Research studies conducted in the laboratory have investigated the properties of silymarin or its isomer silybin using cell lines and animal models. Other substances in milk thistle have not been extensively studied.

Several research studies have investigated the effects of silymarin or silybin in a noncancer context. These studies have tested silymarin or silybin:

• In healthy animal liver and kidney cells.
• As a prophylaxis against toxic chemicals.
• In stimulating detoxification pathways (enzyme concentrations and activity).
• For antioxidant properties.

Silymarin or silybin has also been investigated in cancer models. The effects of silymarin and/or silybin have been investigated in prostate (DU 145, LNCaP, PC-3),[1-5] breast (MDA-MB 468, MCF-7),[6-8] hepatic (HepG2),[9,10] epidermoid (A431),[10] colon (Caco-2),[11] ovarian (OVCA 433, A2780),[12] histiocytic lymphoma (U-937),[13] and leukemia (HL-60) [14,15] cells. In animal tumor models, tongue cancer,[16] skin cancer,[17-22] bladder cancer,[23] and adenocarcinoma of the colon [24,25] and small intestine [25] have been investigated. These studies have tested the ability of silymarin or silibinin to:

• Mitigate the toxicity associated with chemotherapy agents.
• Enhance the efficacy of chemotherapy agents.
• Inhibit the growth of cancer cell lines and inhibit tumor initiation or tumor promotion.

Although many of these studies have produced encouraging results, none of the findings have been replicated in human clinical trials.

Laboratory data suggest that silymarin and silybin protect the liver from damage induced by toxic chemicals. Animal studies have found that liver cells treated with silybin and then exposed to toxins do not incur cell damage or death at the same rate as liver cells that are not treated with silybin. This finding suggests that silybin can prevent toxins from entering the cell or effectively exports toxins out of the cell before damage ensues.[10,26-30] Alternatively, this may be related to the effect of silymarin on detoxification systems. In vitro data have shown silybin to stimulate and/or inhibit phase I detoxification pathways in silybin-treated human liver cells. However, this effect was found to be dose-dependent, and these levels are not physiologically attainable with the current manufacturer dose recommendations.[31,32]

Silymarin has been shown to stimulate phase II detoxification pathways in mice. Administration of silymarin (100 or 200 mg /kg body weight/day) to SENCAR mice for 3 days significantly increased glutathione S-transferase activity in the liver (P < .01–.001), lung (P < .05–.01), stomach (P < .05), small bowel (P < .01), and skin (P < .01). This effect appeared to be dose-dependent.[33] Administration of silymarin to rats challenged with a toxin (50 mg/kg body weight) resulted in higher levels of glutathione in liver cells, decreased levels of oxidative stress (measured by malondialdehyde concentrations), and less elevated liver function tests (measured by levels of aspartate aminotransferase [AST] and alanine aminotransferase [ALT]).[30] Silymarin and silybin have also been found to accelerate cell regeneration in the liver through stimulation of precursors to DNA synthesis and enhancement of production of the cellular enzymes required for synthesis of DNA.[34-39] Laboratory studies have also shown silymarin and silybin to be potent antioxidants.[27,28,40-47] Silymarin has been shown to mitigate oxidative stress in cells treated with pro-oxidant compounds.

A number of laboratory studies have investigated the effect of silymarin or silybin on the efficacy and toxicity of chemotherapy agents or have measured their direct cytotoxic activity. In an investigation of the effect of a variety of flavonoids on the formation of DNA damage, silymarin did not induce DNA damage in colon (Caco-2) cells, hepatoma (HepG2) cells, and human lymphocytes.[11] At higher concentrations of silymarin (400–1,000 μmol/L) DNA damage was induced in an epithelial cell line (HeLa cells). At higher concentrations (1,000 μmol/L) DNA damage was observed in human lymphocytes. Cell growth was inhibited as the flavonoid concentration was increased in human lymphocytes and HeLa cells. Only at very high concentrations was cell viability affected by silymarin in HepG2 cells. Although this study demonstrated that the flavonolignans of Silybum marianum (L.) are capable of inhibiting cellular proliferation and inducing DNA strand breaks, the results were obtained at very high concentrations that may be difficult to achieve in humans. This study also showed that silymarin does not stimulate cell growth in the HeLa, Burkitt lymphoma, and human hepatoma cell lines.

Silymarin has also been investigated as a possible adjunctive agent in mitigating some of the toxicity associated with chemotherapy agents. Silybin and silychristin exerted a protective effect on monkey kidney cells exposed to vincristine and especially cisplatin chemotherapy.[48] Silybin (200 mg/kg body weight) administration with cisplatin in rats resulted in statistically significant reductions in measures of kidney toxicity.[49] Significant decreases in weight loss, faster recovery of urinary osmolality measures, and depressions in the increase in activity of urinary alanine aminopeptidase ([AAP], a marker of kidney toxicity) were observed. Silybin had no effect on magnesium excretion or glomerular function. Silybin (2 g /kg body weight) administration in rats receiving cisplatin prevented reductions in creatinine clearance, increases in urea plasma levels, and large increases in urinary AAP.[50] No effect on magnesium excretion was observed. Silybin did not interfere with the antineoplastic effects of cisplatin or ifosfamide in germ cell tumors. In experiments with ovarian and breast cancer cell lines, silybin potentiated the effect of cisplatin and doxorubicin.[12] IdB 1,016, a form of silybin bound to a phospholipid complex, was found to enhance the activity of cisplatin against A2780 ovarian cancer cells but had no effect on its own.[51] Silybin increased the chemosensitivity of DU 145 prostate cancer cells resistant to chemotherapy.[52]

Studies have also investigated the effect of silymarin on tumor initiation and promotion. Silymarin appears to have a chemopreventive effect through perturbations in the cell cycle, altering cell signaling that induces cellular proliferation, affecting angiogenesis, or through its anti-inflammatory properties.[1,6,12,18,53] These findings have been supported in human prostate, breast, ectocervical, ovarian, hepatic, leukemia, and epidermoid cell lines.[4,6,8,9,14,54] An investigation of the effect of silymarin on ultraviolet B radiation-induced nonmelanoma skin cancer in mice found that silymarin treatment significantly reduced tumor incidence (P < .003), tumor multiplicity (P < .0001), and tumor volume (P < .0001).[18] These findings suggest that silymarin plays a prominent role in the reduction of cancer cells and in preventing the formation of cancer cells. A number of studies have investigated the mechanism through, which silymarin may affect tumor promotion in mouse skin tumor models. Studies have found that silymarin reduces transcription of markers of tumor promotion and activity,[18] inhibits transcription of tumor promoters,[55] stimulates antioxidant activities,[18,22] interferes with cell signaling,[54] inhibits inflammatory actions,[18,21] and modulates cell-cycle regulation.[56]

In prostate cancer cell lines, silybin has been shown to inhibit growth factors and stimulate cell growth,[1-3,5] promote cell cycle arrest,[1,4] and inhibit antiapoptotic activity.[52] In rats with azoxymethane -induced colon cancer, dietary silymarin resulted in a reduction in the incidence and multiplicity of adenocarcinoma of the colon in a dose-dependent manner.[24,25] Dietary silymarin had no effect on small intestinal adenocarcinoma,[25] but exerted a preventive effect in mice with N-butyl-N-(4-hydroxybutyl) nitrosamine –induced bladder cancer [23] and in F344 rats with 4-nitroquinoline 1-oxide –induced cancer of the tongue.[16] Dietary silybin administered to nude mice with prostate carcinoma increased production of insulin-like growth factor-binding protein-3 in the plasma of mice and significantly inhibited tumor volume (P < .05).[2]

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