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Expression of neuroimmune semaphorins 4A and 4D and their receptors in the lung is enhanced by allergen and vascular endothelial growth factor.

Smith EP, Shanks K, Lipsky MM, DeTolla LJ, Keegan AD, Chapoval SP - BMC Immunol. (2011)

Bottom Line: Sema4A and Sema4D were the first semaphorins found to be expressed on immune cells and were termed "immune semaphorins".We found that besides accessory-like cells, Sema4A was also detected on bronchial epithelial and smooth muscle cells, whereas Sema4D expression was high on immune cells such as T and B lymphocytes.CD72 was found on lung immune, inflammatory, and epithelial cells.

View Article: PubMed Central - HTML - PubMed

Affiliation: Center for Vascular and Inflammatory Diseases, University of Maryland School of Medicine, Baltimore, Maryland 21201, USA.

ABSTRACT

Background: Semaphorins were originally identified as molecules regulating a functional activity of axons in the nervous system. Sema4A and Sema4D were the first semaphorins found to be expressed on immune cells and were termed "immune semaphorins". It is known that Sema4A and Sema4D bind Tim-2 and CD72 expressed on leukocytes and PlexinD1 and B1 present on non-immune cells. These neuroimmune semaphorins and their receptors have been shown to play critical roles in many physiological and pathological processes including neuronal development, immune response regulation, cancer, autoimmune, cardiovascular, renal, and infectious diseases. However, the expression and regulation of Sema4A, Sema4D, and their receptors in normal and allergic lungs is undefined.

Results: Allergen treatment and lung-specific vascular endothelial growth factor (VEGF) expression induced asthma-like pathologies in the murine lungs. These experimental models of allergic airway inflammation were used for the expression analysis of immune semaphorins and their receptors employing immunohistochemistry and flow cytometry techniques. We found that besides accessory-like cells, Sema4A was also detected on bronchial epithelial and smooth muscle cells, whereas Sema4D expression was high on immune cells such as T and B lymphocytes. Surprisingly, under inflammation various cell types including macrophages, lymphocytes, and granulocytes in the lung expressed Tim-2, a previously defined marker for Th2 cells. CD72 was found on lung immune, inflammatory, and epithelial cells. Bronchial epithelial cells were positive for both plexins, whereas some endothelial cells selectively expressed Plexin D1. Plexin B1 expression was also detected on lung DC. Both allergen and VEGF upregulated the expression of neuroimmune semaphorins and their receptors in the lung tissue. However, the lung tissue Sema4A-Tim2 expression was rather weak, whereas Sema4D-CD72 ligand-receptor pair was vastly upregulated by allergen. Soluble Sema4D protein was present in the lung lysates and a whole Sema4A protein plus its dimer were readily detected in the bronchoalveolar (BAL) fluids under inflammation.

Conclusions: This study clearly shows that neuroimmune semaphorins Sema4A and Sema4D and their receptors might serve as potential markers for the allergic airway inflammatory diseases. Our current findings pave the way for further investigations of the role of immune semaphorins in inflammation and their use as potential therapeutic targets for the inflammatory lung conditions.

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Regulation of lung Tim-2 expression by allergen and VEGF. Immunohistochemical (A-B) and flow cytometric (C) detection of Tim-2 expression in lung (A) and lymphoid (B) tissues, and on lung T cells (C). (A-B) Note positive Tim-2 staining on different lung cells besides lymphocytes in allergen- or VEGF-exposed mouse lungs. Red arrows point to Tim-2+ APC-like cells and granulocyte. Inserts show high magnification fields (100x) with marker-positive cells (lymphocytes). (C) Flow cytometry analysis of cells from lung enzymatic digests obtained from PBS- and allergen-treated mice showed an increase in lung Tim-2+ T cells in OVA-treated mice. The histograms show the percentage of Tim-2+CD3+ cells in the lung (clear histogram) as compared to the appropriate isotype control stained CD3+ cells (gray histogram).
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Figure 4: Regulation of lung Tim-2 expression by allergen and VEGF. Immunohistochemical (A-B) and flow cytometric (C) detection of Tim-2 expression in lung (A) and lymphoid (B) tissues, and on lung T cells (C). (A-B) Note positive Tim-2 staining on different lung cells besides lymphocytes in allergen- or VEGF-exposed mouse lungs. Red arrows point to Tim-2+ APC-like cells and granulocyte. Inserts show high magnification fields (100x) with marker-positive cells (lymphocytes). (C) Flow cytometry analysis of cells from lung enzymatic digests obtained from PBS- and allergen-treated mice showed an increase in lung Tim-2+ T cells in OVA-treated mice. The histograms show the percentage of Tim-2+CD3+ cells in the lung (clear histogram) as compared to the appropriate isotype control stained CD3+ cells (gray histogram).

