HIV infection impairs the host response to Mycobacterium tuberculosis infection by altering surfactant protein D function in the human lung alveolar mucosa.

Tuberculosis is the leading cause of death for people living with HIV (PLWH). We hypothesized that altered functions of innate immune components in the human alveolar lining fluid of PLWH (HIV-ALF) drive susceptibility to Mycobacterium tuberculosis (M.tb) infection. Our results indicate a significant increase in oxidation of innate proteins and chemokine levels and significantly lower levels and function of complement components and Th1/Th2/Th17 cytokines in HIV-ALF versus control-ALF (non-HIV-infected people). We further found a deficiency of surfactant protein D (SP-D) and reduced binding of SP-D to M.tb that had been exposed to HIV-ALF. Primary human macrophages infected with M.tb exposed to HIV-ALF were significantly less capable of controlling the infection, which was reversed by SP-D replenishment in HIV-ALF. Thus, based on the limited number of participants in this study, our data suggest that PLWH without antiretroviral therapy (ART) have declining host innate defense function in their lung mucosa, thereby favoring M.tb and potentially other pulmonary infections.

Histone H4 potentiates neutrophil inflammatory responses to influenza A virus: Down-modulation by H4 binding to C-reactive protein and Surfactant protein D.

Neutrophils participate in the early phase of the innate response to uncomplicated influenza A virus (IAV) infection but also are a major component in later stages of severe IAV or COVID 19 infection where neutrophil extracellular traps (NETs) and associated cell free histones are highly pro-inflammatory. It is likely that IAV interacts with histones during infection. We show that histone H4 binds to IAV and aggregates viral particles. In addition, histone H4 markedly potentiates IAV induced neutrophil respiratory burst responses. Prior studies have shown reactive oxidants to be detrimental during severe IAV infection. C reactive protein (CRP) and surfactant protein D (SP-D) rise during IAV infection. We now show that both of these innate immune proteins bind to histone H4 and significantly down regulate respiratory burst and other responses to histone H4. Isolated constructs composed only of the neck and carbohydrate recognition domain of SP-D also bind to histone H4 and partially limit neutrophil responses to it. These studies indicate that complexes formed of histones and IAV are a potent neutrophil activating stimulus. This finding could account for excess inflammation during IAV or other severe viral infections. The ability of CRP and SP-D to bind to histone H4 may be part of a protective response against excessive inflammation in vivo.

Acrolein in cigarette smoke attenuates the innate immune responses mediated by surfactant protein D.

BACKGROUND: Surfactant proteins (SP) A and D belong to collectin family proteins, which play important roles in innate immune response in the lung. We previously demonstrated that cigarette smoke (CS) increases the acrolein modification of SP-A, thereby impairing the innate immune abilities of this protein. In this study, we focused on the effects of CS and its component, acrolein, on the innate immunity role of another collectin, SP-D. METHODS: To determine whether aldehyde directly affects SP-D, we examined the lungs of mice exposed to CS for 1 week and detected aldehyde-modified SP-D using an aldehyde reactive probe. The structural changes in CS extract (CSE) or acrolein-exposed recombinant human (h)SP-D were determined by western blot, liquid chromatography-electrospray ionization tandem mass spectrometry, and blue native-polyacrylamide gel electrophoresis analyses. Innate immune functions of SP-D were determined by bacteria growth and macrophage phagocytosis. RESULTS: Aldehyde-modified SP-D as well as SP-A was detected in the lungs of mice exposed to CS for 1 week. Exposure of hSP-D to CSE or acrolein induced an increased higher-molecular -weight of hSP-D and acrolein induced modification of five lysine residues in hSP-D. These modifications led to disruption of the multimer structure of SP-D and attenuated its ability to inhibit bacterial growth and activate macrophage phagocytosis. CONCLUSION: CS induced acrolein modification in SP-D, which in turn induced structural and functional defects in SP-D. GENERAL SIGNIFICANCE: These results suggest that CS-induced structural and functional defects in SP-D contribute to the dysfunction of innate immune responses in the lung following CS exposure.

