Aerosol Biologics for the Treatment of Eosinophilic Asthma

Asthma is a heterogeneous chronic airway disease comprising of distinct phenotypes characterized by different immunopathophysiologic pathways, clinical features, disease severity, and response to treatment. The phenotypes of asthma include eosinophilic, neutrophilic, mixed cellularity, and paucigranulocytic asthma. Eosinophilic asthma is principally a T helper type 2 (Th2)-mediated airway disease. However, several other immune and structural cells secrete the cytokines implicated in the pathogenesis of eosinophilic asthma. Innate type 2 lymphoid cells, mast cells, basophils, and eosinophils secrete Th2 cytokines, such as interleukin-4 (IL-4), IL-13, and IL-5. Additionally, airway epithelial cells produce alarmin cytokines, including IL-25, IL-33, and thymic stromal lymphopoietin (TSLP). Alarmins are the key initiators of allergic inflammation at the sentinel mucosal surfaces. Innovative biotherapeutic research has led to the discovery of monoclonal antibodies which target and inhibit the immunopathological effects of the cytokines involved in the pathogenesis of eosinophilic asthma. Parenteral biologics targeting the inciting interleukins, include mepolizumab and reslizumab (anti-IL-5), benralizumab (anti-IL-5Rα), dupilumab (anti-4Rα), and tezelizumab (anti-TSLP). They have been shown to significantly reduce annualized exacerbation rates, improve asthma control, lung function, and quality of life. Currently, there are no pulmonary delivered aerosol biologics for topical treatment of asthma. CSJ117 is a potent neutralizing antibody Fab fragment against TSLP, formulated as a PulmoSol TM engineered powder, and is delivered to the lungs by a dry powder inhaler. Phase 2 placebo-controlled clinical trial evaluated the efficacy and safety of CSJ117. CSJ117 delivered as an inhaler attenuated the late asthmatic response (LAR), and the early asthmatic response (EAR) after allergen inhalation challenge (AIC) at day 84 of treatment. The maximum decrease in FVE1 from pre-AIC were significantly lower in the CSJ117 group compared to placebo (P = 029), during LAR. CSJ117 also significantly reduced fractional exhaled nitric oxide before AIC at day 83; and significantly reduced the allergen-induced increase in % sputum eosinophil count. Pulmonary delivery of biologics directly to the airway mucosal surface has several advantages over parenteral routes, particularly in treating airway diseases such as asthma. Inhaler delivered biologics, such as CSJ117 are innovative and attractive methods of future precision treatment of asthma, and other respiratory diseases.

An immunologically active, adipose-derived extracellular matrix biomaterial for soft tissue reconstruction: concept to clinical trial

Soft tissue reconstruction remains an intractable clinical challenge as current surgical options and synthetic implants may produce inadequate outcomes. Soft tissue deficits may be surgically reconstructed using autologous adipose tissue, but these procedures can lead to donor site morbidity, require multiple procedures, and have highly variable outcomes. To address this clinical need, we developed an “off-the-shelf” adipose extracellular matrix (ECM) biomaterial from allograft human tissue (Acellular Adipose Tissue, AAT). We applied physical and chemical processing methods to remove lipids and create an injectable matrix that mimicked the properties of lipoaspirate. Biological activity was assessed using cell migration and adipogenesis assays. Characterization of regenerative immune properties in a murine muscle injury model revealed that allograft and xenograft AAT induced pro-regenerative CD4+ T cells and macrophages with xenograft AAT additionally attracting eosinophils secreting interleukin 4 (Il4). In immunocompromised mice, AAT injections retained similar volumes as human fat grafts but lacked cysts and calcifications seen in the fat grafts. The combination of AAT with human adipose-derived stem cells (ASCs) resulted in lower implant volumes. However, tissue remodeling and adipogenesis increased significantly in combination with ASCs. Larger injected volumes of porcine-derived AAT demonstrated biocompatibility and greater retention when applied allogeneicly in Yorkshire cross pigs. AAT was implanted in healthy volunteers in abdominal tissue that was later removed by elective procedures. AAT implants were well tolerated in all human subjects. Implants removed between 1 and 18 weeks demonstrated increasing cellular infiltration and immune populations, suggesting continued tissue remodeling and the potential for long-term tissue replacement.

