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Anti-HIV-1 activity, protease inhibition and safety profile of extracts prepared from Rhus parviflora.

Modi M - BMC Complement Altern Med (2013)

Bottom Line: Both the extracts did not disturb the integrity of monolayer formed by intestinal epithelial Caco-2 cells.The extracts when tested up to 100 μg/ml did not significantly reduce the viability of L. plantarum, L. fermentum, L. rhamnosus and L. casei.The studies reported herein showed in vitro anti-HIV activity and preliminary safety profile of the extracts prepared from the leaves of R. parviflora.

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ABSTRACT

Background: In the present study, extracts prepared from the leaves of Rhus parviflora Roxb. (Anacardiaceae) were evaluated for their anti-HIV activity, which have been traditionally used for the treatment of neurological disorders such as anxiety, insomnia and epilepsy.

Methods: Aqueous and 50% ethanolic extracts prepared from leaves of the plant were tested for their cytotoxicity and anti-HIV property using reporter gene based assays as well as human peripheral blood lymphocytes (PBLs). Further these extracts were evaluated for their ability to inhibit HIV-1 reverse transcriptase (RT) and protease activity. Safety profile of the extracts was determined on viability of Lactobacillus sp., secretion of pro-inflammatory cytokines by vaginal keratinocytes and transepithelial resistance.

Results: Both aqueous (IC50 = 15 μg/ml) and 50% ethanolic (IC50 = 26 μg/ml) extracts prepared from leaves of R. parviflora showed anti-HIV activity in TZM-bl cells wherein the virus was treated with the extracts prior to infection. Further, both the extracts also inhibited virus load in HIV infected CEM-GFP cells and human PBLs. The anti-HIV activity is mediated through inhibition of HIV-1 protease activity. Both the extracts did not disturb the integrity of monolayer formed by intestinal epithelial Caco-2 cells. The extracts when tested up to 100 μg/ml did not significantly reduce the viability of L. plantarum, L. fermentum, L. rhamnosus and L. casei. The extracts (100 μg/ml) did not reveal any cytotoxic effect on vaginal keratinocytes (Vk2/E6E7). Levels of pro-inflammatory cytokines secreted by Vk2/E6E7 cells treated with both the plant extracts were within the non-inflammatory range.

Conclusions: The studies reported herein showed in vitro anti-HIV activity and preliminary safety profile of the extracts prepared from the leaves of R. parviflora.

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Reduction in virus load by leaves extracts of R. parviflora using CEM-GFP cells. Cells were infected with HIV-1NL4.3 (0.05 MOI) and treated with extracts at various concentrations followed by 8 day incubation. Culture supernatant was collected on 8th day for p24 estimation by ELISA. AZT (10 μM) was used as positive control that reduced p24 level by 97%. Data is represented as mean ± SE of three independent experiments performed in duplicates. Statistical significance between the extract treated groups as compared to untreated control is shown by asterisk (*p < 0.001).
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Figure 1: Reduction in virus load by leaves extracts of R. parviflora using CEM-GFP cells. Cells were infected with HIV-1NL4.3 (0.05 MOI) and treated with extracts at various concentrations followed by 8 day incubation. Culture supernatant was collected on 8th day for p24 estimation by ELISA. AZT (10 μM) was used as positive control that reduced p24 level by 97%. Data is represented as mean ± SE of three independent experiments performed in duplicates. Statistical significance between the extract treated groups as compared to untreated control is shown by asterisk (*p < 0.001).

Mentions: The inhibitory activity of the extracts against HIV infection is sometimes, a result of their toxic effects and consequently might result in an erroneous conclusion. Hence to exclude the non-specific antiviral effect, the toxicity of plant extracts on TZM-bl and CEM-GFP cells was assessed using MTT assay. The CC50 values of 50% ethanolic as well as aqueous extracts prepared from leaves of R. parviflora were > 331 μg/ml on TZM-bl cells and >196 μg/ml on CEM-GFP cells (Table 1). Both aqueous (IC50; 15 μg/ml) and 50% ethanolic extract (IC50; 26 μg/ml) showed dose dependent inhibition in HIV-1 infection using TZM-bl cell-based assay (Table 1). Treatment of HIV-1 infected CEM-GFP cells with the 50% ethanolic as well as aqueous extracts of the leaves of R. parviflora also showed a dose dependent inhibition of HIV infection with IC50 values of 29 and 65 μg/ml, respectively (Table 1). Both the extracts reduced the virus released by infected CEM-GFP cells as estimated by p24 in the culture supernatants collected from the infected cells (Figure 1). The higher efficacy of the extracts in TZM-bl cells as compared to CEM-GFP cells-based assay may be due to an additional step of virus pre-treatment with extracts in TZM-bl cells-based assay format and hence suggests the presence of additional virucidal phytochemicals. The therapeutic index (TI) of a drug is the ratio between the toxic and the therapeutic dose and is used as a measure of its relative safety. The TI values of the extracts were in a range of 6.8 to 25.5 (Table 1).


