Surfactant Protein A Concentration Research Articles

Surfactant Protein A can Affect the Surface Tension of the Eustachian Tube and Macrophage Migration.

To assess the role and possible mechanism of surfactant protein A (SPA) in the pathogenesis of otitis media with effusion (OME). This was a multi-part study with both an in vivo mouse model study as well as an in vitro study. The control and study groups (OME group) received phosphate-buffered saline and inactivated Streptococcus pneumoniae, respectively, via external auditory meatus injections. Changes in the surface tension of secretions from the eustachian tube (ET) and SPA expression were measured in both groups. A transwell assay was performed to observe the effect of different concentrations of SPA on the migration ability of macrophages. We examined the differentially expressed genes related to SPA-treated macrophages using RNA-seq analysis. On Day 3, the surface tension of the OME group was higher than that of the control group (p=0.014). The variation intensity of SPA in the ET of the OME group was significantly lower than that of the control group (p < 0.001). Surface tension was correlated with SPA (r=-0.525, p=0.037). The expression of SPA and macrophages in the ET was different between the two groups. In vitro experiments revealed that macrophages showed different migration abilities with SPA concentration changes (p < 0.05). RNA-seq and western blotting were performed after macrophages were treated with SPA. The results showed that RhoA and Rac1/2/3 were differentially expressed. SPA can change the surface tension of secretions from the ET and affect macrophage migration to alter the function of the ET. Although research in this field of OME is nascent, initial work suggests that SPA likely plays an important role in OME progression. NA Laryngoscope, 133:1726-1733, 2023.

Surfactant Protein A in particles in exhaled air (PExA), bronchial lavage and bronchial wash - a methodological comparison

IntroductionAt present, there are few methods available for monitoring respiratory diseases affecting distal airways. Bronchoscopy is the golden standard for sampling the lower airways. The recently developed method for collecting non-volatile material from exhaled air – PExA (Particles in Exhaled air) is a promising new tool, but no direct comparison between the two methods has yet been performed. The aim of the present study was to compare sampling using PExA with bronchial wash (BW) representing the larger more proximal airways and broncho-alveolar lavage (BAL) representing the distal airways.Methods15 healthy non-smoking subjects (7 female/8 male), age 28 ± 4 years, with normal lung function were included in the study. PExA-sampling (2 × 250 ng particles) and bronchoscopy with BW (2 × 20 ml) and BAL (3 × 60 ml sterile saline) was performed. Albumin and Surfactant Protein A (SP-A) were analyzed with ELISA, and analyses of correlation were performed.ResultsA significant association was found between BAL-fluid albumin and PExA-albumin (rs:0.65 p = 0.01). There was also an association between SP-A in PExA and BAL, when corrected for albumin concentration (rs:0.61, p = 0.015). When correlating concentrations of albumin and SP-A in bronchial wash and PExA respectively, no associations were found.ConclusionsThis is the first direct comparison between the bronchoscopy-based BW/BAL-fluids and material collected using the PExA methodology. Both albumin and albumin-corrected SP-A concentrations were significantly associated between BAL and PExA, however, no such association was found in either marker between BW and PExA. These results indicate that the PExA method samples the distal airways. PExA is thus considered a new promising non-invasive assessment for monitoring of the distal airways.

Modified Da-chai-hu Decoction regulates the expression of occludin and NF-κB to alleviate organ injury in severe acute pancreatitis rats