Hyaluronic Acid Present in the Tumor Microenvironment Can Negate the Pro-apototic Effect of a Recombinant Fragment of Human Surfactant Protein D on Breast Cancer Cells.

Human surfactant protein D (SP-D) belongs to the family of collectins that is composed of a characteristic amino-terminal collagenous region and a carboxy-terminal C-type lectin domain. Being present at the mucosal surfaces, SP-D acts as a potent innate immune molecule and offers protection against non-self and altered self, such as pathogens, allergens, and tumor. Here, we examined the effect of a recombinant fragment of human SP-D (rfhSP-D) on a range of breast cancer lines. Breast cancer has four molecular subtypes characterized by varied expressions of estrogen (ER), progesterone (PR), and epidermal growth factor (EGF) receptors (HER2). The cell viability of HER2-overexpressing (SKBR3) and triple-positive (BT474) breast cancer cell lines [but not of a triple-negative cell line (BT20)] was reduced following rfhSP-D treatment at 24 h. Upregulation of p21/p27 cell cycle inhibitors and p53 phosphorylation (Ser15) in rfhSP-D-treated BT474 and SKBR3 cell lines signified G2/M cell cycle arrest. Cleaved caspases 9 and 3 were detected in rfhSP-D-treated BT474 and SKBR3 cells, suggesting an involvement of the intrinsic apoptosis pathway. However, rfhSP-D-induced apoptosis was nullified in the presence of hyaluronic acid (HA) whose increased level in breast tumor microenvironment is associated with malignant tumor progression and invasion. rfhSP-D bound to solid-phase HA and promoted tumor cell proliferation. rfhSP-D-treated SKBR3 cells in the presence of HA showed decreased transcriptional levels of p53 when compared to cells treated with rfhSP-D only. Thus, HA appears to negate the anti-tumorigenic properties of rfhSP-D against HER2-overexpressing and triple-positive breast cancer cells.

Berberine-mediated up-regulation of surfactant protein D facilitates cartilage repair by modulating immune responses via the inhibition of TLR4/NF-ĸB signaling.

The innate immune system drives inflammatory joint damage in osteoarthritis (OA) and regulates cartilage repair. Berberine chloride (BBR) is an isoquinoline alkaloid that shows immunomodulatory activity in a variety of cell lines. However, the immunomodulatory mechanisms of BBR in chondrocytes during OA are largely unknown. Herein, we assessed the ability of BBR to mediate chondroprotection through its effects on innate immunity. We found that BBR up-regulated the expression of surfactant protein D (SP-D) in OA cartilage, a key regulator of inflammation and innate immunity both in the airways and extrapulmonary tissues, including joint cartilage. To further explore these findings, we used recombinant adeno-associated virus (rAAV)-mediated knockdown of SP-D. Silencing was assessed in rat model of surgically-induced OA in the presence or absence of BBR treatment, 10 weeks post-surgery. We observed a clear improvement in histological scores of BBR-treated animals compared to those treated with BBR and the rAAV-SP-D vector. In addition, animals co-treated with BBR + recombinant human SP-D (rhSP-D) exhibited significantly lower histological scores than those treated with BBR alone. BBR treatment led to significantly reduced immune cell infiltration mediated through TLR4, F4/80, CD68 and CD34, whilst SP-D silencing reversed this improvement. In contrast, rhSP-D treatment enhanced the protective phenotype. We further explored how BBR influences SP-D and other OA-associated genes in vitro. We observed an up-regulation of SP-D and a marked decline in TRAF6, TLR4, MD-2 and MyD88 expression, as well as NF-κB p65 and IκBα phosphorylation in chondrocytes treated with sodium nitroprusside. siRNAs specific for SP-D were able to partially reverse this phenotype, whilst both rhSP-D and the TLR4 inhibitor TAK-242 enhanced the effects. Together, these results are consistent with a model wherein SP-D has therapeutic potential for OA treatment. Concomitantly, BBR modulates immune responses and decreases cartilage degradation. These findings suggest that BBR achieves this function through releasing SP-D from MD2/SP-D complexes and through the inhibition of TLR4/NF-κB signaling.

