Sex differences in the acute in vivo effects of different human SP-A variants on the mouse alveolar macrophage proteome

Phelps, David S.; Umstead, Todd M.; Floros, Joanna

doi:10.1016/j.jprot.2014.06.007

Cited by 27 publications

(71 citation statements)

References 125 publications

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“…The mice were returned to their cages after recovery from anesthesia. In previous studies [21,22,26] we have found this method to be very consistent and reproducible for introducing SP-A (and other fluids) to the lungs.…”

Section: Treatment Of Mice With Exogenous Sp-a1mentioning

confidence: 73%

“…The exogenous SP-A1 preparation contained SP-A1 (10 μg) in 50 μl of sterile saline with 1 mM CaCl 2 . We have used this dose of exogenous SP-A in previous rescue studies [26]. Control animals received 50 μl of vehicle (saline and 1 mM CaCl 2 ) alone.…”

Section: Treatment Of Mice With Exogenous Sp-a1mentioning

confidence: 99%

“…In humans, however, unlike in rodents, there are two genes, Sftpa1 and Sftpa2, encoding SP-A1 and SP-A2, respectively, and these two gene products have been shown to have a differential impact on several AM functions. These functions include bacterial phagocytosis and cytokine production by AM [18,24,25], actin polymerization in the AM [7], and effects on the AM proteome and miRNome [26][27][28]. Moreover, the effects of SP-A variants on the regulation of the AM proteome and miRNome, survival, and pulmonary mechanics after infection, vary with sex [26][27][28][29][30].…”

Section: Introductionmentioning

confidence: 99%

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Using toponomics to characterize phenotypic diversity in alveolar macrophages from male mice treated with exogenous SP-A1

et al. 2020

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Background: We used the Toponome Imaging System (TIS) to identify "patterns of marker expression", referred to here as combinatorial molecular phenotypes (CMPs) in alveolar macrophages (AM) in response to the innate immune molecule, SP-A1. Methods: We compared 114 AM from male SPA deficient mice. One group (n = 3) was treated with exogenous human surfactant protein A1 (hSP-A1) and the other with vehicle (n = 3). AM obtained by bronchoalveolar lavage were plated onto slides and analyzed using TIS to study the AM toponome, the spatial network of proteins within intact cells. With TIS, each slide is sequentially immunostained with multiple FITC-conjugated antibodies. Images are analyzed pixel-by-pixel identifying all of the proteins within each pixel, which are then designated as CMPs. CMPs represent organized protein clusters postulated to contribute to specific functions. Results: 1) We compared identical CMPs in KO and SP-A1 cells and found them to differ significantly (p = 0.0007). Similarities between pairs of markers in the two populations also differed significantly (p < 0.0001). 2) Focusing on the 20 most abundant CMPs for each cell, we developed a method to generate CMP "signatures" that characterized various groups of cells. Phenotypes were defined as cells exhibiting similar signatures of CMPs. i) AM were extremely diverse and each group contained cells with multiple phenotypes. ii) Among the 114 AM analyzed, no two cells were identical. iii) However, CMP signatures could distinguish among cell subpopulations within and between groups. iv) Some cell populations were enriched with SP-A1 treatment, some were more common without SP-A1, and some seemed not to be influenced by the presence of SP-A1. v) We also found that AM were more diverse in mice treated with SP-A1 compared to those treated with vehicle. Conclusions: AM diversity is far more extensive than originally thought. The increased diversity of SP-A1-treated mice points to the possibility that SP-A1 enhances or activates several pathways in the AM to better prepare it for its innate immune functions and other functions shown previously to be affected by SPA treatment. Future studies may identify key protein(s) responsible for CMP integrity and consequently for a given function, and target it for therapeutic purposes.

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Section: Treatment Of Mice With Exogenous Sp-a1mentioning

confidence: 73%

Section: Treatment Of Mice With Exogenous Sp-a1mentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Using toponomics to characterize phenotypic diversity in alveolar macrophages from male mice treated with exogenous SP-A1

et al. 2020

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“…Both SP-A1 and SP-A2 genes are very similar in sequence, and the gene-specific SP-A1 and SP-A2 variants are distinguished by four amino acids within the collagen-like domain (11,35); the SP-A1 and SP-A2 variants carry a few more amino acid differences that are not gene-specific and these are located in other regions of the molecule, except the neck region, which is conserved among all variants. Functional, structural, and biophysical differences between SP-A1 and SP-A2 have been observed (14,(36)(37)(38)(39)(40)(41). It has been previously described that residue 85, one of the key differences between SP-A1 and SP-A2, is a cysteine in SP-A1, whereas it is an arginine in SP-A2.…”

Section: Introductionmentioning

confidence: 99%

Human Pulmonary Surfactant Protein SP-A1 Provides Maximal Efficiency of Lung Interfacial Films

