Inhibitory effect of aqueous Dandelion extract on HIV-1 replication and reverse transcriptase activity

Abstract

Background: Acquired immunodeficiency syndrome (AIDS), which is caused by the human immunodeficiency virus (HIV), is an immunosuppressive disease that results in life-threatening opportunistic infections. The general problems in current therapy include the constant emergence of drug-resistant HIV strains, adverse side effects and the unavailability of treatments in developing countries. Natural products from herbs with the abilities to inhibit HIV-1 life cycle at different stages, have served as excellent sources of new anti-HIV-1 drugs. In this study, we aimed to investigate the anti-HIV-1 activity of aqueous dandelion extract.

Methods: The pseudotyped HIV-1 virus has been utilized to explore the anti-HIV-1 activity of dandelion, the level of HIV-1 replication was assessed by the percentage of GFP-positive cells. The inhibitory effect of the dandelion extract on reverse transcriptase activity was assessed by the reverse transcriptase assay kit.

Results: Compared to control values obtained from cells infected without treatment, the level of HIV-1 replication and reverse transcriptase activity were decreased in a dose-dependent manner. The data suggest that dandelion extract has a potent inhibitory activity against HIV-1 replication and reverse transcriptase activity. The identification of HIV-1 antiviral compounds from Taraxacum officinale should be pursued.

Conclusions: The dandelion extract showed strong activity against HIV-1 RT and inhibited both the HIV-1 vector and the hybrid-MoMuLV/MoMuSV retrovirus replication. These findings provide additional support for the potential therapeutic efficacy of Taraxacum officinale. Extracts from this plant may be regarded as another starting point for the development of an antiretroviral therapy with fewer side effects.

Figure 1

Construction of retroviral and HIV-1 replication reporter vector system. (A) The pLNCX2 -EGFP vector, constructed by inserting the EGFP gene into the multiple cloning site of the pLNCX2 plasmid, a retroviral vector contains elements derived from Moloney murine leukemia virus (MoMuLV) and Moloney murine sarcoma virus (MoMuSV). The HIV-1 based lentiviral vector pLL3.7 is commercially available, and EGFP is driven by the CMV promoter. (B) The development of stable RetroPackPT67 packaging cells encoding the EGFP genes, which was used as the EGFP virus-producing cell line. (C) Outline of the anti-HIV-1 activity assay for DWE.

Figure 2

Cytotoxicity assay for different cells cultured with different concentrations of DWE. (A) NIH/3T3 cell survival curves from the CCK-8 assay. The negative control was untreated cells, and the blank control contained the Cell Counting Kit-8 reagent and no cells. The curve represents the average of three independent experiments with SD bars. (B) PBMCs were incubated at 37°C in 5% CO₂ overnight in media supplemented with IL-2 at a concentration of 100 IU/ml. The cells were then exposed to the DWE for 24 h, and aliquots of cells were stained with trypan blue dye to determine the total numbers of cells as well as viable cells. PBMC cells, cultured in the absence of DWE were used as control. These data represent three independent experiments.

Figure 3

Fluorescence micrograph of GFP proteins in infected cells treated with DWE. NIH/3T3 cells expressing EGFP following infection with VSV-G pseudotyped HIV-1 virus. (A) NIH/3T3 cells infected with the pLL3.7 lentivirus without DWE (untreated). Figures show NIH/3T3 cells infected with the pLL3.7 lentivirus in the presence of DWE at a concentration of (B) 0.25 mg/ml (C) 0.5 mg/ml (D) 1 mg/ml. In the upper panels, cells are visualized under normal light, while in the lower panels, the same cells are visualized by fluorescent microscope.

Figure 4

Inhibitory effects of DWE on HIV-1 and another retrovirus. Figure A and B shows the effects of DWE on HIV-1 and another retrovirus pLNCX2 -EGFP replication. Gene expression of NIH/3T3 cells were infected with two pseudotyped retroviruses for 12 h and treated with different concentrations of DWE. Different concentrations of AZT (from 0 μM to 10 μM) and aqueous Herba Artemisiae Scopariae extract (HASWE) were used as positive control and negative control respectively. (A) NIH/3T3 cells infected with VSV-G pseudotyped HIV-1; (B) NIH/3T3 cells infected with MoMuLV/MoMuSV hybrid retrovirus; (C) Percentage of inhibition of HIV-1 RT by DWE. 2.5 μM AZT was used as the positive control with 98% inhibitory rate while lysis buffer with no HIV-1 RT was used as one negative control, lysis buffer with HIV-1 RT but no DWE was used as another negative control. The data represent the mean ± SD of triplicate experiments

Figure 5

HPLC chromatograms. HPLC chromatogram of Taraxacum officinale and Herba Artemisiae Scopariae which are (1) caffeic acid, (2), chlorogenic acid.