Membrane Interactome of a Recombinant Fragment of Human Surfactant Protein D Reveals GRP78 as a Novel Binding Partner in PC3, a Metastatic Prostate Cancer Cell Line.

Surfactant protein-D (SP-D), a member of the collectin family has been shown to induce apoptosis in cancer cells. SP-D is composed of an N-terminal collagen-like domain and a calcium-dependent carbohydrate recognition domain (CRD). Recently, we reported that a recombinant fragment of human SP-D (rfhSP-D), composed of homotrimeric CRD region, induced intrinsic apoptotic pathway in prostate cancer cells. Here, we analyzed the membrane interactome of rfhSP-D in an androgen-independent prostate cancer cell line, PC3, by high resolution mass spectrometry and identified 347 proteins. Computational analysis of PPI network of this interactome in the context of prostate cancer metastasis and apoptosis revealed Glucose Regulated Protein of 78 kDa (GRP78) as an important binding partner of rfhSP-D. Docking studies suggested that rfhSP-D (CRD) bound to the substrate-binding domain of glycosylated GRP78. This was further supported by the observations that human recombinant GRP78 interfered with the binding of rfhSP-D to anti-SP-D polyclonal antibodies; GRP78 also significantly inhibited the binding of recombinant full-length human SP-D with a monoclonal antibody specific to the CRD in a dose-dependent manner. We conclude that the interaction with rfhSP-D is likely to interfere with the pro-survival signaling of GRP78.

Early Identification of Acute Lung Injury in a Porcine Model of Hemorrhagic Shock.

BACKGROUND: Acute lung injury (ALI) is a frequent complication after severe trauma. Lung-protective ventilation strategies and damage control resuscitation have been proposed for the prevention of ALI; however, there are no clinical or laboratory parameters to predict who is at risk of developing ALI after trauma. In the present study, we explored pulmonary inflammatory markers as a potential predictor of ALI using a porcine model of hemorrhagic shock. MATERIALS AND METHODS: Female swine were randomized to mechanical ventilation with low tidal volume (VT) (6 mL/kg) or high VT (12 mL/kg). After equilibration, animals underwent pressure-controlled hemorrhage (mean arterial pressure [MAP] 35 ± 5 mmHg) for 1 h, followed by resuscitation with fresh whole blood or Hextend. They were maintained at MAP of 50 ± 5 mmHg for 3 h in the postresuscitation phase. Bronchoalveolar lavage fluids were collected hourly and analyzed for inflammatory markers. Lung samples were taken, and porcine neutrophil antibody staining was used to evaluate the presence of neutrophils. ELISA evaluated serum porcine surfactant protein D levels. Sham animals were used as negative controls. RESULTS: Pigs that underwent hemorrhagic shock had higher heart rates, lower cardiac output, lower MAPs, and worse acidosis compared with sham at the early time points (P < 0.05 each). There were no significant differences in central venous pressure or pulmonary capillary wedge pressure between groups. Pulmonary neutrophil infiltration, as defined by neutrophil antibody staining on lung samples, was greater in the shock groups regardless of resuscitation fluid (P < 0.05 each). Bronchoalveolar lavage fluid neutrophil levels were not different between groups. There were no differences in levels of porcine surfactant protein D between groups at any time points, and the levels did not change over time in each respective group. CONCLUSIONS: Our study demonstrates the reproducibility of a porcine model of hemorrhagic shock that is consistent with physiologic changes in humans in hemorrhagic shock. Pulmonary neutrophil infiltration may serve as an early marker for ALI; however, the practicality of this finding has yet to be determined.