HLA-G in Allergy: Does It Play an Immunoregulatory Role?

Allergy is an inflammatory process determined by a cascade of immune events characterized by T-helper 2 lymphocytes polarization leading to interleukin-4 upregulation, IgE secretion, and mast cell and eosinophil activation. HLA-G molecules, both in membrane-bound and in soluble forms, are known to play a key immunoregulatory role and their involvement in allergic diseases is supported by increasing literature data. HLA-G expression and secretion is specifically induced in peripheral blood mononuclear cells of allergic patients after in vitro incubation with the causal allergen. Elevated levels of soluble HLA-G molecules are detected in serum of patients with allergic rhinitis correlating with allergen-specific IgE levels, clinical severity, drug consumption and response to allergen-specific immunotherapy. HLA-G genetic polymorphisms confer susceptibility to allergic asthma development and high levels of soluble HLA-G molecules are found in plasma and bronchoalveolar lavage fluid of patients with allergic asthma correlating with allergen-specific IgE levels. Interestingly, allergic pregnant women have lower plasma sHLA-G levels than non-allergic women during the 3rd trimester of pregnancy and at delivery. Finally, in allergic patients with atopic dermatitis HLA-G molecules are expressed by T cells, monocytes-macrophages and Langerhans cells infiltrating the dermis. Although at present is difficult to completely define the role of HLA-G molecules in allergic diseases, it may be suggested that they are specifically expressed and secreted by immune cells during the allergic reaction in an attempt to suppress allergic inflammation.

Kinetics of LYVE-1-Positive M2-Like Macrophages in Developing and Repairing Dental Pulp in Vivo and Their Pro-Angiogenic Activity in Vitro

Tissue-resident macrophages expressing lymphatic vessel endothelial hyaluronan receptor-1 (LYVE-1) are found in multiple tissues and organs. We aimed to evaluate the dynamics and biological function of LYVE-1+ macrophages in dental pulp during post-injury tissue remodeling. Immunofluorescence staining of mouse embryos revealed that LYVE-1+ macrophages colonized dental pulp before birth. In mature rat molar dental pulp, LYVE-1+ macrophages were the main subset of macrophages expressing CD163, an M2 marker, and were distributed throughout the tissue. In response to dental pulp injury induced by cavity preparation, LYVE-1+ macrophages quickly disappeared from the affected area of the pulp and gradually repopulated during the wound healing process. RAW264.7 mouse macrophages cultured with a mixture of macrophage colony-stimulating factor, interleukin-4, and dexamethasone increased LYVE-1 expression, whereas lipopolysaccharide-stimulation decreased LYVE-1 expression. Enforced expression of Lyve1 in RAW264.7 cells resulted in increased mRNA expression of matrix metalloproteinase 2 (Mmp2), Mmp9, and vascular endothelial growth factor A (Vegfa). Lyve1-expressing macrophages promoted the migration and tube formation of human umbilical vein endothelial cells. In conclusion, LYVE-1+ tissue-resident M2-like macrophages in dental pulp showed dynamism in response to pulp injury, and possibly play an important role in angiogenesis during wound healing and tissue remodeling.

Characterization of the Pathology, Biochemistry, and Immune Response in Kunming (KM) Mice Following Fasciola gigantica Infection