Modified Da-chai-hu Decoction (MDD), a traditional Chinese medicinal formulation, which was empirically generated from Da-chai-hu decoction, has been utilized to treat severe acute pancreatitis (SAP) for decades. The aim of the present study was to explore its potential organprotective mechanism in SAP. In the present study, rat SAP model was induced by retrograde injection of 3.5% sodium taurocholate into the biliopancreatic duct, MDD (23.35 g/kg body weight, twelve times the clinical dose) were orally given at 2 h before and 10 h after injection. At 12 h after model induction, blood was taken from vena cava for analysis of amylase, diamine oxidase (DAO), pulmonary surfactant protein-A (SP-A), and C-reactive protein (CRP). Histopathological change of pancreas, ileum and lung was assayed by H&E staining, myeloperoxidase (MPO) activity were determinated using colorimetric assay, and the expressions of occludin and nuclear factor-κB (NF-κB) were detected by real-time RT-PCR and western blot, respectively. In addition, the tissue concentrations of tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), and monocyte chemoattractant protein-1 (MCP-1) were measured by enzyme-linked immunosorbent assay (ELISA). The results showed that in SAP rats, MDD significantly alleviated histopathological damage, depressed the MPO activity and the concentrations of TNF-α, IL-1β, and MCP-1 of pancreas, ileum and lung, and reduced the serum levels of amylase [(3283.4 ± 585.5) U·L-1vs (5626.4 ± 795.1)U·L-1], DAO [(1100.1 ± 334.3) U·L-1vs (1666.4 ± 525.3) U·L-1] and CRP [(7.6 ± 1.2) μg·mL-1vs (17.8 ± 3.8) μg·mL-1]. However, the serum SP-A concentration [(106.1 ± 16.6) pg·mL-1vs (90.1 ± 14.9) pg·mL-1] was elevated when treated SAP rats with MDD. Furthermore, MDD increased the occludin expression and reduced the NF-κB expression in pancreas, ileum and lung of SAP rats. Our findings suggested that MDD administration was an effective therapeutic approach for SAP treatment. It could up-regulate occludin expression to protect intercellular tight junction and down-regulate NF-κB expression to inhibit inflammatory reaction of pancreas, ileum and lung.

The role of erlotinib in the acute lung injury and the expression of surfactant protein A

Objective To investigate the role of erlotinib in the expression of surfactant protein A (SP-A) in LPS-induced acute lung injury (ALI) of mice model. Methods C57BL/6 mice were randomly (random number)divided into control group (n=6), ER group (n=6), LPS group (n=6), and ER+LPS group (n=6). In the LPS group, 2 mg/kg LPS was instilled into trachea of mice to induce lung injury. In control group, normal saline was instilled into trachea of mice instead. In the ER+LPS group and ER group, 100 mg/kg of erlotinib was instilled into stomach of mice, and one hour later, 2 mg/kg LPS was instilled into trachea of mice in ER+PLS group to induce lung injury. Twenty-four hours later, bronchoalveolar lavage fluid (BALF) and lung tissue of mice in four groups were collected. HE staining were used for evaluating pathological changes of lung injury. Lung wet/dry weight ratio, protein concentrations and total cell numbers in the BALF were measured to determine the degree of pulmonary edema. Immunohistochemical staining and Western Blot were used for testing the protein expression of SP-A, Data of multiple groups were analyzed by one way variance (ANOVA) and inter-group comparisons were made by the least significant difference (LSD) tests. Results There was no significant difference in lung injury score (LIS) between control group (0.056±0.008) and ER (0.064±0.037) group, The LIS in LPS group (0.846±0.047) was higher than that in control group, however the LIS in ER+LPS group (0.279±0.020) was significant lower than that in LPS group (P < 0.05). Lung wet/dry weight, SP-A concentration and total cell numbers in the bronchoalveolar lavage fluid revealed that the degree of pulmonary edema in LPS group was higher than that in control group, and this pulmonary edema was reversed by erlotinib treatment. Immunohistochemical staining and Western blot showed that the expression of SP-A in LPS group was decreased compared with control group, but it was recovered after erlotinib treatment (P < 0.05). Conclusions Erlotinib could protect the LPS-induced ALI, and it may be related to the regulation of SP-A. Key words: Acute lung injury; Mice; Lipopolysaccharide; Erlotinib; Alveolar epithelial cells; Surfactant protein A; Inflammatory response; Epidermal growth factor receptor; Drug treatment

Expression of surfactant Protein-A in the Haemophilus influenzae-induced otitis media in a rat model