A Strategy for Suppressing Macrophage-mediated Rejection in Xenotransplantation.

Although xenografts are one of the most attractive strategies for overcoming the shortage of organ donors, cellular rejection by macrophages is a substantial impediment to this procedure. It is well known that macrophages mediate robust immune responses in xenografts. Macrophages also express various inhibitory receptors that regulate their immunological function. Recent studies have shown that the overexpression of inhibitory ligands on porcine target cells results in the phosphorylation of tyrosine residues on intracellular immunoreceptor tyrosine-based inhibitory motifs on macrophages, leading to the suppression of xenogenic rejection by macrophages. It has also been reported that myeloid-derived suppressor cells, a heterogeneous population of immature myeloid cells, suppress not only NK and cytotoxic T lymphocyte cytotoxicity but also macrophage-mediated cytotoxicity. This review is focused on the recent findings regarding strategies for inhibiting xenogenic rejection by macrophages.

Enhanced Antiviral Activity of Human Surfactant Protein D by Site-Specific Engineering of the Carbohydrate Recognition Domain.

Innate immunity is critical in the early containment of influenza A virus (IAV) infection and surfactant protein D (SP-D) plays a crucial role in innate defense against IAV in the lungs. Multivalent lectin-mediated interactions of SP-D with IAVs result in viral aggregation, reduced epithelial infection, and enhanced IAV clearance by phagocytic cells. Previous studies showed that porcine SP-D (pSP-D) exhibits distinct antiviral activity against IAV as compared to human SP-D (hSP-D), mainly due to key residues in the lectin domain of pSP-D that contribute to its profound neutralizing activity. These observations provided the basis for the design of a full-length recombinant mutant form of hSP-D, designated as "improved SP-D" (iSP-D). Inspired by pSP-D, the lectin domain of iSP-D has 5 amino acids replaced (Asp324Asn, Asp330Asn, Val251Glu, Lys287Gln, Glu289Lys) and 3 amino acids inserted (326Gly-Ser-Ser). Characterization of iSP-D revealed no major differences in protein assembly and saccharide binding selectivity as compared to hSP-D. However, hemagglutination inhibition measurements showed that iSP-D expressed strongly enhanced activity compared to hSP-D against 31 different IAV strains tested, including (pandemic) IAVs that were resistant for neutralization by hSP-D. Furthermore, iSP-D showed increased viral aggregation and enhanced protection of MDCK cells against infection by IAV. Importantly, prophylactic or therapeutic application of iSP-D decreased weight loss and reduced viral lung titers in a murine model of IAV infection using a clinical isolate of H1N1pdm09 virus. These studies demonstrate the potential of iSP-D as a novel human-based antiviral inhalation drug that may provide immediate protection against or recovery from respiratory (pandemic) IAV infections in humans.

The Dual Role of Surfactant Protein-D in Vascular Inflammation and Development of Cardiovascular Disease.

Cardiovascular disease (CVD) is responsible for 31% of all global deaths. Atherosclerosis is the major cause of cardiovascular disease and is a chronic inflammatory disorder in the arteries. Atherosclerosis is characterized by the accumulation of cholesterol, extracellular matrix, and immune cells in the vascular wall. Recently, the collectin surfactant protein-D (SP-D), an important regulator of the pulmonary immune response, was found to be expressed in the vasculature. Several in vitro studies have examined the role of SP-D in the vascular inflammation leading to atherosclerosis. These studies show that SP-D plays a dual role in the development of atherosclerosis. In general, SP-D shows anti-inflammatory properties, and dampens local inflammation in the vessel, as well as systemic inflammation. However, SP-D can also exert a pro-inflammatory role, as it stimulates C-C chemokine receptor 2 inflammatory blood monocytes to secrete tumor necrosis-factor α and increases secretion of interferon-γ from natural killer cells. In vivo studies examining the role of SP-D in the development of atherosclerosis agree that SP-D plays a proatherogenic role, with SP-D knockout mice having smaller atherosclerotic plaque areas, which might be caused by a decreased systemic inflammation. Clinical studies examining the association between SP-D and cardiovascular disease have reported a positive association between circulatory SP-D level, carotid intima-media thickness, and coronary artery calcification. Other studies have found that circulatory SP-D is correlated with increased risk of both total and cardiovascular disease mortality. Both in vitro, in vivo, and clinical studies examining the relationship between SP-D and CVDs will be discussed in this review.

