, induces systemic inflammation, and amplifies allergen-induced neutrophilic airway inflammation. This suggests that systemic inflammation induced by SAA-high HDL may augment disease severity in asthma.

Apolipoprotein E is a concentration-dependent pulmonary danger signal that activates the NLRP3 inflammasome and IL-1ß secretion by bronchoalveolar fluid macrophages from asthmatic subjects.

BACKGROUND: House dust mite (HDM)-challenged Apoe-/- mice display enhanced airway hyperreactivity and mucous cell metaplasia. OBJECTIVE: We sought to characterize the pathways that induce apolipoprotein E (APOE) expression by bronchoalveolar lavage fluid (BALF) macrophages from asthmatic subjects and identify how APOE regulates IL-1ß secretion. METHODS: Macrophages were isolated from asthmatic BALF and derived from THP-1 cells and human monocytes. RESULTS: HDM-derived cysteine and serine proteases induced APOE secretion from BALF macrophages through protease-activated receptor 2. APOE at concentrations of less than 2.5 nmol/L, which are similar to levels found in epithelial lining fluid from healthy adults, did not induce IL-1ß release from BALF macrophages. In contrast, APOE at concentrations of 25 nmol/L or greater induced nucleotide-binding oligomerization domain, leucine-rich repeat-containing protein (NLRP) 3 and pro-IL-1ß expression by BALF macrophages, as well as the caspase-1-mediated generation of mature IL-1ß secreted from cells. HDM acted synergistically with APOE to both prime and activate the NLRP3 inflammasome. In a murine model of neutrophilic airway inflammation induced by HDM and polyinosinic-polycytidylic acid, APOE reached a concentration of 32 nmol/L in epithelial lining fluid, with associated increases in BALF IL-1ß levels. APOE-dependent NLRP3 inflammasome activation in macrophages was primarily mediated through a potassium efflux-dependent mechanism. CONCLUSION: APOE can function as an endogenous, concentration-dependent pulmonary danger signal that primes and activates the NLPR3 inflammasome in BALF macrophages from asthmatic subjects to secrete IL-1ß. This might represent a mechanism through which APOE amplifies pulmonary inflammatory responses when concentrations in the lung are increased to greater than normal levels, which can occur during viral exacerbations of HDM-induced asthma characterized by neutrophilic airway inflammation.

Low-density lipoprotein receptor-related protein 1 attenuates house dust mite-induced eosinophilic airway inflammation by suppressing dendritic cell-mediated adaptive immune responses.

BACKGROUND: Low-density lipoprotein receptor-related protein 1 (LRP-1) is a scavenger receptor that regulates adaptive immunity and inflammation. LRP-1 is not known to modulate the pathogenesis of allergic asthma. OBJECTIVE: We sought to assess whether LRP-1 expression by dendritic cells (DCs) modulates adaptive immune responses in patients with house dust mite (HDM)-induced airways disease. METHODS: LRP-1 expression on peripheral blood DCs was quantified by using flow cytometry. The role of LRP-1 in modulating HDM-induced airways disease was assessed in mice with deletion of LRP-1 in CD11c+ cells (Lrp1fl/fl; CD11c-Cre) and by adoptive transfer of HDM-pulsed CD11b+ DCs from Lrp1fl/fl; CD11c-Cre mice to wild-type (WT) mice. RESULTS: Human peripheral blood myeloid DC subsets from patients with eosinophilic asthma have lower LRP-1 expression than cells from healthy nonasthmatic subjects. Similarly, LRP-1 expression by CD11b+ lung DCs was significantly reduced in HDM-challenged WT mice. HDM-challenged Lrp1fl/fl; CD11c-Cre mice have a phenotype of increased eosinophilic airway inflammation, allergic sensitization, TH2 cytokine production, and mucous cell metaplasia. The adoptive transfer of HDM-pulsed LRP-1-deficient CD11b+ DCs into WT mice generated a similar phenotype of enhanced eosinophilic inflammation and allergic sensitization. Furthermore, CD11b+ DCs in the lungs of Lrp1fl/fl; CD11c-Cre mice have an increased ability to take up HDM antigen, whereas bone marrow-derived DCs display enhanced antigen presentation capabilities. CONCLUSION: This identifies a novel role for LRP-1 as a negative regulator of DC-mediated adaptive immune responses in the setting of HDM-induced eosinophilic airway inflammation. Furthermore, the reduced LRP-1 expression by circulating myeloid DCs in patients with eosinophilic asthma suggests a possible role for LRP-1 in modulating type 2-high asthma.

A Chronic Cardiac Ischemia Model in Swine Using an Ameroid Constrictor.

Cardiovascular disease remains the number one cause of mortality in the United States. There are numerous approaches to treating these diseases, but regardless of the approach, an in vivo model is needed to test each treatment. The pig is one of the most used large animal models for cardiovascular disease. Its heart is very similar in anatomy and function to that of a human. The ameroid placement technique creates an ischemic area of the heart, which has many useful applications in studying myocardial infarction. This model has been used for surgical research, pharmaceutical studies, imaging techniques, and cell therapies. There are several ways of inducing an ischemic area in the heart. Each has its advantages and disadvantages, but the placement of an ameroid constrictor remains the most widely used technique. The main advantages to using the ameroid are its prevalence in existing research, its availability in various sizes to accommodate the anatomy and size of the vessel to be constricted, the surgery is a relatively simple procedure, and the post-operative monitoring is minimal, since there are no external devices to maintain. This paper provides a detailed overview of the proper technique for the placement of the ameroid constrictor.