The objective of this study was to investigate the expression and the role of surfactant protein A (SP-A) in the middle ear (ME) mucosa in response to bacterial infection in a rat model. Otitis media (OM) was induced by surgical inoculation of non-typeable Haemophilus influenza (NTHi) into the ME cavity of Sprague-Dawley rats. The rats were divided into an NTHi-induced OM group and a phosphate-buffered saline-injected control group. The NTHi-induced OM and control groups were subdivided into sets of 6 rats, one for each of the 6 time points (0, 1, 2, 4, 7, and 14 days post-inoculation), at which point the rats were euthanized after inoculation. The concentrations of SP-A in the ME effusion were determined by an enzyme-linked immunosorbent assay (ELISA). Tissue expression of SP-A, interleukin-1β (IL-1β), interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α) in infected ME mucosa was assessed by immunohistochemical staining. For mRNA expression quantification, RNA was extracted from the ME mucosa and SP-A expression was monitored and compared between the control and OM groups using quantitative polymerase chain reaction (PCR). Expression of IL-1β, IL-6, and TNF-α in the ME mucosa was also evaluated. SP-A expression was evaluated in the effusion of pediatric OM patients (70 ears) who received ventilation-tube insertion by ELISA. SP-A was detected in normal rat ME mucosa before bacterial inoculation. SP-A expression was up-regulated in the NTHi-induced OM group (p = 0.046). Immunohistochemical staining revealed increased SP-A expression on post-inoculation day 1, 2, and 4 in the OM group. Expression of proinflammatory cytokines (IL-1β, IL-6, and TNF-α) in the ME also increased significantly on post-inoculation day 1, 2, and 4 in the OM group. It correlated with changes in SP-A expression. Expression of SP-A was also identified in the ME effusion of humans. SP-A exists in the ME of the rat and was up-regulated in the ME of NTHi-induced OM. Expression of IL-1β, IL-6, and TNF-α was increased in the ME of the bacteria-induced OM in the rat model. The results suggest that SP-A may play a significant role in the early phase of OM induction and subsequent recovery from it.

Clinical value of surfactant protein-A in exudate pleural effusion

To evaluate the clinical value of surfactant protein-A (SP-A) in exudate pleural effusion (EPE). Methods: This clinical study was prospective, observational and cross-sectional. Two hundred and fifteen patients with pleural effusion were divided into the transudate pleural effusions (TPE) group and the EPE group. TPE patients served as the control group. The concentrations of pleuraleffusions SP-A (SP-Apl) and serum SP-A (SP-Ase) were measured by ELISA, and receiver operator characteristic (ROC) curve and multivarate Cox analysis of SP-A was analysed for its clinical value. Results: SP-Apl concentrations in the EPE group were significantly higher than that in the TPE group [(189.8±43.4) ng/mL vs (22.3±5.1) ng/mL, P<0.01]; SP-Ase concentrations in the EPE group were higher than that in the TPE group [(78.9±11.3) ng/mL vs (25.8±12.4) ng/mL, P<0.05]; SP-Apl concentrations were significantly higher than the concentrations of SP-Ase in the EPE group (P<0.01). In EPE group, SP-Apl and SP-Ase concentration in the patients with primary lung adenocarcinomas were the highest. The cut off value of SP-Apl concentrations was more than 484.5 ng/mL, yielding a 85.4% sensitivity and 95.2% specificity for diagnosing primary lung adenocarcinomas, with an area under the curve (AUC) of 0.943 (95% CI 0.852 to 0.934, P<0.01); when SP-Ase concentration was more than 84.2 ng/mL, it yielded a 76.4% sensitivity and 94.3% specificity for diagnosing primary lung adenocarcinomas, with an AUC of 0.910 (95% CI 0.921 to 0.953, P<0.01). Conclusion: While SP-Apl concentration is more than 484.5 ng/mL and/or SP-Ase concentration is more than 84.2 ng/mL, it may be helpful for the diagnosis of primary lung adenocarcinomas with the usage of pleural effusion.

Effects of probiotics on ghrelin and lungs in children with acute lung injury: A double-blind randomized, controlled trial.

To assess the effects of probiotics on serum ghrelin levels and protection for lungs in children with acute lung injury (ALI). This study was performed as a double-blind, randomized, and controlled trial in a pediatric intensive care unit (PICU). The eligible children with ALI were assigned to either probiotic treatment or an identical placebo for 10 days. Serum ghrelin, SP-A(surfactant protein-A), TNF-α, and IL-6 concentrations were assessed at baseline and at the end of trial. Meanwhile, pulmonary function test and echocardiography were examined, then VPEF (volume to peak tidal expiratory flow), TPEF/TE (the ratio of time taken to reach peak expiratory flow to total expiratory time), MAP (mean arterial pressure), and PAP (pulmonary artery pressure) were recorded. Eighty participants fulfilled the study requirements with 40 children for each group. The groups were comparable in baseline characteristics. Serum SP-A, TNF-α, and IL-6 levels in the probiotic group were 212.6 ± 52.9 ng/mL, 401.9 ± 56.4 pg/mL, and 245.1 ± 55.1 pg/mL on day 10, respectively, significantly lower levels compared to the control group where the same parameters were 248.2 ± 57 ng/mL, 449.4 ± 60.1 pg/mL, and 308.3 ± 92.2 pg/mL (P < 0.01). However, ghrelin concentrations were elevated in the intervention group (P < 0.05). On measurement of pulmonary function, the probiotic group demonstrated a VPEF of 26.1 ± 4.2 mL and TPEF/TE of 29.1 ± 4.7%, which were higher than the control group (24.7 ± 4.3 mL and 26.9 ± 4.7%, respectively) (P < 0.05). MAP and PAP also improved in the probiotic group (P < 0.05). Furthermore, ghrelin, SP-A, TNF-α, IL-6, and PAP were negatively correlated. Positive correlations were found between ghrelin, TPEF/TE, and MAP. There were no probiotic-associated adverse events during the observation. Probiotics administrated to children with ALI alleviates the inflammation of lungs, improves pulmonary function and circulation by ghrelin.