López-Rodríguez

Pascual

Arroyo

et al. 2016

Biophysical Journal

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Pulmonary surfactant is a lipoprotein complex that reduces surface tension to prevent alveolar collapse and contributes to the protection of the respiratory surface from the entry of pathogens. Surfactant protein A (SP-A) is a hydrophilic glycoprotein of the collectin family, and its main function is related to host defense. However, previous studies have shown that SP-A also aids in the formation and biophysical properties of pulmonary surfactant films at the air-water interface. Humans, unlike rodents, have two genes, SFTPA1 and SFTPA2. The encoded proteins, SP-A1 and SP-A2, differ quantitatively or qualitatively in function. It has been shown that both gene products are necessary for tubular myelin formation, an extracellular structural form of lung surfactant. The goal of this study was to investigate potential differences in the biophysical properties of surfactants containing human SP-A1, SP-A2, or both. For this purpose, we have studied for the first time, to our knowledge, the biophysical properties of pulmonary surfactant from individual humanized transgenic mice expressing human SP-A1, SP-A2, or both SP-A1 and SP-A2, in the captive bubble surfactometer. We observed that pulmonary surfactant containing SP-A1 reaches lower surface tension after postexpansion interfacial adsorption than surfactants containing no SP-A or only SP-A2. Under interfacial compression-expansion cycling conditions, surfactant films containing SP-A1 also performed better, particularly with respect to the reorganization of the films that takes place during compression. On the other hand, addition of recombinant SP-A1 to a surfactant preparation reconstituted from the hydrophobic fraction of a porcine surfactant made it more resistant to inhibition by serum than the addition of equivalent amounts of SP-A2. We conclude that the presence of SP-A1 allows pulmonary surfactant to adopt a particularly favorable structure with optimal biophysical properties.

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“…Moreover, the levels of 14-3-3 isoforms ␤/␣, ε, ␥, and /␦ have been shown to change in the alveolar macrophage proteome in response to a single in vivo treatment with exogenous SP-A1 and/or SP-A2 in SP-A knockout mice. The changes were more pronounced when SP-A1 and SP-A2 were administered together than when SP-A1 was administered alone (39). Further experimentation is needed to decipher the underlying mechanisms.…”

Section: Discussionmentioning

confidence: 99%

14-3-3 isoforms bind directly exon B of the 5′-UTR of human surfactant protein A2 mRNA

Noutsios

Ghattas

Bennett

et al. 2015

American Journal of Physiology-Lung Cellular and Molecular Phys

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Human surfactant protein (SP) A (SP-A), an innate immunity molecule, is encoded by two genes, SFTPA1 and SFTPA2. The 5'-untranslated splice variant of SP-A2 (ABD), but not SP-A1 (AD), contains exon B (eB). eB is an enhancer for transcription and translation and contains cis-regulatory elements. Specific trans-acting factors, including 14-3-3, bind eB. The 14-3-3 protein family contains seven isoforms that have been found by mass spectrometry in eB electromobility shift assays (Noutsios et al. Am J Physiol Lung Cell Mol Physiol 304: L722-L735, 2013). We used four different approaches to investigate whether 14-3-3 isoforms bind directly to eB. 1) eB RNA pulldown assays showed that 14-3-3 isoforms specifically bind eB. 2) RNA electromobility shift assay complexes were formed using purified 14-3-3 isoforms β, γ, ε, η, σ, and τ, but not isoform ζ, with wild-type eB RNA. 3 and 4) RNA affinity chromatography assays and surface plasmon resonance analysis showed that 14-3-3 isoforms β, γ, ε, η, σ, and τ, but not isoform ζ, specifically and directly bind eB. Inhibition of 14-3-3 isoforms γ, ε, η, and τ/θ with shRNAs in NCI-H441 cells resulted in downregulation of SP-A2 levels but did not affect SP-A1 levels. However, inhibition of 14-3-3 isoform σ was correlated with lower levels of SP-A1 and SP-A2. Inhibition of 14-3-3 isoform ζ/δ, which does not bind eB, had no effect on expression levels of SP-A1 and SP-A2. In conclusion, the 14-3-3 protein family affects differential regulation of SP-A1 and SP-A2 by binding directly to SP-A2 5'-UTR mRNA.

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Sex differences in the acute in vivo effects of different human SP-A variants on the mouse alveolar macrophage proteome

Cited by 27 publications

References 125 publications

Using toponomics to characterize phenotypic diversity in alveolar macrophages from male mice treated with exogenous SP-A1

Using toponomics to characterize phenotypic diversity in alveolar macrophages from male mice treated with exogenous SP-A1

Human Pulmonary Surfactant Protein SP-A1 Provides Maximal Efficiency of Lung Interfacial Films

14-3-3 isoforms bind directly exon B of the 5′-UTR of human surfactant protein A2 mRNA

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