Molecular and cytogenetic effects of Thai royal jelly: modulation through c-MYC, h-TERT, NRF2, HO-1, BCL2, BAX and cyclins in human lymphocytes in vitro.

Royal jelly (RJ) is widely used as a food supplement for anti-aging and beauty. However, its use has been linked to asthma and hemorrhagic colitis. Since its mechanisms of toxicity have not been fully identified, we conducted an investigation to elucidate its molecular and cytogenetic effects. Using human lymphocytes in vitro, treatments with RJ (0.0005-5 mg/ml) for 3 h did not induce sister chromatid exchanges until 5 mg/ml was used. Treatments for 24 h showed a dose-dependent reduction in BCL2/BAX, c-MYC/BAX and HO-1/BAX ratios. The exception was the NRF2/BAX ratio, showing a dose-dependent reduction at low doses, but a marked increase at the highest dose. The hTERT/BAX ratio was maintained at approximately a 1.2-fold increase but decreased to nearly normal at the highest dose. Our findings indicated that the lowest dose of RJ treatment provided maximum benefits, mainly through hTERT activation relating to prolonged lifespan. The highest dose of RJ inhibited cell survival, cell proliferation and an antioxidative enzyme; nevertheless, it still activated an antioxidative response through NRF2 and maintained telomeres during cell crisis. RJ treatment at 0.05 mg/ml increased cyclin E, BCL2 and BAX to maximum levels indicating that throughout the active cell cycle, both cell survival and cell apoptosis increased. Using the gene expression ratios over BAX, similar to BCL2/BAX, provided more informative data than using individual protein levels alone. With these informative ratios, our results confirm the potential benefits of RJ in enhancing lifespan and activation antioxidative power. Further, in vivo mechanistic studies will be useful in validating these results.

In vitro enhancement of doxorubicin genotoxic activities and interference with cell cycle delay by Plumbago indica root ethanolic extract in human lymphocytes.

BACKGROUND: Combinations of modern medicines with herbal medicines are being developedfor more effectiveness. Data on the safety and drug-herb interactions are needed to be clarified. Ethanolic extract of Plumbago indica root (EEPIR) is medicinally usedfor cancer treatment in Asian traditional medicine. However its mechanism of action is still inconclusive. Our previous study demonstrated that EEPIR was genotoxic and induced cell cycle delay in human lymphocytes in vitro. OBJECTIVE: To investigate genotoxic potency and interference with cell cycle of EEPIR in combination with doxorubicin (DXR), a standard chemotherapeutic agent, in human lymphocytes by in vitro sister chromatid exchange (SCE) assay. MATERIAL AND METHOD: Human lymphocytes were pretreated with EEPIR at 6.25-100 mcg/mlfollowed by DXR (0.1 mcg/ml). SCE levels and cell cycle kinetics were evaluated. RESULTS: EEPIR pretreatments (6.5-50 mcg/ml) significantly enhanced genetic damage induced by DXR (p<0. 05). Delaying of the cell cycle was detected and related to EEPIR concentration. EEPIR at 100 mcg/ml, on the contrary, did not enhance DXR-induced genotoxicity but tended to lower genotoxicity compared to DXR treatment alone. It significantly delayed cell cycle the most (p<0.05). CONCLUSION: EEPIR pretreatments at proper doses enhanced genotoxic damage induced by DXR in human lymphocytes. Delaying cell cycle by EEPIR could lower that potency. Usage of EEPIR, therefore, should be adjusted for safety. Combination of EEPIR with DXR might be usefulfor more efficient cancer treatment with less DXR toxicity. Further in vivo study is needed to support this in vitro evidence.

Synergistic Genotoxic Effects and Modulation of Cell Cycle by Ginger Ethanolic Extracts in Adjunct to Doxorubicin in Human Lymphocytes In Vitro.