Concentrations of SP-A and HSP70 are associated with polarization of macrophages in pleural effusions of non-small cell lung cancer

Damage-associated molecular pattern (DAMP) molecules can initiate an immune response through Toll-like receptors (TLRs). DAMPs are released from cells as a response to the extracellular danger and can be by-products of tissue damage. In cancer microenvironment necrotic cells release debris which has potency to become DAMPs. Non-small cell lung cancer (NSCLC) is often accompanied by pleural effusion (PE), which contains a variety of DAMPs. Surfactant protein A (SP-A) and heat shock protein 70 (Hsp70) are important DAMPs in the respiratory tract.The aim of this study was to determine a correlation between SP-A or Hsp70 and development of PE in the course of NSCLC. Moreover, we aimed to determine relationships between DAMPs and certain humoral factors associated with formation and persistence of PE as well as pleural-residing macrophages. In 34 PE samples, we estimated concentration of SP-A, Hsp70, IL-6, IL-18, G-CSF, M-CSF, SCF, SDF1α, VEGF as well as the fraction of macrophages and their pattern of polarization.We have found correlations between the concentration of the SP-A and Hsp70 and the percentage of PE-derived macrophages, also between concentrations of SP-A and Hsp70, and cytokines which participate in inflammation and processes involved in remodeling of extracellular matrix (ECM). Our data indicate an important role of SP-A during the development of PE associated with NSCLC. We suggest that measurement of concentration level of SP-A can be helpful in the course of diagnosis of malignant PE associated with NSCLC.

P239 Diagnostic potential of MMP-1 and SP-A as biomarkers to distinguish chronic hypersensitivity pneumonitis from Idiopathic pulmonary fibrosis

Introduction: Hypersensitivity pneumonitis (HP) is an inflammatory interstitial lung disease caused by a wide variety of organic particles. In general, the diagnosis of chronic HP represents a challenge for clinicians that need to use a constellation of clinical, laboratory, radiographic and pathological data to assess a confident diagnosis. Importantly, the clinical features cHP can be indistinguishable from those observed in idiopathic pulmonary fibrosis (IPF). Objective: To determine whether serum levels of surfactant protein A (SP-A) and matrix metalloprotease-1 (MMP-1) may help to differentiate cHP from IPF. Methods: This was a cross-sectional study including patients with IPF (n = 31), cHP (n = 31) and 17 healthy subjects. SP-A and MMP-1 serum levels were determined by ELISA and were evaluated for the differential diagnosis between HP and IPF, and also correlated with values of pulmonary function tests at diagnosis. Results: MMP-1 and SP-A were significantly elevated in cHP compared with the other two groups. MMP-1 (HP= 11.7 ± 14 ng/ml versus FPI= 16 ± 20.7 ng/ml, p< 0.001) and versus Controls= (0.4 ± 0.3 ng/ml, p < 0.0001), ANOVA between groups < 0.0001. SP-A (HP= 74 ± 30 pg/ml versus FPI= 54 ± 20 pg/ml, p< 0.02) and versus Controls= (35 ± 6 pg/ml, p < 0.0001), ANOVA between groups < 0.0001. There was no correlation between MMP-1 or SP-A levels with the abnormalities in pulmonary function tests. Conclusions: Our findings indicate that patients with cHP display the highest concentration of MMP-1 and SP-A which may be useful biomarkers to distinguish it from IPF.

