Expression of TGF-β1 and TGF-β1 receptor.

<p>Liver lysates obtained from the NZB/W F1 mice receiving PBS, NS1, VP1u or VP2 were probed with antibodies against (A) TGF-<i>β</i>1 and TGF-β1 receptor, respectively. Bars represent the relative protein quantification of (B) TGF-β1 and (C) TGF-β1 on the basis of β-actin. Serum level of (D) TGF-β1 was determined using an EIA kit. Similar results were observed in three independent experiments, and * indicates the significant difference, P<0.05.</p

Schematic illustration of the molecular mechanisms involved in the B19 NS1-induced hepatic fibrosis.

<p>B19 NS1 increases the expression of TGF-β isoforms, which elicit late activation of Smad2/3 signaling. Activated Smad2/3 cooperates with Smad4 and Sp1 to induce the expressions of fiboriss related proteins, including PAI-1 and smooth-muscle-specific (á-SMA).</p

Representative photographs of cross-sections of liver with trichrome staining.

<p>Fibrosis area (blue) in liver section from NZB/W F1 mice receiving NS1 were revealed as compared to those mice receiving PBS, VP1u or VP2, respectively. (magnification, 200). Similar results were observed in three independent experiments.</p

Exacerbating Effects of Human Parvovirus B19 NS1 on Liver Fibrosis in NZB/W F1 Mice

<div><p>Systemic lupus erythematosus (SLE) is an autoimmune disorder with unknown etiology that impacts various organs including liver. Recently, human parvovirus B19 (B19) is recognized to exacerbate SLE. However, the effects of B19 on liver in SLE are still unclear. Herein we aimed to investigate the effects of B19 on liver in NZB/W F1 mice by injecting subcutaneously with PBS, recombinant B19 NS1, VP1u or VP2, respectively. Our experimental results revealed that B19 NS1 protein significantly enhanced the TGF-β/Smad fibrotic signaling by increasing the expressions of TGF-β, Smad2/3, phosphorylated Smad2/3, Smad4 and Sp1. The consequent fibrosis-related proteins, PAI-1 and α-SMA, were also significantly induced in livers of NZB/W F1 mice receiving B19 NS1 protein. Accordingly, markedly increased collagen deposition was also observed in livers of NZB/W F1 mice receiving B19 NS1 protein. However, no significant difference was observed in livers of NZB/W F1 mice receiving B19 VP1u or VP2 as compared to the controls. These findings indicate that B19 NS1 plays a crucial role in exacerbating liver fibrosis in NZB/W F1 mice through enhancing the TGF-â/Smad fibrotic signaling.</p></div

Expression of á-SMA.

<p>Liver lysates obtained from the NZB/W F1 mice receiving PBS, NS1, VP1u or VP2 were probed with antibodies against (A)á-SMA. Bars represent the relative protein quantification of (B)á-SMA on the basis of β-actin. Similar results were observed in three independent experiments, and * indicates the significant difference, P<0.05.</p

Expression of iNOS and COX-2.

<p>Liver lysates obtained from the NZB/W F1 mice receiving PBS, NS1, VP1u or VP2 were probed with antibodies against (A) iNOS and (B) COX-2. Bars represent the relative protein quantification of (A) iNOS and (B) COX-2 on the basis of β-actin. Similar results were observed in three independent experiments, and * indicates the significant difference, P<0.05.</p

Expression of TNF-α and TNF-α receptor.

<p>Liver lysates obtained from the NZB/W F1 mice receiving PBS, NS1, VP1u or VP2 were probed with antibodies against (A) TNF-α and (B) TNF-α receptor. Bars represent the relative protein quantification of (A) TNF-α and (B) TNF-α receptor on the basis of β-actin. (C) Serum TNF-αwas determined with ELISA kit as described in materials and methods (S1). Similar results were observed in three independent experiments, and * indicates the significant difference, P<0.05.</p

Expression of Smad2/3, phosphorylated Smad2/3 (p-Smad2/3) and Smad4.

<p>Liver lysates obtained from the NZB/W F1 mice receiving PBS, NS1u, VP1u or VP2 were probed with antibodies against (A) smad2/3, phosphorylated smad2/3 and smad4, respectively. Bars represent the relative protein quantification of (B) p-smad2/3/on the basis of Smad2/3 and (C) Smad4 on the basis of β-actin. Similar results were observed in three independent experiments, and * indicates the significant difference, P<0.05.</p

Expression of PAI-1.

<p>Liver lysates obtained from the NZB/W F1 mice receiving PBS, NS1, VP1u or VP2 were probed with antibodies against (A) PAI-1. Bars represent the relative protein quantification of (B) PAI-1 on the basis of β-actin. Serum level of (C) PAI-I was determined using an ELISA kit. Similar results were observed in three independent experiments, and * indicates the significant difference, P<0.05.</p

Expression of Sp1.

<p>Liver lysates obtained from the NZB/W F1 mice receiving PBS, NS1, VP1u or VP2 were probed with antibodies against (A) Sp1. Bars represent the relative protein quantification of (B) Sp1 on the basis of β-actin. Similar results were observed in three independent experiments, and * indicates the significant difference, P<0.05.</p