Ozone Inhalation Attenuated the Effects of Budesonide on Aspergillus fumigatus-Induced Airway Inflammation and Hyperreactivity in Mice.

Inhaled glucocorticoids form the mainstay of asthma treatment because of their anti-inflammatory effects in the lung. Exposure to the air pollutant ozone (O3) exacerbates chronic airways disease. We and others showed that presence of the epithelial-derived surfactant protein-D (SP-D) is important in immunoprotection against inflammatory changes including those induced by O3 inhalation in the airways. SP-D synthesis requires glucocorticoids. We hypothesized here that O3 exposure impairs glucocorticoid responsiveness (including SP-D production) in allergic airway inflammation. The effects of O3 inhalation and glucocorticoid treatment were studied in a mouse model of allergic asthma induced by sensitization and challenge with Aspergillus fumigatus (Af) in vivo. The role of O3 and glucocorticoids in regulation of SP-D expression was investigated in A549 and primary human type II alveolar epithelial cells in vitro. Budesonide inhibited airway hyperreactivity, eosinophil counts in the lung and bronchoalveolar lavage (BAL) and CCL11, IL-13, and IL-23p19 release in the BAL of mice sensitized and challenged with Af (p < 0.05). The inhibitory effects of budesonide were attenuated on inflammatory changes and were completely abolished on airway hyperreactivity after O3 exposure of mice sensitized and challenged with Af. O3 stimulated release of pro-neutrophilic mediators including CCL20 and IL-6 into the airways and impaired the inhibitory effects of budesonide on CCL11, IL-13 and IL-23. O3 also prevented budesonide-induced release of the immunoprotective lung collectin SP-D into the airways of allergen-challenged mice. O3 had a bi-phasic direct effect with early (<12 h) inhibition and late (>48 h) activation of SP-D mRNA (sftpd) in vitro. Dexamethasone and budesonide induced sftpd transcription and translation in human type II alveolar epithelial cells in a glucocorticoid receptor and STAT3 (an IL-6 responsive transcription factor) dependent manner. Our study indicates that O3 exposure counteracts the effects of budesonide on airway inflammation, airway hyperreactivity, and SP-D production. We speculate that impairment of SP-D expression may contribute to the acute O3-induced airway inflammation. Asthmatics exposed to high ambient O3 levels may become less responsive to glucocorticoid treatment during acute exacerbations.

Biomarkers of skin and lung fibrosis in systemic sclerosis.

Introduction: Excessive fibrosis is the hallmark of systemic sclerosis (SSc) and numerous experts are investing efforts into identifying parameters that could predict disease course and prognosis. Here, we review the available and potential biomarkers of lung and skin fibrosis in SSc.Areas covered: Specific autoantibodies are important for the determination of clinical subsets of SSc, making them routine in clinical practice. Physical parameters, such as modified Rodnan skin score (mRSS) and pulmonary function tests are standardized in evaluating the skin and lung involvement. High resolution computed tomography is the gold standard for SSc-related interstitial lung disease (ILD) diagnostics, as well as progress evaluation. Nowadays, the main focus is on specific autoantibodies, various genetic pathways, and different cytokines. In addition to the profibrotic role of interleukin 6 and transforming growth factor-ß, newer studies stress on glycoprotein Krebs von den Lungen-6 (KL-6), surfactant protein-D (SP-D) and chemokine (C-C motif) ligand 18 (CCL18) as potential biomarkers of skin and lung fibrosis in SSc.Expert opinion: Skin and lung biomarkers in SSc frequently mirror the typical signs of fibrosis, overlapping sporadically. There is an urgent need for better diagnostic distinction and evaluation; therefore, further investigations are critical to establish more suitable biomarkers of SSc.