Effects of bone marrow-derived mesenchymal stem cells transfected with survivin on pulmonary fibrosis in mice

The aim of the present study was to investigate the effects of bone marrow-derived mesenchymal stem cells (BMSCs) transfected with survivin on lung fibrosis in mice. Mice with bleomycin-induced pulmonary fibrosis were allocated at random to group A, B or C, and injected with 1×106 survivin gene-expressing BMSCs, 1×106 BMSCs or normal saline, respectively. A total of 6 mice were sacrificed from each group on days 7, 14 and 28 after treatment. The extent of alveolitis and pulmonary fibrosis was assessed and the apoptotic rates of the BMSCs and survivin-expressing BMSCs were detected. The content of surfactant protein A (SP-A) in the lung and hydroxyproline (Hyp) in the serum was measured. The mRNA expression levels of transforming growth factor (TGF)-β1 and matrix metalloproteinase (MMP)-9 in the lung tissue of the mice was detected. Furthermore, the protein expression levels of caspase-3 and -9 were detected. The apoptotic rates of the BMSCs (group B) and survivin-expressing BMSCs (group A) were 14.466±1.953 and 7.718±0.493%, respectively. The degree of lung fibrosis in groups A and B was reduced compared with that in group C. The hydroxyproline content in groups A and B was reduced compared with that in group C, and the SP-A content in groups A and B was increased compared with that in group C. The mRNA expression levels of TGF-β1 in group A were reduced compared with those in group B, and the levels in group B were reduced compared with those in group C. By contrast, the mRNA expression levels of MMP-9 in group A were increased compared with those in groups B and C, and the levels in group B were increased compared with those in group A. The expression levels of caspase-3 and -9 in group A were elevated compared with those in groups B and C. In conclusion, BMSCs are effective in preventing bleomycin-induced lung fibrosis, and survivin may enhance the protective effects of BMSCs.

Protective effects of different modes of ventilation on lungs on operated side during one-lung ventilation in patients undergoing thoracic surgery

Objective To evaluate the protective effects of different modes of ventilation on the lungs on the operated side during one-lung ventilation （OLV ） in patients undergoing thoracic surgery .Methods Forty-five ASA physical status Ⅰ or Ⅱ patients of both sexes ,aged 45-64 yr ,weighing 65-80 kg ,were randomly divided into 3 groups （n=15 each） using a random number table ：group A ,group B and group C .After induction of anesthesia ,the patients were intubated with double-lumen tube and OLV was performed .During OLV ,the lung on the operated side was collapsed naturally in group A ,positive pressure ventilation （FGF 2 L/min） was applied in the lung on the operated side in group B ,and high-frequency jet ventilation （frequency 100 beats/min ,driving pressure 0.5 kg/cm2 ） was used in the lung on the operated side in group C .Immediately after intubation （T0 ） , and at 1.5 h （T1 ） and 2 h （T2 ） of OLV ,blood samples were taken from the central vein and radial artery for determination of the serum interleukin-6 （IL-6 ） and IL-8 concentrations .The net release of IL-6 and IL-8 was calculated .Blood samples were taken from the radial artery at T0-2 for blood gas analysis and for determination of surfactant protein A （SP-A） concentration in the serum .Respiratory index （RI） was calculated .The non-cancer tissues 1.0 cm × 1.0 cm × 1.0 cm which were extracted from the lung cancer specimens were used for microscopic examination of the pathological changes of lungs which were scored .Results Compared with group A ,the net release of IL-6 and IL-8 ,serum SP-A concentration ,RI and pathological scores were significantly decreased at T1 ,2＆nbsp;in B and C groups （ P〈0.05） .Compared with group B ,the serum SP-A concentration and RI were significantly decreased at T1 ,2 , and the net release of IL-6 was increased at T2 in group C （ P〈 0.05 ） .Conclusion Continuous positive ventilation and high-frequency jet ventilation both can effectively protect the lungs on the operated side during OLV in patients undergoing thoracic surgery ,and the efficacy of high-frequency jet ventilation is better . Key words: Continuous positive airway pressure; High-frequency jet ventilation

Surfactant Protein-A Concentration in Sera from Dogs with Pulmonary Parenchymal Diseases