Mentions: As it has been shown previously, Tim-2 is a marker for activated T cells, preferentially Th2 cells [5-7], whereas CD72 is found to be expressed on B cells and DC [9-12]. To our surprise, using Ab for IHC specified in the Methods section, Tim-2 staining was detected on many cells in the lungs of allergen-treated mice (Figure 4 A) including inflammatory macrophages and a subset of granulocytes. This staining was specific when compared to isotype control Ab staining (Additional files, Figure 1). Some lymphocytes were also Tim-2-positive. In VEGF tg mice Tim-2 was targeted predominantly to the tissue lymphocytes with a weaker staining detected in macrophages and granulocytes (Figure 4 A). In contrast to the lung tissue, strong Tim-2 expression was readily detected in spleen and local lymph nodes of allergen-treated WT mice (Figure 4 B). Flow cytometry data had shown an increase in Tim-2-positive T cells in allergic lung (mean of fluorescence intensity from 7.94 to 25.5%, PBS- vs OVA-treated mice, respectively) (Figure 4 C). The physiological mean of Tim-2 expression on other cell types besides T cells has to be determined.


Expression of neuroimmune semaphorins 4A and 4D and their receptors in the lung is enhanced by allergen and vascular endothelial growth factor.

Smith EP, Shanks K, Lipsky MM, DeTolla LJ, Keegan AD, Chapoval SP - BMC Immunol. (2011)

Regulation of lung Tim-2 expression by allergen and VEGF. Immunohistochemical (A-B) and flow cytometric (C) detection of Tim-2 expression in lung (A) and lymphoid (B) tissues, and on lung T cells (C). (A-B) Note positive Tim-2 staining on different lung cells besides lymphocytes in allergen- or VEGF-exposed mouse lungs. Red arrows point to Tim-2+ APC-like cells and granulocyte. Inserts show high magnification fields (100x) with marker-positive cells (lymphocytes). (C) Flow cytometry analysis of cells from lung enzymatic digests obtained from PBS- and allergen-treated mice showed an increase in lung Tim-2+ T cells in OVA-treated mice. The histograms show the percentage of Tim-2+CD3+ cells in the lung (clear histogram) as compared to the appropriate isotype control stained CD3+ cells (gray histogram).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3118960&req=5