As a putative model of Fasciola gigantica infection, detailed data in Kunming (KM) mice infected with F. gigantica are lacking. In this study, KM mice were orally infected with 15 metacercaria for 8 weeks. Macroscopic and microscopic changes, serum biochemistry, cytokine responses, and changes in parasite-specific immunoglobulin G (IgG) antibody levels were monitored at 1, 3, 5, 7, and 8 weeks post-infection (wpi), respectively. The serum levels of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) increased after infection, while that of albumin (ALB) decreased, which was positively correlated with the degree of liver damage. Between 5 and 7 wpi, the mice showed symptoms of anemia and weight loss, possibly caused by the decrease of alkaline phosphatase (ALP). Moreover, the changing tendencies of the levels of globulin (GLB) and parasite-specific IgG antibody were similar, suggesting a potential correlation between GLB production and adaptive immune response in the host. Coordinated variations in interferon gamma (IFN-γ) and interleukin 4 (IL-4) indicated a mixed T helper 1 (Th1)/Th2 cellular immune response. Furthermore, the serum IgG antibody increased after infection and peaked at 5 wpi, and it was positively correlated with the average parasite burdens. The worms collected from mice were approximately 1 cm in length at 8 wpi, their digestive and reproductive systems were well developed, and no eggs were found in the uterus. To the best of our knowledge, this is the first report describing detailed histological, biochemical, and immunological indices in KM mice infected with F. gigantica, which provides basic information on KM mice against infection with F. gigantica.

Interleukin-4 Aggravates LPS-Induced Striatal Neurodegeneration In Vivo via Oxidative Stress and Polarization of Microglia/Macrophages

The present study investigated the effects of interleukin (IL)-4 on striatal neurons in lipopolysaccharide (LPS)-injected rat striatum in vivo. Either LPS or PBS as a control was unilaterally injected into the striatum, and brain tissues were processed for immunohistochemical and Nissl staining or for hydroethidine histochemistry at the indicated time points after LPS injection. Analysis by NeuN and Nissl immunohistochemical staining showed a significant loss of striatal neurons at 1, 3, and 7 days post LPS. In parallel, IL-4 immunoreactivity was upregulated as early as 1 day, reached a peak at 3 days, and was sustained up to 7 days post LPS. Increased levels of IL-4 immunoreactivity were exclusively detected in microglia/macrophages, but not in neurons nor astrocytes. The neutralizing antibody (NA) for IL-4 significantly protects striatal neurons against LPS-induced neurotoxicity in vivo. Accompanying neuroprotection, IL-4NA inhibited activation of microglia/macrophages, production of reactive oxygen species (ROS), ROS-derived oxidative damage and nitrosative stress, and produced polarization of microglia/macrophages shifted from M1 to M2. These results suggest that endogenous IL-4 expressed in LPS-activated microglia/macrophages contributes to striatal neurodegeneration in which oxidative/nitrosative stress and M1/M2 polarization are implicated.

Perilla Leaf Extract (PLE) Attenuates COPD Airway Inflammation via the TLR4/Syk/PKC/NF-κB Pathway In Vivo and In Vitro

Chronic obstructive pulmonary disease (COPD) is a complex and heterogeneous disease characterized by persistent airflow limitation but still lacking effective treatments. Perilla frutescens (L.) Britt., an important traditional medicinal plant with excellent antioxidant and anti-inflammatory properties, is widely used for the treatment of respiratory disease in China. However, its protective activity and mechanism against COPD airway inflammation have not been fully studied. Here, the anti-inflammatory effects of the PLE were investigated, and its underlying mechanisms were then elucidated. The presented results suggested a notable effect of the PLE on airway inflammation of COPD, by significantly ameliorating inflammatory cell infiltration in lung tissue, lessening leukocytes (lymphocytes, neutrophils, and macrophages) and inflammatory mediators (interleukin 4 (IL-4), IL-6, IL-17A, interferon γ (IFN-γ), and tumor necrosis factor α (TNF-α)) in the bronchoalveolar lavage fluid (BALF) of cigarette smoke (CS)/lipopolysaccharide (LPS)-induced COPD mice in vivo and inhibiting the production of inflammatory factors (nitric oxide (NO), IL-6, and TNF-α) and intracellular reactive oxygen species (ROS) in LPS-stimulated RAW264.7 cells in vitro. For further extent, PLE treatment significantly suppressed the expression and phosphorylation of TLR4, Syk, PKC, and NF-κB p65 in vivo and their mRNA in vitro. Subsequently, by co-treating with their inhibitors in vitro, its potential mechanism via TLR4/Syk/PKC/NF-κB p65 signals was disclosed. In summary, the obtained results indicated a noteworthy effective activity of the PLE on COPD inflammation, and partly, the TLR4/Syk/PKC/NF-κB p65 axis might be the potential mechanism.