Anti-HIV-1 activity, protease inhibition and safety profile of extracts prepared from Rhus parviflora.

Modi M - BMC Complement Altern Med (2013)

Reduction in virus load by leaves extracts of R. parviflora using CEM-GFP cells. Cells were infected with HIV-1NL4.3 (0.05 MOI) and treated with extracts at various concentrations followed by 8 day incubation. Culture supernatant was collected on 8th day for p24 estimation by ELISA. AZT (10 μM) was used as positive control that reduced p24 level by 97%. Data is represented as mean ± SE of three independent experiments performed in duplicates. Statistical significance between the extract treated groups as compared to untreated control is shown by asterisk (*p < 0.001).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3716979&req=5

Figure 1: Reduction in virus load by leaves extracts of R. parviflora using CEM-GFP cells. Cells were infected with HIV-1NL4.3 (0.05 MOI) and treated with extracts at various concentrations followed by 8 day incubation. Culture supernatant was collected on 8th day for p24 estimation by ELISA. AZT (10 μM) was used as positive control that reduced p24 level by 97%. Data is represented as mean ± SE of three independent experiments performed in duplicates. Statistical significance between the extract treated groups as compared to untreated control is shown by asterisk (*p < 0.001).
Mentions: The inhibitory activity of the extracts against HIV infection is sometimes, a result of their toxic effects and consequently might result in an erroneous conclusion. Hence to exclude the non-specific antiviral effect, the toxicity of plant extracts on TZM-bl and CEM-GFP cells was assessed using MTT assay. The CC50 values of 50% ethanolic as well as aqueous extracts prepared from leaves of R. parviflora were > 331 μg/ml on TZM-bl cells and >196 μg/ml on CEM-GFP cells (Table 1). Both aqueous (IC50; 15 μg/ml) and 50% ethanolic extract (IC50; 26 μg/ml) showed dose dependent inhibition in HIV-1 infection using TZM-bl cell-based assay (Table 1). Treatment of HIV-1 infected CEM-GFP cells with the 50% ethanolic as well as aqueous extracts of the leaves of R. parviflora also showed a dose dependent inhibition of HIV infection with IC50 values of 29 and 65 μg/ml, respectively (Table 1). Both the extracts reduced the virus released by infected CEM-GFP cells as estimated by p24 in the culture supernatants collected from the infected cells (Figure 1). The higher efficacy of the extracts in TZM-bl cells as compared to CEM-GFP cells-based assay may be due to an additional step of virus pre-treatment with extracts in TZM-bl cells-based assay format and hence suggests the presence of additional virucidal phytochemicals. The therapeutic index (TI) of a drug is the ratio between the toxic and the therapeutic dose and is used as a measure of its relative safety. The TI values of the extracts were in a range of 6.8 to 25.5 (Table 1).

Bottom Line: Both the extracts did not disturb the integrity of monolayer formed by intestinal epithelial Caco-2 cells.The extracts when tested up to 100 μg/ml did not significantly reduce the viability of L. plantarum, L. fermentum, L. rhamnosus and L. casei.The studies reported herein showed in vitro anti-HIV activity and preliminary safety profile of the extracts prepared from the leaves of R. parviflora.

View Article: PubMed Central - HTML - PubMed

ABSTRACT

Background: In the present study, extracts prepared from the leaves of Rhus parviflora Roxb. (Anacardiaceae) were evaluated for their anti-HIV activity, which have been traditionally used for the treatment of neurological disorders such as anxiety, insomnia and epilepsy.

Methods: Aqueous and 50% ethanolic extracts prepared from leaves of the plant were tested for their cytotoxicity and anti-HIV property using reporter gene based assays as well as human peripheral blood lymphocytes (PBLs). Further these extracts were evaluated for their ability to inhibit HIV-1 reverse transcriptase (RT) and protease activity. Safety profile of the extracts was determined on viability of Lactobacillus sp., secretion of pro-inflammatory cytokines by vaginal keratinocytes and transepithelial resistance.

Results: Both aqueous (IC50 = 15 μg/ml) and 50% ethanolic (IC50 = 26 μg/ml) extracts prepared from leaves of R. parviflora showed anti-HIV activity in TZM-bl cells wherein the virus was treated with the extracts prior to infection. Further, both the extracts also inhibited virus load in HIV infected CEM-GFP cells and human PBLs. The anti-HIV activity is mediated through inhibition of HIV-1 protease activity. Both the extracts did not disturb the integrity of monolayer formed by intestinal epithelial Caco-2 cells. The extracts when tested up to 100 μg/ml did not significantly reduce the viability of L. plantarum, L. fermentum, L. rhamnosus and L. casei. The extracts (100 μg/ml) did not reveal any cytotoxic effect on vaginal keratinocytes (Vk2/E6E7). Levels of pro-inflammatory cytokines secreted by Vk2/E6E7 cells treated with both the plant extracts were within the non-inflammatory range.

Conclusions: The studies reported herein showed in vitro anti-HIV activity and preliminary safety profile of the extracts prepared from the leaves of R. parviflora.

Show MeSH
Related in: MedlinePlus