Surfactant protein-D modulation of pulmonary macrophage phenotype is controlled by S-nitrosylation.

Surfactant protein-D (SP-D) is a regulator of pulmonary innate immunity whose oligomeric state can be altered through S-nitrosylation to regulate its signaling function in macrophages. Here, we examined how nitrosylation of SP-D alters the phenotypic response of macrophages to stimuli both in vivo and in vitro. Bronchoalveolar lavage (BAL) from C57BL6/J and SP-D-overexpressing (SP-D OE) mice was incubated with RAW264.7 cells ± LPS. LPS induces the expression of the inflammatory genes Il1b and Nos2, which is reduced 10-fold by SP-D OE-BAL. S-nitrosylation of the SP-D OE-BAL (SNO-SP-D OE-BAL) abrogated this inhibition. SNO-SP-D OE-BAL alone induced Il1b and Nos2 expression. PCR array analysis of macrophages incubated with SP-D OE-BAL (±LPS) shows increased expression of repair genes, Ccl20, Cxcl1, and Vcam1, that was accentuated by LPS. LPS increases inflammatory gene expression, Il1a, Nos2, Tnf, and Ptgs2, which was accentuated by SNO-SP-D OE-BAL but inhibited by SP-D OE-BAL. The transcription factor NF-κB was identified as a target for SNO-SP-D by IPA, which was confirmed by Trans-AM ELISA in vitro. In vivo, SP-D overexpression increases the burden of infection in a Pneumocystis model while increasing cellular recruitment. Expression of iNOS and the production of NO metabolites were significantly reduced in SP-D OE mice relative to C57BL6/J. Inflammatory gene expression was increased in infected C57BL6/J mice but decreased in SP-D OE. SP-D oligomeric structure was disrupted in C57BL6/J infected mice but unaltered within SP-D OE. Thus SP-D modulates macrophage phenotype and the balance of multimeric to trimeric SP-D is critical to this regulation.

Chronic rhinosinusitis in unified airway disease: surfactant proteins as mediators of respiratory immunity.

PURPOSE OF REVIEW: The aim of this review is to describe the co-occurrence of chronic rhinosinusitis (CRS) with other inflammatory illnesses of the lower respiratory system characterised by airway obstruction and hyperresponsiveness, such as asthma, cystic fibrosis (CF), and chronic obstructive pulmonary disease (COPD) in the context of the unified airway disease (UAD). We also sought to discuss the novel role of surfactant proteins as mediators of innate immunity in the sinonasal epithelium and their potential as therapeutic interventions. RECENT FINDINGS: Different epidemiological and physiological studies in CRS and asthma have outlined that there are common clustering patterns in the phenotypes/endotypes of both diseases, reinforcing the notion of the UAD. Also, surfactant proteins A (SP-A) and SP-D have now emerged as novel innate immunity molecules in bacterial sinusitis and allergic fungal sinusitis patients, respectively. SUMMARY: CRS and asthma coexist and are interconnected. Therefore, management of CRS and asthma must be jointly carried out as one functional entity. SP-A and SP-D bridge the innate and adaptive immunity mechanisms of the sinonasal epithelium to bring together a well-orchestrated mechanism that effectively fights pathogens. The use of SP-A to ameliorate the innate immune responses in CRS is a new concept and is likely to lead to new horizons in CRS therapeutic regimens.  .

Serial change in serum biomarkers during treatment of Non-HIV Pneumocystis pneumonia.

