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Local synthesis of interferon-alpha in lupus nephritis is associated with type I interferons signature and LMP7 induction in renal tubular epithelial cells.

Castellano G, Cafiero C, Divella C, Sallustio F, Gigante M, Pontrelli P, De Palma G, Rossini M, Grandaliano G, Gesualdo L - Arthritis Res. Ther. (2015)

Bottom Line: Type I interferons signature was characterized by MXA-specific staining in renal tubular epithelial cells; in addition, in situ hybridization showed that renal tubular epithelial cells were the major producers of interferon-alpha, indicating a potential autocrine effect.Whole-genome expression profile showed interferon-alpha induced up-regulation of genes involved in innate immunity, protein ubiquitination and switching to immunoproteasome.Our data indicate that type I interferons might have a pathogenic role in lupus nephritis characterized by an autocrine effect of interferon-alpha on renal tubular epithelial cells.

View Article: PubMed Central - PubMed

Affiliation: Renal, Dialysis and Transplantation Unit, Department of Emergency and Organ Transplantation, University of Bari, Piazza Giulio Cesare 11, 70124, Bari, Italy. giuseppe.castellano@uniba.it.

ABSTRACT

Introduction: Type I interferons are pivotal in the activation of autoimmune response in systemic lupus erythematous. However, the pathogenic role of interferon-alpha in patients affected by lupus nephritis remains uncertain. The aim of our study was to investigate the presence of a specific interferon signature in lupus nephritis and the effects of interferon-alpha at renal level.

Methods: We performed immunohistochemical analysis for MXA-protein and in situ hybridization to detect interferon-alpha signature and production in human lupus nephritis. Through microarray studies, we analyzed the gene expression profile of renal tubular epithelial cells, stimulated with interferon-alpha. We validated microarray results through real-time polymerase chain reaction, flow cytometry on renal tubular epithelial cells, and through immunohistochemical analysis and confocal microscopy on renal biopsies.

Results: Type I interferons signature was characterized by MXA-specific staining in renal tubular epithelial cells; in addition, in situ hybridization showed that renal tubular epithelial cells were the major producers of interferon-alpha, indicating a potential autocrine effect. Whole-genome expression profile showed interferon-alpha induced up-regulation of genes involved in innate immunity, protein ubiquitination and switching to immunoproteasome. In accordance with the in vitro data, class IV lupus nephritis showed up-regulation of the immunoproteasome subunit LMP7 in tubular epithelial cells associated with type I interferon signature.

Conclusions: Our data indicate that type I interferons might have a pathogenic role in lupus nephritis characterized by an autocrine effect of interferon-alpha on renal tubular epithelial cells. Therefore we hypothesize that inhibition of type I interferons might represent a therapeutic target to prevent tubulo-interstitial damage in patients with lupus nephritis.

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Immunoproteasome subunit LMP7 induction in tubular epithelial cells from SLE patients with nephritis. Increased expression of LMP7 was found in renal biopsies of patients with class IV lupus nephritis (B,D) compared to those with class I (A,C). LMP7 expression was found at the tubular interstitial level but not on glomerular cells (G) and vessels (V). Co-localization of LMP7 (red H, K) and MXA (green G, J) in tubular epithelium of patients with lupus nephritis was investigated by immunofluorescence/confocal microscopy (E-L). We found co-localization of LMP7 and MXA in class IV lupus nephritis (F,I,L) but not in class I, where MXA was absent and LMP7 had very low expression (E); nuclei were stained with TO-PRO-3 (blue); (L) zoom of figure F. (M) Quantification of immunohistochemical staining was carried out as described in Methods section. The histograms represent the increased tissue expressions of MXA (P <0.0001) and LMP7 proteins (P <0.0001) in eight class IV lupus nephritis patients compared to eight patients of class I/II. SLE, systemic lupus erythematous.
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Fig4: Immunoproteasome subunit LMP7 induction in tubular epithelial cells from SLE patients with nephritis. Increased expression of LMP7 was found in renal biopsies of patients with class IV lupus nephritis (B,D) compared to those with class I (A,C). LMP7 expression was found at the tubular interstitial level but not on glomerular cells (G) and vessels (V). Co-localization of LMP7 (red H, K) and MXA (green G, J) in tubular epithelium of patients with lupus nephritis was investigated by immunofluorescence/confocal microscopy (E-L). We found co-localization of LMP7 and MXA in class IV lupus nephritis (F,I,L) but not in class I, where MXA was absent and LMP7 had very low expression (E); nuclei were stained with TO-PRO-3 (blue); (L) zoom of figure F. (M) Quantification of immunohistochemical staining was carried out as described in Methods section. The histograms represent the increased tissue expressions of MXA (P <0.0001) and LMP7 proteins (P <0.0001) in eight class IV lupus nephritis patients compared to eight patients of class I/II. SLE, systemic lupus erythematous.

Mentions: Moreover, we investigated whether the genes and the relative proteins modulated in vitro by IFN-alpha were also detectable in patients affected by lupus nephritis. First, we investigated the presence of LMP7 by immunohistochemistry (Figure 4A to D). We observed that LMP7 was expressed at very low levels in tubules of patients with class I/II lupus nephritis, without any expression in glomeruli and blood vessels (Figure 4A and C). On the contrary, we found high LMP7 expression in class IV lupus nephritis, mainly localized at the tubulo-interstitial level (Figure 4B and D). Confocal microscopy analysis showed that the co-localization of MXA with LMP7 was present only in patients with class IV lupus nephritis (Figure 4F,I and L). Quantification of LMP7 and MXA protein expression by immunohistochemistry analysis demonstrated that the difference between the classes was statistically significant (Figure 4M).Figure 4


Local synthesis of interferon-alpha in lupus nephritis is associated with type I interferons signature and LMP7 induction in renal tubular epithelial cells.

