Immunomodulatory activities of various medicinal plant extracts: effects on human lymphocytes apoptosis.

Plant extracts have been widely evaluated for biological properties. In the present study extracts of several native plants in Iran was investigated for their possible immunomodulatory effects. Peripheral blood lymphocytes separated from healthy individuals were stimulated with phytohemagglutinin (PHA) and cultured with different concentrations of the extracts. Comparison of the cell proliferation in treated cultures showed the highest inhibitory effect due to exposure with Linum persicum. This extract caused a strong dose-dependent decrease in lymphocyte proliferation (p < 0.001). Lymphocytes treated with Cirsium bracteosum were inhibited in a dose dependent manner (SI range 0.9-0.2). Similarly, Echinophora cinerea-treated lymphocytes showed a significant reduction in proliferation compared to that in non-treated cells. Among the extracts, Dionysia termeana, Salvia macrociphon and Ferulago angulata had a mild stimulatory effect on the lymphocytes at concentrations less than 1 microg/ml (p < 0.05). At higher doses all these extracts showed significant inhibitory effects on the proliferation of PHA-treated cells (SI range 0.81 to 0.04). In cell cycle analysis performed by flow cytometry, the strongest appearance of apoptotic cells at sub-G1 phase in various extract-treated cultures was found for D. termeana (14.6 +/- 0.5%). The Percentage of cells undergoing apoptosis in cultures treated with L. persicum was more than 11 compared to that of the control (1.7 +/- 0.08). In DNA analysis, D. termeana and L. persicum showed typical DNA laddering, indicating that these extracts induced apoptosis of lymphocytes. In conclusion, all the extracts studied showed lymphocyte inhibitory effects at high concentrations. These inhibitory effects for some of the plants seem to be due to induction of apoptosis.

Calycopterin, an immunoinhibitory compound from the extract of Dracocephalum kotschyi.

Medicinal plants have been widely investigated for their various effects. Dracocephalum kotschyi Boiss (Labiatae) is used in Iranian traditional medicine for the treatment of rheumatoid diseases. The inhibitory effect of D. kotschyi on the lectin-induced cellular immune response has been demonstrated previously. In this study, mitogen-treated lymphocytes were exposed to the extract of D. kotschyi and analysed for the induction of apoptosis using flow cytometry and gel electrophoresis. The data obtained indicated a dose-dependent increase of cells in the sub-G1 phase of cell cycle. Study of internucleosomal DNA fragmentation showed a typical DNA laddering in agarose gels. A bioactivity-guided fractionation assay to find the active components responsible for the inhibitory effect of D. kotschyi on mitogen-induced lymphocyte proliferation led to the isolation of calycopterin from the ethyl acetate extract of D. kotschyi. Its structure was identified by spectroscopic methods including( 1)H-NMR, (13)C-NMR, MS and UV spectra. Calycopterin inhibited lymphocyte proliferation in a dose-dependent manner with an IC(50) value of 1.7 microg/mL. In conclusion, the results of this study suggest that D. kotschyi extract has the capacity to induce apoptosis in the lymphocytes and that isolated calycopterin is responsible for the inhibitory effect of D. kotschyi on lymphocyte proliferation.

Antitumor activity and apoptosis induction in human cancer cell lines by Dionysia termeana.

Studies have demonstrated that plant extracts possess various biological effects including antitumor activity. In the present study, the antitumor activity of Dionysia termeana, a plant native to Iran, was investigated. Cytotoxic activity of the extract on tumor cell lines using MTT colorimetric assay was determined. Cell cycle analysis by flow cytometry and DNA fragmentation analysis on sensitive cell lines was then carried out. Results obtained indicated that the highest activity of D. termeana was against K562 leukemia cell line with IC50 less than 20 microg/mL. Fifty-five percent inhibition of Jurkat cells due to exposure to D. termeana was found at 200 microg/mL of the extract. A549, a lung carcinoma cell, and Fen bladder carcinoma cell line were less affected. In flow cytometry analysis, D. termeana induced apoptosis in the K562 and Jurkat cells. In DNA fragmentation analysis the extract produced ladder formation in both cells. In conclusion, these results indicated that the extract used in this study have antitumor activity through induction of apoptosis particularly in the leukemia cell lines.

Induction of apoptosis in leukemia cell lines by Linum persicum and Euphorbia cheiradenia.

PURPOSE: In the present study two medicinal herbs Linum persicum and Euphorbia cheiradenia that are native to Iran were tested for their possible anticancer effect and induction of apoptosis on human tumor cell lines including leukemia cell lines. METHODS: The effect of methanolic extracts of the herbs on the inhibition of cell proliferation was assessed by MTT colorimetric assay. K562 and Jurkat cell lines treated with the extracts were analyzed for the induction of apoptosis by flow cytometry using propidium iodide staining. DNA fragmentation analysis was performed. RESULTS: Various concentrations of L. persicum and E. cheiradenia showed inhibitory effects on different cell lines. Two leukemic lines including K562 and Jurkat were the most sensitive cells for L. persicum with IC50 of 0.1 and 10 mug/ml, respectively. In the cultures of tumor cell lines treated with E. cheiradenia, the main inhibitory effects was for Jurkat cells with IC50 of 12.5 microg/ml. Results indicated a dose-dependent accumulation of cells in the sub-G1 phase. Study of internucleosomal DNA fragmentation showed a typical DNA laddering in agarose gels for both extracts. CONCLUSION: The present study showed cytotoxic activity of both herbs on tumor cell lines and suggests that this effect may in part be due to the induction of apoptosis in leukemic cells.

