Расторопши: семена ранней потенциальных

Discussion

Oxidative stress refers to an imbalance between pro-oxidant and anti-oxidant factors, which are controlled by multiple components. Oxidative stress may lead to cellular damage. ROS play a key role in oxidative stress and are generated as by-products of cellular metabolism, primarily in the mitochondria [20]. Elevated levels of mitochondrial ROS have been shown to be sufficient to trigger apoptosis [21]. In the present study, the apoptotic effects of SIL in HepG2 cells were associated with rapid increases in the levels of intracellular ROS. Additionally, SIL-induced ROS generation was associated with significant depletions of intracellular GSH, which is a major non-protein cellular antioxidant that can eliminate intracellular ROS [2], [22]. The degree of exposure to ROS and perturbations in the GSH redox balance play critical roles in determining whether cells undergo pro-survival or pro-death responses [3]. Tumor cells are significantly more sensitive to changes in the levels of GSH because tumor cells have heightened basal levels of ROS-mediated signals, which contribute to their increased rates of growth, metabolism and proliferation. Therefore, tumor cells may be more vulnerable to oxidative stress [20]. In the present study, T-AOC was remarkably decreased in SIL-treated HepG2 cells. SIL may have exhausted the total cellular antioxidant capacity and increased the ROS levels beyond a threshold, which may have contributed to the induction of apoptosis in HepG2 cells.

Studies have shown that components of the Notch pathway are overexpressed during HCC progression, as observed for other genes that are known to be involved in the survival of cancer cells [16], [23], [24]. Ahn and colleagues found that Notch1 and Notch4 are markers for poor prognoses during hepatocellular carcinoma [16]. Dill and colleagues confirmed that constitutive Notch2 signaling induces hepatic tumors in mice [23]. Downregulating Notch1 may be an effective approach to inactivating Snail/E-cadherin by regulating cyclooxygenase-2, which results in the inhibition of HCC cell invasion and migration [24]. The overexpression of the key transcription factor RBP-Jκ and its downstream target Hes1 has also been previously reported in HCC [15], [25]. Our results indicate that SIL treatment decreased NICD, RBP-Jκ, and Hes1 in HepG2 cells. When combined with Notch1 siRNA in vitro or DAPT in vivo, silybin further decreased the viability of HCC cells and/or inhibited tumor growth. Additionally, recombinant Jagged1 protein (a known Notch ligand in vitro) effectively attenuated the antitumor activity of SIL. These data suggest that the antitumor activity of SIL in HCC cells is exerted in part through the inhibition of the Notch signaling pathway.

The ability to induce cellular apoptosis is an important property of many candidate anticancer drugs [21]. Apoptosis is a tightly regulated process that involves at least one of the caspase-dependent signaling pathways, i.e., the cell death receptor pathway or the mitochondrial pathway. In the mitochondrial pathway, a variety of death signals trigger the release and translocation of several pro-apoptotic proteins from the mitochondria to the cytosol. Among the numerous factors known to modulate apoptosis in cancer cells, the proteins of the Bcl2 family are considered to be the main regulators of apoptosis. Bcl2 is an anti-apoptotic protein, whereas Bax is a crucial pro-apoptotic and tumor suppressor protein. The ratio of anti-apoptotic to pro-apoptotic molecules, such as the Bcl2/Bax ratio, indicates the threshold sensitivity of cells to the induction of apoptosis via the intrinsic pathway [20]. The activation of caspases is a pivotal step in the apoptotic process and is triggered by signals from death factors, mitochondrial alterations or DNA damage due to external and/or internal insults [26]. Regardless of whether apoptosis is mediated via the cell death receptor pathway or the mitochondrial pathway, both of the pathways ultimately activate caspase3, which in turn induces DNA fragmentation, the characteristic morphological change that is associated with apoptotic cells. Caspase3 activation indicates a key and irreversible point in the induction of apoptosis [27]. Our results indicated that SIL treatment not only downregulated Bcl2 protein expression but also upregulated Bax protein expression and caspase3 activity in HepG2 cells. These results indicated that SIL-induced apoptosis occurred via the mitochondrial pathway and verified the results of previous studies that indicated that the inhibition of Notch signaling is associated with the induction of the mitochondrial apoptotic pathway in human cancer cells [28], [29].

The results of the present study also indicate that the viability of HepG2 cells was decreased by the use of Notch1 siRNA (in vitro) or DAPT (in vivo) in combination with SIL treatment. These decreases were associated with decreases in NICD, RBP-Jκ, Hes1, and Bcl2 expression, as well as with increases in Bax expression. These results indicate that the inhibition of Notch signaling may be a novel means both of enhancing the effects of chemotherapy and of delaying chemoresistance in patients with cancer. In addition, Notch1 siRNA or DAPT in combination with SIL treatment further suppressed the expression of cyclin D1 and survivin, two pro-survival factors that are targets of Notch1 and have been reported to play roles in sensitization to anticancer drugs [15], [30], [31]. These results suggest that the inhibition of Notch signaling may sensitize HCC cells to SIL treatment by preventing the activation of pro-survival pathways. The concentration of DAPT (10 mg/kg) that did not affect tumor volume significantly (compared with the control group) was selected based on our preliminary experiments. As expected, DAPT treatment alone significantly decreased NICD expression (compared with the control group); however, DAPT alone only slight affected Cyclin D1, Bcl2, and Bax gene expression, while SIL had very potent effects on tumor suppression and on the expression of these proteins. These results indicate that Notch signaling is not the only pathway that mediates the function of SIL.

In conclusion, these experiments provide mechanistic evidence that the Notch pathway is inhibited in HCC cells in response to SIL treatment. The downregulation of Notch signaling sensitizes HCC cells to SIL treatment, and upregulation of Notch signaling desensitizes HCC cells to SIL treatment. Additionally, this potentiation of chemosensitivity by Notch inhibition may be related to the downregulation of pro-survival pathways. Therefore, we propose that the inhibition of Notch signaling may be a novel strategy that can be used to prevent the induction of cancer survival mechanisms in advanced HCC.

Силибинин является основным биологически активных flavonolignan в настоящее расторопши (Silybum marianum), который обладает антиоксидантным, противовоспалительным и антиканцерогенным деятельности. Однако точное базовый механизм остается до конца не изучены. Настоящее исследование было разработано для исследования молекулярных механизмов, лежащих в для antitumorigenic потенциал силибинин против химически индуцированных опухолей кожи в швейцарских мышей-альбиносов. В свете важной роли ядерного фактора-kappaB (NF-b), циклооксигеназы-2 (ЦОГ-2), iNOS, провоспалительных цитокинов, фактора роста сосудистого эндотелия и окислительного стресса в канцерогенез, химические эффективность силибинин против 7, 12-dimethylbenz[a]антрацен/12-O-tetradecanoylphorbol-13-ацетат-индуцированных 2-этап канцерогенеза кожи изучали с точки зрения цитопротекторной активности ферментов, липидов, воспалительных реакций, и выражение различных молекулярных маркеров в ткани кожи. Мы обнаружили, что местное применение силибинин в дозе 9 мг/мышь эффективно подавляют окислительный стресс и нерегулируемого активации медиаторов воспаления и опухолей. Таким образом, результаты настоящего исследования показывают, что химические эффект силибинин связана с повышенной эндогенных цитопротекторных механизмов и down-регуляции воспалительных медиаторов (оксид азота, фактор некроза опухоли-α, интерлейкин-6, интерлейкин-1β, COX-2, iNOS, и NF-b).