Figure 4: Regulation of lung Tim-2 expression by allergen and VEGF. Immunohistochemical (A-B) and flow cytometric (C) detection of Tim-2 expression in lung (A) and lymphoid (B) tissues, and on lung T cells (C). (A-B) Note positive Tim-2 staining on different lung cells besides lymphocytes in allergen- or VEGF-exposed mouse lungs. Red arrows point to Tim-2+ APC-like cells and granulocyte. Inserts show high magnification fields (100x) with marker-positive cells (lymphocytes). (C) Flow cytometry analysis of cells from lung enzymatic digests obtained from PBS- and allergen-treated mice showed an increase in lung Tim-2+ T cells in OVA-treated mice. The histograms show the percentage of Tim-2+CD3+ cells in the lung (clear histogram) as compared to the appropriate isotype control stained CD3+ cells (gray histogram).
Mentions: As it has been shown previously, Tim-2 is a marker for activated T cells, preferentially Th2 cells [5-7], whereas CD72 is found to be expressed on B cells and DC [9-12]. To our surprise, using Ab for IHC specified in the Methods section, Tim-2 staining was detected on many cells in the lungs of allergen-treated mice (Figure 4 A) including inflammatory macrophages and a subset of granulocytes. This staining was specific when compared to isotype control Ab staining (Additional files, Figure 1). Some lymphocytes were also Tim-2-positive. In VEGF tg mice Tim-2 was targeted predominantly to the tissue lymphocytes with a weaker staining detected in macrophages and granulocytes (Figure 4 A). In contrast to the lung tissue, strong Tim-2 expression was readily detected in spleen and local lymph nodes of allergen-treated WT mice (Figure 4 B). Flow cytometry data had shown an increase in Tim-2-positive T cells in allergic lung (mean of fluorescence intensity from 7.94 to 25.5%, PBS- vs OVA-treated mice, respectively) (Figure 4 C). The physiological mean of Tim-2 expression on other cell types besides T cells has to be determined.

Bottom Line: Sema4A and Sema4D were the first semaphorins found to be expressed on immune cells and were termed "immune semaphorins".We found that besides accessory-like cells, Sema4A was also detected on bronchial epithelial and smooth muscle cells, whereas Sema4D expression was high on immune cells such as T and B lymphocytes.CD72 was found on lung immune, inflammatory, and epithelial cells.

View Article: PubMed Central - HTML - PubMed

Affiliation: Center for Vascular and Inflammatory Diseases, University of Maryland School of Medicine, Baltimore, Maryland 21201, USA.

ABSTRACT

Background: Semaphorins were originally identified as molecules regulating a functional activity of axons in the nervous system. Sema4A and Sema4D were the first semaphorins found to be expressed on immune cells and were termed "immune semaphorins". It is known that Sema4A and Sema4D bind Tim-2 and CD72 expressed on leukocytes and PlexinD1 and B1 present on non-immune cells. These neuroimmune semaphorins and their receptors have been shown to play critical roles in many physiological and pathological processes including neuronal development, immune response regulation, cancer, autoimmune, cardiovascular, renal, and infectious diseases. However, the expression and regulation of Sema4A, Sema4D, and their receptors in normal and allergic lungs is undefined.

Results: Allergen treatment and lung-specific vascular endothelial growth factor (VEGF) expression induced asthma-like pathologies in the murine lungs. These experimental models of allergic airway inflammation were used for the expression analysis of immune semaphorins and their receptors employing immunohistochemistry and flow cytometry techniques. We found that besides accessory-like cells, Sema4A was also detected on bronchial epithelial and smooth muscle cells, whereas Sema4D expression was high on immune cells such as T and B lymphocytes. Surprisingly, under inflammation various cell types including macrophages, lymphocytes, and granulocytes in the lung expressed Tim-2, a previously defined marker for Th2 cells. CD72 was found on lung immune, inflammatory, and epithelial cells. Bronchial epithelial cells were positive for both plexins, whereas some endothelial cells selectively expressed Plexin D1. Plexin B1 expression was also detected on lung DC. Both allergen and VEGF upregulated the expression of neuroimmune semaphorins and their receptors in the lung tissue. However, the lung tissue Sema4A-Tim2 expression was rather weak, whereas Sema4D-CD72 ligand-receptor pair was vastly upregulated by allergen. Soluble Sema4D protein was present in the lung lysates and a whole Sema4A protein plus its dimer were readily detected in the bronchoalveolar (BAL) fluids under inflammation.

Conclusions: This study clearly shows that neuroimmune semaphorins Sema4A and Sema4D and their receptors might serve as potential markers for the allergic airway inflammatory diseases. Our current findings pave the way for further investigations of the role of immune semaphorins in inflammation and their use as potential therapeutic targets for the inflammatory lung conditions.

Show MeSH
Related in: MedlinePlus