BACKGROUND: For patients with non-human immunodeficiency virus (HIV) Pneumocystis pneumonia (PCP), data are limited on serial changes in serum biomarkers and the correlations with clinical outcomes. OBJECTIVE: This study evaluated serial change in serum biomarkers and clinical outcomes of non-HIV PCP. METHODS: We retrospectively reviewed data from 63 patients treated for non-HIV PCP at Toho University Omori Medical Center. The patients were classified as survivors and nonsurvivors on the basis of 60-day PCP mortality. The groups were compared for clinical course and levels of serum biomarkers (ß-D glucan, Krebs von den Lungen-6 antigen [KL-6], and surfactant protein-D [SP-D]), which were measured at baseline, and 7 days and 14 days after starting treatment. In addition, serial changes in serum biomarkers were analyzed in survivors and nonsurvivors. RESULTS: There were 14 PCP nonsurvivors and 49 survivors. Biomarker values were not different between groups at baseline. At 7 and 14 days after starting treatment, the proportions of patients with elevated ß-D glucan and KL-6 did not significantly differ between groups; however, the proportion of patients with elevated SP-D was significantly lower among survivors than among nonsurvivors (57.1% vs. 100%, p = 0.009; 30% vs. 100%, p < 0.001; respectively). SP-D on day 14 was significantly lower than that at baseline among survivors (99.6 [61.0-190.3] vs. 156 [100.8-283.5]; p = 0.045) but significantly higher among nonsurvivors (974 [744.5-1565] vs. 317 [211-448]; p = 0.03). CONCLUSION: Serum SP-D value continues to increase after failure of treatment for non-HIV PCP and may thus be associated with outcomes for non-HIV PCP patients.

Surfactant Protein D Reverses the Gene Signature of Transepithelial HIV-1 Passage and Restricts the Viral Transfer Across the Vaginal Barrier.

Effective prophylactic strategy against the current epidemic of sexually transmitted HIV-1 infection requires understanding of the innate gatekeeping mechanisms at the genital mucosa. Surfactant protein D (SP-D), a member of the collectin family of proteins naturally present in the vaginal tract, is a potential HIV-1 entry inhibitor at the cellular level. Human EpiVaginal tissues compartmentalized in culture inserts were apically exposed to HIV-1 and/or a recombinant fragment of human SP-D (rfhSP-D) and viral passage was assessed in the basal chamber containing mononuclear leukocytes. To map the gene signature facilitating or resisting the transepithelial viral transfer, microarray analysis of the HIV-1 challenged EpiVaginal tissues was performed in the absence or presence of rfhSP-D. Mucosal biocompatibility of rfhSP-D was assessed ex vivo and in the standard rabbit vaginal irritation model. The passage of virus through the EpiVaginal tissues toward the underlying target cells was associated with a global epithelial gene signature including differential regulation of genes primarily involved in inflammation, tight junctions and cytoskeletal framework. RfhSP-D significantly inhibited HIV-1 transfer across the vaginal tissues and was associated with a significant reversal of virus induced epithelial gene signature. Pro-inflammatory NF-κB and mTOR transcripts were significantly downregulated, while expression of the tight junctions and cytoskeletal genes was upheld. In the absence of virus, rfhSP-D directly interacted with the EpiVaginal tissues and upregulated expression of genes related to structural stability of the cell and epithelial integrity. There was no increment in the viral acquisition by the PBMCs present in basal chambers wherein, the EpiVaginal tissues in apical chambers were treated with rfhSP-D. The effective concentrations of rfhSP-D had no effect on lactobacilli, epithelial barrier integrity and were safe on repeated applications onto the rabbit vaginal mucosa. This pre-clinical safety data, coupled with its efficacy of restricting viral passage via reversal of virus-induced gene expression of the vaginal barrier, make a strong argument for clinical trials of rfhSP-D as a topical anti-HIV microbicide.

The lung environment controls alveolar macrophage metabolism and responsiveness in type 2 inflammation.