Pulmonary surfactant protein A (SP-A) is used as a biomarker to understand the clinical features of pulmonary diseases and associated prognostic indices in human medicine. This study was conducted to investigate whether or not serum SP-A concentration can be used as a biomarker for identifying pulmonary parenchymal diseases in dogs. Thirty-two dogs with pulmonary parenchymal diseases, 34 with nonrespiratory diseases and 57 healthy dogs were included. Serum SP-A concentration was measured in all dogs using sandwich enzyme linked immunosorbent assay with an anti-dog SP-A polyclonal antibody. Median serum SP-A concentration in healthy dogs was <2.0 ng/ml, whereas that in dogs with aspiration pneumonia (n=11), primary lung tumors (n=9) and blunt traumatic lung injury (BTLI; n=12) was 3.1, 7.2 and 2.6 ng/ml, respectively; these values were significantly higher than those in healthy dogs. The serum SP-A concentration in dogs with nonrespiratory diseases was comparable with that in healthy dogs. No correlation was observed between the serum SP-A and plasma C-reactive protein concentrations in dogs with aspiration pneumonia and BTLI. There was a significant correlation between the serum SP-A concentration and thoracic radiographic changes in dogs with BTLI. These findings suggest that the serum SP-A concentration may be a useful clinical biomarker of alveolar damage that can be used for differential diagnosis of pulmonary parenchymal diseases and nonrespiratory diseases in dogs.

Surfactant Protein A (SP-A) and Angiotensin Converting Enzyme (ACE) as Early Biomarkers for Pulmonary Edema Formation in Ventilated Human Lung Lobes

Ex vivo perfused and ventilated lung lobes frequently develop pulmonary edema. We were looking for a suitable and early detectable biomarker in the perfusion fluid indicating lung cell damage and loss of tissue integrity in ventilated human lung lobes. Therefore, we elucidated whether surfactant protein A (SP-A) and angiotensin-converting enzyme (ACE) were measurable in the perfusion fluid and whether they were suitable indicators for edema formation occurring within the experimental time frame of 1-2 h. Patients (n = 39) undergoing a lobectomy, bilobectomy or pneumonectomy due to primary bronchial cell carcinoma were included in the studies. Lung lobes were extracorporally ventilated and perfused for up to 2 h. Two different perfusion fluids were used, plain perfusion buffer and perfusion buffer containing packed erythrocytes or buffy coats. Perfusion fluid samples were analyzed for SP-A and ACE using immunoassays served as perfusion fluids. SP-A and ACE concentrations were analyzed in fluid sample sets of 39 and 33 perfusion experiments, respectively. Degrees of edema formation were arbitrarily classified into three groups (≤ 29, 30-59, ≥ 60 % weight gain). The maximum increase of SP-A and ACE concentrations in the perfusate was significantly higher for more pronounced edemas in case of perfusions using a mixture of blood components and buffer. Interestingly, the time courses of ACE and SP-A were highly similar. We suggest that SP-A and ACE are promising early biochemical markers for the development for pulmonary edema formation in the ex vivo lung lobe perfusion.

Proteome Analysis of Bronchoalveolar Lavage Fluid in Chronic Hypersensitivity Pneumonitis

Hypersensitivity pneumonitis (HP) is an immune-mediated lung disease induced by inhalation of numerous antigens. Pathologically, chronic HP tends to show usual interstitial pneumonia (UIP) and fibrotic nonspecific interstitial pneumonia (fNSIP) patterns. Patients with UIP pattern present insidious onset and a risk for acute exacerbations. To evaluate the proteomic differences of bronchoalveolar lavage fluid (BALF) between UIP and fNSIP patterns, BALF from seven patients with UIP pattern and four patients with fNSIP pattern was examined using two-dimensional gel electrophoresis and mass spectrometry. By individually comparing each BALF sample, we found that the protein levels of surfactant protein A (SP-A), immunoglobulin heavy chain α, α-2 heat shock glycoprotein, haptoglobin β, and immunoglobulin J chain were significantly higher in the patients with UIP pattern than those in the patients with fNSIP pattern. In contrast, the protein levels of glutathione s-transferase, vitamin D-binding protein, and β-actin were significantly higher in the patients with fNSIP pattern than those in the patients with UIP pattern. To confirm the results of SP-A in the BALF proteome, we performed enzyme-linked immunosorbent assay in a larger group. The concentrations of SP-A in BALF from the patients with UIP pattern were significantly higher than those from the patients with fNSIP pattern (2.331 ± 1.656 μg/ml vs. 1.319 ± 1.916 μg/ml, p = 0.034). We identified several proteins that may play roles in the development of pathological differences between UIP and fNSIP patterns of chronic HP.

