Dataset Information

The Pathogenic Aspects of Human Parvovirus B19 NS1 Protein in Chronic and Inflammatory Diseases.

ABSTRACT:

Background

The nonstructural protein (NS1) of human parvovirus B19 (hPVB19) is considered to be a double-edged sword in its pathogenesis. NS1 protein promotes cell death by apoptosis in erythroid-lineage cells and is also implicated in triggering and the progression of various inflammation and autoimmune disorders.

Objectives

We investigated the possible role of hPVB19 NS1 in the modulation of proinflammatory cytokines in nonpermissive HEK-293T cells.

Methods

A plasmid containing the fully sequenced NS1 gene (pCMV6-AC-GFP-NS1) was transfected into HEK-293T cells. Transfection efficiency was assessed by fluorescent microscopy over time. Mock (pCMV6-AC-GFP) transfected cells were used as controls. The percentage of apoptotic cells was measured by flow cytometry at 24, 48, and 72 h posttransfection. Interleukin 6 (IL-6) mRNA, as a pleiotropic cytokine, was measured by real-time PCR. Furthermore, cellular supernatants were collected to determine the type and quantity of cytokines produced by mock- and NS1-transfected cells using flow cytometry.

Results

Fold change in the expression level of IL-6 mRNA in transfected cells after 72 hr of incubation was found to be 3.01 when compared with mock-transfected cells; however, cell apoptosis did not happen over time. Also, the concentration of cytokines such as IL-2, IL-6, IL-9, IL-17A, IL-21, IL-22, interferon (IFN)-γ, and tumor necrosis factor α (TNF-α) increased in NS1-transfected cells.

Conclusions

Overall, our results indicated that proinflammatory cytokine levels had increased following the expression of hPVB19 NS1 in HEK-293T cells, consistent with a role for NS1 expression facilitating the upregulation of inflammatory reactions. Therefore, hPVB19 NS1 function may play a role in the progression of some chronic and inflammatory diseases.

SUBMITTER: Jalali S

PROVIDER: S-EPMC9192293 | biostudies-literature | 2022

REPOSITORIES: biostudies-literature

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Publications

The Pathogenic Aspects of Human Parvovirus B19 NS1 Protein in Chronic and Inflammatory Diseases.

Jalali Sedigheh S Farhadi Ali A Rafiei Dehbidi Gholamreza G Farjadian Shirin S Sharifzadeh Sedigheh S Ranjbaran Reza R Seyyedi Noorossadat N Namdari Sepide S Behzad-Behbahani Abbas A

Interdisciplinary perspectives on infectious diseases 20220606

<h4>Background</h4>The nonstructural protein (NS1) of human parvovirus B19 (hPVB19) is considered to be a double-edged sword in its pathogenesis. NS1 protein promotes cell death by apoptosis in erythroid-lineage cells and is also implicated in triggering and the progression of various inflammation and autoimmune disorders.<h4>Objectives</h4>We investigated the possible role of hPVB19 NS1 in the modulation of proinflammatory cytokines in nonpermissive HEK-293T cells.<h4>Methods</h4>A plasmid cont ...[more]

PMID: 35707129

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Project description:Human parvovirus B19 (B19V) has been strongly associated with a variety of inflammatory disorders, such as rheumatoid arthritis (RA), inflammatory bowel disease and systemic lupus erythematosus. Non‑structural protein 1 (NS1) of B19V has been demonstrated to play essential roles in the pathological processes of B19V infection due to its regulatory properties on inflammatory cytokines. Celastrol, a quinone methide isolated from Tripterygium wilfordii, has displayed substantial potential in treating inflammatory diseases, such as psoriasis and RA. However, little is known about the effects of celastrol on B19V NS1‑induced inflammation. Therefore, cell viability assay, migration assay, phagocytosis analysis, zymography assay, ELISA and immunoblotting were conducted to verify the influences of celastrol on macrophages. The present study reported the attenuating effects of celastrol on B19V NS1‑induced inflammatory responses in macrophages derived from human acute monocytic leukemia cell lines, U937 and THP‑1. Although the migration was not significantly decreased by celastrol in both U937 and THP‑1 macrophages, significantly decreased viability, migration and phagocytosis were detected in both B19V NS1‑activated U937 and THP‑1 macrophages in the presence of celastrol. Additionally, celastrol significantly decreased MMP‑9 activity and the levels of inflammatory cytokines, including IL‑6, TNF‑α and IL‑1β, in B19V NS1‑activated U937 and THP‑1 cells. Notably, significantly decreased levels of NLR family pyrin domain‑containing 3, apoptosis‑associated speck‑like, caspase‑1 and IL‑18 proteins were observed in both B19V NS1‑activated U937 and THP‑1 cells in the presence of celastrol, indicating the involvement of the inflammasome pathway. To the best of our knowledge, the present study is the first to report on the attenuating effects of celastrol on B19V NS1‑induced inflammatory responses in macrophages, suggesting a therapeutic role for celastrol in B19V NS1‑related inflammatory diseases.

Project description:Parvovirus B19 infections are associated with diverse clinical manifestations, ranging from no symptoms to severe symptoms. The virus shows an extreme tropism for replication in erythroid progenitor cells, possibly due to the activity of the only functional promoter (p6) of the B19 virus genome in combination with both cell- and cell cycle-specific factors and the trans-activator protein NS1. As presented here, p6 promoter sequences derived from several B19 virus isolates proved to be highly conserved. Furthermore, mutations did not affect any of the potential binding sites for transcription factors. One variation of the base at position 223 was identified only in B19 virus isolates derived from patients with persistent infection or chronic arthritis. To determine promoter activity and to characterize regulatory elements, sequences spanning the total p6 promoter and subfragments of them were introduced into a eukaryotic expression vector upstream of the luciferase gene (from Photinus pyralis). After transfection into HeLa, CEM, BJAB, and K562 cells, the p6 promoter was found to be highly active. When introduced into the erythroid cell line K562, p6-controlled transcription exceeded that of the simian virus 40 promoter-enhancer used as a control by more than 25-fold. Sequence elements relevant for promoter activity mapped to the regions from nucleotides (nt) 100 to 190 and 233 to 298. Also, the segment from nt 343 to 400 downstream of the TATA box was important for transcriptional activity in HeLa and K562 cells. By transfecting the promoter-luciferase constructs into a HeLa cell line stably carrying the viral NS1 gene under the control of an inducible promoter, transcriptional activity mediated by the p6 promoter rose significantly after induction of NS1 expression. The region from nt 100 to 160 proved to be essential for NS1-mediated transcriptional activation. Furthermore, NS1-mediated transactivation was dependent on the presence of two GC-rich elements arranged in tandem upstream of the TATA box. These data indicate that NS1-mediated p6 transactivation is dependent on a multicomponent complex combining NS1 with ATF, NF-kappaB/c-Rel, and GC-box binding cellular factors.

Dataset Information

The Pathogenic Aspects of Human Parvovirus B19 NS1 Protein in Chronic and Inflammatory Diseases.

Background

Objectives

Methods

Results

Conclusions

Publications

The Pathogenic Aspects of Human Parvovirus B19 NS1 Protein in Chronic and Inflammatory Diseases.

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