Fine control of macrophage activation is needed to prevent inflammatory disease, particularly at barrier sites such as the lungs. However, the dominant mechanisms that regulate the activation of pulmonary macrophages during inflammation are poorly understood. We found that alveolar macrophages (AlvMs) were much less able to respond to the canonical type 2 cytokine IL-4, which underpins allergic disease and parasitic worm infections, than macrophages from lung tissue or the peritoneal cavity. We found that the hyporesponsiveness of AlvMs to IL-4 depended upon the lung environment but was independent of the host microbiota or the lung extracellular matrix components surfactant protein D (SP-D) and mucin 5b (Muc5b). AlvMs showed severely dysregulated metabolism relative to that of cavity macrophages. After removal from the lungs, AlvMs regained responsiveness to IL-4 in a glycolysis-dependent manner. Thus, impaired glycolysis in the pulmonary niche regulates AlvM responsiveness during type 2 inflammation.

Lipophilic Allergens, Different Modes of Allergen-Lipid Interaction and Their Impact on Asthma and Allergy.

Molecular allergology research has provided valuable information on the structure and function of single allergenic molecules. There are several allergens in food and inhalant allergen sources that are able to interact with lipid ligands via different structural features: hydrophobic pockets, hydrophobic cavities, or specialized domains. For only a few of these allergens information on their associated ligands is already available. Several of the allergens are clinically relevant, so that it is highly probable that the individual structural features with which they interact with lipids have a direct effect on their allergenic potential, and thus on allergy development. There is some evidence for a protective effect of lipids delaying the enzymatic digestion of the peanut (Arachis hypogaea) allergen Ara h 8 (hydrophobic pocket), probably allowing this molecule to get to the intestinal immune system intact (sensitization). Oleosins from different food allergen sources are part of lipid storage organelles and potential marker allergens for the severity of the allergic reaction. House dust mite (HDM), is more often associated with allergic asthma than other sources of inhalant allergens. In particular, lipid-associated allergens from Dermatophagoides pteronyssinus which are Der p 2, Der p 5, Der p 7, Der p 13, Der p 14, and Der p 21 have been reported to be associated with severe allergic reactions and respiratory symptoms such as asthma. The exact mechanism of interaction of these allergens with lipids still has to be elucidated. Apart from single allergens glycolipids have been shown to directly induce allergic inflammation. Several-in parts conflicting-data exist on the lipid (and allergen) and toll-like receptor interactions. For only few single allergens mechanistic studies were performed on their interaction with the air-liquid interface of the lungs, in particular with the surfactant components SP-A and SP-D. The increasing knowledge on protein-lipid-interaction for lipophilic and hydrophobic food and inhalant allergens on the basis of their particular structure, of their capacity to be integral part of membranes (like the oleosins), and their ability to interact with membranes, surfactant components, and transport lipids (like the lipid transfer proteins) are essential to eventually clarify allergy and asthma development.

The Lung Mucosa Environment in the Elderly Increases Host Susceptibility to Mycobacterium tuberculosis Infection.

As we age, there is an increased risk for the development of tuberculosis (TB) caused by Mycobacterium tuberculosis (Mtb) infection. Few studies consider that age-associated changes in the alveolar lining fluid (ALF) may increase susceptibility by altering soluble mediators of innate immunity. We assessed the impact of adult or elderly human ALF during Mtb infection in vitro and in vivo. We identified amplification of pro-oxidative and proinflammatory pathways in elderly ALF and decreased binding capability of surfactant-associated surfactant protein A (SP-A) and surfactant protein D (SP-D) to Mtb. Human macrophages infected with elderly ALF-exposed Mtb had reduced control and fewer phagosome-lysosome fusion events, which was reversed when elderly ALF was replenished with functional SP-A/SP-D. In vivo, exposure to elderly ALF exacerbated Mtb infection in young mice. Our studies demonstrate how the pulmonary environment changes as we age and suggest that Mtb may benefit from declining host defenses in the lung mucosa of the elderly.