Effect of Salvia mirzayanii on the immune system and induction of apoptosis in peripheral blood lymphocytes.

In this study, the immunomodulatory effects of Salvia mirzayanii were investigated. Study of the effect of this plant on activated human peripheral blood lymphocytes showed stimulatory effects at lower concentrations and inhibitory effects at higher ones (p < 0.01). In flow cytometry analysis, accumulation of apoptotic cells in the sub-G1 phase of the cell cycle of the mitogen-treated lymphocytes exposed to the inhibitory doses of the extract was observed. DNA fragmentation analysis of these cells showed a typical DNA laddering. Immunisation of extract-treated mice with the antigen decreased delayed hypersensitivity skin reaction as well as the antibody titre at higher concentrations (p < 0.007). These results indicated the presence of immunomodulatory compounds in the extract of S. mirzayanii and suggest that the induction of apoptosis in lymphocytes might be the mechanism responsible for the inhibitory effect of the extract that was observed at higher concentrations.

Stimulatory effects of Euphorbia cheiradenia on cell mediated immunity and humoral antibody synthesis.

BACKGROUND: Studies have demonstrated that plant extracts possess various biological characteristics including immunomodulatory activity. OBJECTIVE: Euphorbia cheiradenia Boiss et Hohen (Euphorbiaceae), a medicinal herb native to Iran was investigated for its immunomodulatory effects. METHODS: The methanolic extract of the plant was prepared and added to mitogen-induced human peripheral blood lymphocyte cultures at different concentrations. Effect of E. cheiradenia on in vivo cell-mediated immunity was measured by delayed type hypersensitivity (DTH) reaction. The effect of the extract on humoral antibody synthesis was also measured in immunized mice treated with different extract concentrations. RESULTS: The stimulation index (SI) for cultures treated with 0.01 to 200 microg/ml of the extract ranged from 1.3+/-0.04 to 2.4+/-0.06, (p<0.01) showing a significant stimulatory effect of E. cheiradenia on the lymphocytes. IL-2 secreted from lymphocytes treated with the extract was significantly higher than that from the non-treated cells (p<0.001). Cell cycle analysis on mitogen-treated lymphocytes exposed to different concentrations of the extract showed an increase in the percentage of cells at G2M phase with increases in the concentration of the extract, but the results was not significant. In DTH skin test, the mean footpad thickness of all mice groups treated with 1, 50 and 100 mg/kg of the extract at 24 hours after immunization with antigen was 3.5+/-0.6 mm compared to 2.5+/-0.5 mm for the non-treated group (p=0.005). Moreover, an increase in production of specific antibody in mice immunized with different extract concentrations was also demonstrated. CONCLUSION: Results of this study showed the ability of the E. cheiradenia extract to induce proliferation of lymphocytes and enhance both cellular and humoral specific immune responses.

Immunomodulatory and apoptotic effects of Stachys obtusicrena on proliferative lymphocytes.

BACKGROUND: Several medicinal plants have shown anti-proliferative and apoptotic effects on human lymphocytes. In the present study the immunomodulatory properties of Stachys obtusicrena, a native plant to Iran, was examined and its possible effects on the induction of apoptosis were determined. MATERIAL/METHODS: The in vivo effect of the methanolic extract of S. obtusicrena on delayed type hypersensitivity (DTH) and antibody production was investigated in mice immunized with the antigen. The proliferation of human lymphocytes stimulated with mitogen in the presence of the extract was determined by [3H]-thymidine incorporation assay. Mitogen-treated lymphocytes were exposed to the extract and analyzed for induction of apoptosis using flow cytometry and gel electrophoresis. RESULTS: Data obtained from the DTH and antibody responses indicated a dose-related decrease in both parameters in the extract-treated mice compared with the untreated control group. In the in vitro study performed on the lymphocytes, the extract caused a dose-dependent decline in [3H]-thymidine uptake. The stimulation index of all cultures treated with different concentrations of the extract was less than 1. IL-2 levels in the culture supernatants of the activated lymphocytes showed a dose-related decrease as well. Results of flow cytometry indicated a dose-dependent accumulation of cells in the sub-G1 phase. The internucleosomal DNA fragmentation study showed typical DNA laddering in agarose gels. CONCLUSIONS: The present study showed inhibitory effects of S. obtusicrena on both cellular and humoral immune responses and suggests that this effect may in part be due to the induction of apoptosis in proliferative lymphocytes.