Direct evidence that GM-CSF inhalation improves lung clearance in pulmonary alveolar proteinosis

Autoimmune pulmonary alveolar proteinosis (aPAP) is caused by granulocyte/macrophage-colony stimulating factor (GM-CSF) autoantibodies in the lung. Previously, we reported that GM-CSF inhalation therapy improved alveolar-arterial oxygen difference and serum biomarkers of disease severity in these patients. It is plausible that inhaled GM-CSF improves the dysfunction of alveolar macrophages and promotes the clearance of the surfactant. However, effect of the therapy on components in bronchoalveolar lavage fluid (BALF) remains unclear. To figure out changes in surfactant clearance during GM-CSF inhalation therapy. We performed retrospective analyses of BALF obtained under a standardized protocol from the same bronchus in each of 19 aPAP patients before and after GM-CSF inhalation therapy (ISRCTN18931678, JMA-IIA00013; total dose 10.5-21 mg, duration 12-24 weeks). For evaluation, the participants were divided into two groups, high responders with improvement in alveolar-arterial oxygen difference ≥13 mmHg (n = 10) and low responders with that < 13 mmHg (n = 9). Counts of both total cells and alveolar macrophages in BALF did not increase during the therapy. However, total protein and surfactant protein-A (SP-A) were significantly decreased in high responders, but not in low responders, suggesting that clearance of surfactant materials is correlated with the efficacy of the therapy. Among 94 biomarkers screened in bronchoalveolar lavage fluid, we found that the concentration of interleukin-17 and cancer antigen-125 were significantly increased after GM-CSF inhalation treatment. GM-CSF inhalation decreased the concentration of total protein and SP-A in BALF, and increase interleukin-17 and cancer antigen-125 in improved lung of autoimmune pulmonary alveolar proteinosis.

Surfactant protein A is defective in abrogating inflammation in asthma

Surfactant protein A (SP-A) regulates a variety of immune cell functions. We determined the ability of SP-A derived from normal and asthmatic subjects to modulate the inflammatory response elicited by Mycoplasma pneumoniae, a pathogen known to exacerbate asthma. Fourteen asthmatic and 10 normal control subjects underwent bronchoscopy with airway brushing and bronchoalveolar lavage (BAL). Total SP-A was extracted from BAL. The ratio of SP-A1 to total SP-A (SP-A1/SP-A) and the binding of total SP-A to M. pneumoniae membranes were determined. Airway epithelial cells from subjects were exposed to either normal or asthmatic SP-A before exposure to M. pneumoniae. IL-8 protein and MUC5AC mRNA were measured. Total BAL SP-A concentration did not differ between groups, but the percentage SP-A1 was significantly increased in BAL of asthmatic compared with normal subjects. SP-A1/SP-A significantly correlated with maximum binding of total SP-A to M. pneumoniae, but only in asthma. SP-A derived from asthmatic subjects did not significantly attenuate IL-8 and MUC5AC in the setting of M. pneumoniae infection compared with SP-A derived from normal subjects. We conclude that SP-A derived from asthmatic subjects does not abrogate inflammation effectively, and this dysfunction may be modulated by SP-A1/SP-A.

Development and validation of a sandwich ELISA for use in measuring concentrations of canine surfactant protein A in serum of dogs

To develop and evaluate a sandwich ELISA incorporating rabbit antiserum specific for canine surfactant protein A (SP-A) for use in measuring concentrations of SP-A in serum of dogs. Serum samples obtained from 6 healthy dogs and 3 dogs with pulmonary disease. Rabbit antiserum was prepared against purified canine SP-A. The IgG fraction was isolated via protein G affinity chromatography and was then biotinylated. The sandwich ELISA was performed by use of anti-SP-A antibody (IgG) preabsorbed with sera from healthy dogs. Validity of the ELISA was confirmed by determination of the detection limit, precision, reproducibility, and accuracy. Serum SP-A concentrations were measured in 6 healthy dogs and 3 dogs with pulmonary disease. Detection limit of the ELISA was 2.0 ng/mL. Within- and between-assay coefficients of variation ranged from 3.8% to 14.1% and from 15.5% to 35.6%, respectively. The observed-to-expected recovery ratio ranged from 77.1% to 89.9%. Serum SP-A concentrations measured by use of the ELISA were ≤ 2.3 ng/mL in the 6 healthy dogs, 25.6 ng/mL in a dog with severe cardiac pulmonary edema, 8.3 ng/mL in a dog with pneumonia, and 10.1 ng/mL in a dog with lung lobe torsion. The sandwich ELISA was found to be useful for measuring purified canine SP-A concentrations and canine SP-A concentrations in serum samples. The ELISA was precise, reproducible, and accurate. The ELISA may be beneficial in assessing serum concentrations of canine SP-A as a potential biomarker of pulmonary diseases in dogs.