BACKGROUND: Several natural phytochemicals are increasingly used, as an adjunct to chemotherapy, to reduce drug adverse effects. Zingiber officinale rhizome (ginger) product has been reported to be effective against nausea and vomiting in patients receiving emetogenic chemotherapy such as cisplatin/doxorubicin (DXR). In addition, its ethanolic extract of Zingiber officinale rhizome (EEZOR) has been reported to possess anticarcinogenic properties. However, the mechanism for anticancer activity of ginger especially when used in combination with chemotherapy has not well elucidated, one of its possible mechanisms might involve its genotoxicity. OBJECTIVE: To investigate genotoxic and cytotoxic potentials of EEZOR alone and EEZOR pretreatments followed by 0.1 mcg/ ml DXR, a genotoxic chemotherapeutic agent in human lymphocytes by sister chromatid exchange (SCE) assay in vitro. The effect on cell cycle kinetics was also explored. MATERIAL AND METHOD: Human lymphocytes were treated with EEZOR alone at 25-500 mcg/ml and EEZOR pretreated at 12.5-200 mcg/ml followed by 0.1 mcg/ml DXR. SCE levels and cell cycle kinetics were evaluated. RESULTS: EEZOR significantly induced biphasic SCE at 50 and 400 mcg/ml (p < 0.05). However, cytotoxicity manifested at 500 mcg/ml. All EEZOR pretreatments at 12.5, 25, 50, and 100 mcg/ml, except at 200 mcg/ml, prior to DXR, moderately enhanced DXR-induced genotoxicity by 1.3 times (p < 0.05). Both EEZOR alone and EEZOR prior to DXR at certain concentrations delayed cell cycle. CONCLUSION: At specific doses, EEZOR could induce genotoxicity and in pretreatments could moderately enhance DXR-induced genotoxicity and delay cell cycle. This finding suggests that dosage use of EEZOR needs to be adjusted for long-term safety. In addition, EEZOR in adjunct to DXM might have potential benefits not only as an emetic agent but also in chemotherapy. Further in vivo animal and human studies to support this evidence is essentially needed.

Antigenotoxic activity of Thai Sangyod red rice extracts against a chemotherapeutic agent, doxorubicin, in human lymphocytes by sister chromatid exchange (SCE) assay in vitro.

BACKGROUND: Nowadays, anticarcinogenic potential of pigmented brown rice and rice bran varieties have been increasingly stated. However, their mechanisms of action are still inconclusive. One of which might be their antigenotoxic activity that no study in human cells was reported before. OBJECTIVE: To evaluate the antigenotoxic activities of Thai Sangyod red rice extracts against a chemotherapeutic agent, doxorubicin, by sister chromatid exchange (SCE) assay in human lymphocytes in vitro. MATERIAL AND METHOD: Two fractions of water-soluble of Sangyod rice extracts were used: (i) the washed water extract of brown rice (WWBR) and (ii) the water extract of rice bran (WERB). Human lymphocytes were pretreated with each extracts at concentrations of 6.2, 12.5, 25, 50 and 100 microg/ml for 2 h followed by a genotoxic agent, doxorubicin (DXR) (0.1 microg/ml) for 2 h. SCE level, mitotic index (MI) and proliferation index (PI) were evaluated. Statistical analysis by Dunnett's t-test was performed. RESULTS: The results indicated that the pretreatment of WERB fraction only at concentration of 6.2 microg/ml could significantly decrease SCE level as compared to that of the DXR treated alone (p < 0.05). On the other hand, WERB fraction at other concentrations and all WWBR pretreatments could not. In addition, there was no significant difference in MI and PI levels between all pretreated extracts as compared to the DXR treated alone. CONCLUSION: Our data revealed that WERB pretreatment only at specific low concentration of 6.2 microg/ml possessed the antigenotoxic potential against genotoxic damage but not anticytotoxic induced by DXR. Further work is still needed to clarify more the antigenotoxic and anticytotoxic potentials from other fractions of Sangyod rice extracts.

Genotoxicity and interference with cell cycle activities by an ethanolic extract from Thai Plumbago indica roots in human lymphocytes in vitro.

In Thai traditional medicine, Plumbago indica or Jetamul-Pleung-Dang in Thai is known to have health benefit especially for anti-inflammatory, antibacterial, and antitumor activities. However, the mechanisms of its action are still uncertain. One of which might be genotoxic effects. In the present study, we investigated the genotoxicity of an ethanolic extract of Plumbago indica root (EEPIR) by sister chromatid exchange (SCE) assay in human lymphocytes. Results have shown that all treatments with EEPIR (12.5-100 µg/ml) could induce cell cycle delay as shown by significant increase in the number of metaphase cells in the first cell cycle but neither in the second nor the third cell cycle. Only at concentrations of 25, 50, and 100 µg/ml were SCE levels significantly increased above that of the control (p<0.05) . EEPIR at a concentration of 500 µg/ml induced cell death as few mitotic cells were shown. Accordingly, EEPIR (25-100 µg/ml) is genotoxic in human lymphocytes and cytotoxic at concentrations of ≥ 500 µg/ml in vitro. Therefore, these activities of the EEPIR could serve its potential therapeutic effects, especially as an anticancer agent. Further study of EEPIR in vivo is now needed to support this in vitro evidence.