Castellano G, Cafiero C, Divella C, Sallustio F, Gigante M, Pontrelli P, De Palma G, Rossini M, Grandaliano G, Gesualdo L - Arthritis Res. Ther. (2015)

Immunoproteasome subunit LMP7 induction in tubular epithelial cells from SLE patients with nephritis. Increased expression of LMP7 was found in renal biopsies of patients with class IV lupus nephritis (B,D) compared to those with class I (A,C). LMP7 expression was found at the tubular interstitial level but not on glomerular cells (G) and vessels (V). Co-localization of LMP7 (red H, K) and MXA (green G, J) in tubular epithelium of patients with lupus nephritis was investigated by immunofluorescence/confocal microscopy (E-L). We found co-localization of LMP7 and MXA in class IV lupus nephritis (F,I,L) but not in class I, where MXA was absent and LMP7 had very low expression (E); nuclei were stained with TO-PRO-3 (blue); (L) zoom of figure F. (M) Quantification of immunohistochemical staining was carried out as described in Methods section. The histograms represent the increased tissue expressions of MXA (P <0.0001) and LMP7 proteins (P <0.0001) in eight class IV lupus nephritis patients compared to eight patients of class I/II. SLE, systemic lupus erythematous.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4389585&req=5

Fig4: Immunoproteasome subunit LMP7 induction in tubular epithelial cells from SLE patients with nephritis. Increased expression of LMP7 was found in renal biopsies of patients with class IV lupus nephritis (B,D) compared to those with class I (A,C). LMP7 expression was found at the tubular interstitial level but not on glomerular cells (G) and vessels (V). Co-localization of LMP7 (red H, K) and MXA (green G, J) in tubular epithelium of patients with lupus nephritis was investigated by immunofluorescence/confocal microscopy (E-L). We found co-localization of LMP7 and MXA in class IV lupus nephritis (F,I,L) but not in class I, where MXA was absent and LMP7 had very low expression (E); nuclei were stained with TO-PRO-3 (blue); (L) zoom of figure F. (M) Quantification of immunohistochemical staining was carried out as described in Methods section. The histograms represent the increased tissue expressions of MXA (P <0.0001) and LMP7 proteins (P <0.0001) in eight class IV lupus nephritis patients compared to eight patients of class I/II. SLE, systemic lupus erythematous.
Mentions: Moreover, we investigated whether the genes and the relative proteins modulated in vitro by IFN-alpha were also detectable in patients affected by lupus nephritis. First, we investigated the presence of LMP7 by immunohistochemistry (Figure 4A to D). We observed that LMP7 was expressed at very low levels in tubules of patients with class I/II lupus nephritis, without any expression in glomeruli and blood vessels (Figure 4A and C). On the contrary, we found high LMP7 expression in class IV lupus nephritis, mainly localized at the tubulo-interstitial level (Figure 4B and D). Confocal microscopy analysis showed that the co-localization of MXA with LMP7 was present only in patients with class IV lupus nephritis (Figure 4F,I and L). Quantification of LMP7 and MXA protein expression by immunohistochemistry analysis demonstrated that the difference between the classes was statistically significant (Figure 4M).Figure 4

Bottom Line: Type I interferons signature was characterized by MXA-specific staining in renal tubular epithelial cells; in addition, in situ hybridization showed that renal tubular epithelial cells were the major producers of interferon-alpha, indicating a potential autocrine effect.Whole-genome expression profile showed interferon-alpha induced up-regulation of genes involved in innate immunity, protein ubiquitination and switching to immunoproteasome.Our data indicate that type I interferons might have a pathogenic role in lupus nephritis characterized by an autocrine effect of interferon-alpha on renal tubular epithelial cells.

View Article: PubMed Central - PubMed

Affiliation: Renal, Dialysis and Transplantation Unit, Department of Emergency and Organ Transplantation, University of Bari, Piazza Giulio Cesare 11, 70124, Bari, Italy. giuseppe.castellano@uniba.it.

ABSTRACT

Introduction: Type I interferons are pivotal in the activation of autoimmune response in systemic lupus erythematous. However, the pathogenic role of interferon-alpha in patients affected by lupus nephritis remains uncertain. The aim of our study was to investigate the presence of a specific interferon signature in lupus nephritis and the effects of interferon-alpha at renal level.

Methods: We performed immunohistochemical analysis for MXA-protein and in situ hybridization to detect interferon-alpha signature and production in human lupus nephritis. Through microarray studies, we analyzed the gene expression profile of renal tubular epithelial cells, stimulated with interferon-alpha. We validated microarray results through real-time polymerase chain reaction, flow cytometry on renal tubular epithelial cells, and through immunohistochemical analysis and confocal microscopy on renal biopsies.

Results: Type I interferons signature was characterized by MXA-specific staining in renal tubular epithelial cells; in addition, in situ hybridization showed that renal tubular epithelial cells were the major producers of interferon-alpha, indicating a potential autocrine effect. Whole-genome expression profile showed interferon-alpha induced up-regulation of genes involved in innate immunity, protein ubiquitination and switching to immunoproteasome. In accordance with the in vitro data, class IV lupus nephritis showed up-regulation of the immunoproteasome subunit LMP7 in tubular epithelial cells associated with type I interferon signature.

Conclusions: Our data indicate that type I interferons might have a pathogenic role in lupus nephritis characterized by an autocrine effect of interferon-alpha on renal tubular epithelial cells. Therefore we hypothesize that inhibition of type I interferons might represent a therapeutic target to prevent tubulo-interstitial damage in patients with lupus nephritis.

Show MeSH
Related in: MedlinePlus