Assessment of Surfactant Protein A (SP-A) dependent agglutination

BackgroundMonomers of the collectin surfactant associated protein-A (SP-A) are arranged in trimers and higher oligomers. The state of oligomerization differs between individuals and likely affects SP-A's functional properties. SP-A can form aggregates together with other SP-A molecules. Here we report and assess a test system for the aggregate forming properties of SP-A in serum and broncho-alveolar lavage samples.MethodsAnti-SP-A antibodies fixed to latex beads bound SP-A at its N-terminal end and allowed the interaction with other SP-A molecules in a given sample by their C-terminal carbohydrate recognition domain (CRD) to agglutinate the beads to aggregates, which were quantified by light microscopy.ResultsSP-A aggregation was dependent on its concentration, the presence of calcium, and was dose-dependently inhibited by mannose. Unaffected by the presence of SP-D no aggregation was observed in absence of SP-A. The more complex the oligomeric structure of SP-A present in a particular sample, the better was its capability to induce aggregation at a given total concentration of SP-A. SP-A in serum agglutinated independently of the pulmonary disease; in contrast SP-A in lung lavage fluid was clearly inferior in patients with chronic bronchitis and particularly with cystic fibrosis compared to controls.ConclusionsThe functional status of SP-A with respect to its aggregating properties in serum and lavage samples can be easily assessed. SP-A in lung lavage fluid in patients with severe neutrophilic bronchitis was inferior.

Surfactant Protein-A as an Anti-Inflammatory Component in the Amnion: Implications for Human Pregnancy

The mechanism of mouse parturition is thought to involve myometrial infiltration by amniotic fluid (AF) macrophages, activated by surfactant protein-A (SP-A). In humans, the concentration of AF SP-A decreases during labor, and no fetal macrophages are found in the myometrium after labor. Therefore, it appears that the mechanisms of labor in mice and humans are different. We investigated a potential role for SP-A in human pregnancy and parturition by examining SP-A expression patterns in AF and amnion. High molecular mass (>250 kDa) oligomeric SP-A was increased in AF with advancing gestation. Interestingly, these oligomers were more abundant in placental amnion before labor at term, while they increased primarily in reflected amnion during labor (p < 0.05). Immunoblotting showed a binding of high molecular mass SP-A in AF to amnion. In C57BL/6 mice, oligomeric SP-A was also readily detected in AF from E15 onwards, but not in amnion. Macrophage density in mice myometrium did not change with advancing gestational age. Microarray analysis of human amnion explants incubated with SP-A revealed a molecular signature of inhibited cytokine-cytokine receptor interaction with downregulation of IL-1beta, CXCL2, and CXCL5 mRNA expression. The findings in this study strongly suggest that SP-A signals amniotic anti-inflammatory response via AF during pregnancy. We propose that an SP-A interaction among AF, placental amnion, and reflected amnion is a unique mechanism for immunoregulation in human pregnancy akin to that established in lung biology. However, AF SP-A and fetal macrophages by themselves do not seem to be exclusive effectors of parturition in humans.

Surfactant protein A (SP-A) binds to phosphatidylserine and competes with annexin V binding on late apoptotic cells

The role of surfactant protein A (SP-A) in the recognition and clearance of apoptotic cells is well established, but to date, it is still not clear which surface molecules of apoptotic cells are involved in the process. Here we present evidence that phosphatidylserine (PS) is a relevant binding molecule for human SP-A. The binding is Ca(2+)-dependent and is not inhibited by mannose, suggesting that the sugar-binding site of the carbohydrate recognition domain (CRD) of SP-A is not involved. Flow cytometry studies on apoptotic Jurkat cells revealed apparent inhibition of annexin V binding by increasing concentrations of SP-A in late apoptotic but not early apoptotic cells, and this was consistent for Jurkat cells and neutrophils. Supporting these data, confocal microscopy results show a co-localisation of annexin V and SP-A in late apoptotic but not early apoptotic cells. However, we cannot conclude that this inhibition is exclusively due to the binding of SP-A to PS on the cell surface, as annexin V is not wholly specific for PS and SP-A also interacts with other phospholipids that might become exposed on the apoptotic cell surface.