Angiopreventive Efficacy of Pure Flavonolignans from Milk Thistle Extract against Prostate Cancer: Targeting VEGF-VEGFR Signaling

The role of neo-angiogenesis in prostate cancer (PCA) growth and metastasis is well established, but the development of effective and non-toxic pharmacological inhibitors of angiogenesis remains an unaccomplished goal. In this regard, targeting aberrant angiogenesis through non-toxic phytochemicals could be an attractive angiopreventive strategy against PCA. The rationale of the present study was to compare the anti-angiogenic potential of four pure diastereoisomeric flavonolignans, namely silybin A, silybin B, isosilybin A and isosilybin B, which we established previously as biologically active constituents in Milk Thistle extract. Results showed that oral feeding of these flavonolignans (50 and 100 mg/kg body weight) effectively inhibit the growth of advanced human PCA DU145 xenografts. Immunohistochemical analyses revealed that these flavonolignans inhibit tumor angiogenesis biomarkers (CD31 and nestin) and signaling molecules regulating angiogenesis (VEGF, VEGFR1, VEGFR2, phospho-Akt and HIF-1α) without adversely affecting the vessel-count in normal tissues (liver, lung, and kidney) of tumor bearing mice. These flavonolignans also inhibited the microvessel sprouting from mouse dorsal aortas ex vivo, and the VEGF-induced cell proliferation, capillary-like tube formation and invasiveness of human umbilical vein endothelial cells (HUVEC) in vitro. Further studies in HUVEC showed that these diastereoisomers target cell cycle, apoptosis and VEGF-induced signaling cascade. Three dimensional growth assay as well as co-culture invasion and in vitro angiogenesis studies (with HUVEC and DU145 cells) suggested the differential effectiveness of the diastereoisomers toward PCA and endothelial cells. Overall, these studies elucidated the comparative anti-angiogenic efficacy of pure flavonolignans from Milk Thistle and suggest their usefulness in PCA angioprevention

Azadirachta indica leaf extract modulates initiation phase of murine forestomach tumorigenesis

209-215The effects of aqueous Azadirachta indica leaf extract (AAILE) on benzo(a)pyrene [B(a)P]-induced forestomach tumorigenesis, B(a)P-DNA adduct formation and certain parameters of carcinogen biotransformation system in mice have been reported earlier from our laboratory. In this study, the effects of AAILE on the enzymes of B(a)P biotransformation, which play crucial role in initiation of chemical carcinogenesis aryl hydrocarbon hydroxylase (AHH) and uridinediphosphoglucuronosyltransferase (UDP-glucuronosyltransferase) have been evaluated in murine forestomach and liver. In addition, lipid peroxidation (LPO) levels in forestomach as well as liver and the activities of tissue injury marker enzymes lactate dehydrogenase, aspartate aminotransferase and alkaline phosphatase in the serum have also been evaluated. Oral administration of AAILE (100 mg/kg body wt for 2 weeks) reduces the AHH activity and enhances the UDP-glucuronosyltransferase activity in both the tissues, suggesting its potential in decreasing the activation and increasing the detoxification of carcinogens. The LPO levels decrease upon AAILE treatment in the hepatic tissue, suggesting its anti-oxidative and hence anti-carcinogenic effects. Non-significant alterations have been observed in tissue injury marker enzymes upon AAILE treatment, suggesting its safety at the given dose. In conclusion, AAILE appears to modulate initiation phase of carcinogenesis and may be suggested as safe and an effective agent for chemoprevention

Impediment of diethylnitrosamine induced hepatotoxicity in male Balb/c mice by pretreatment with aqueous <i style="">Azadirachta indica </i>leaf extract

359-366Considering the hepatoprotective properties of Azadirachta indica, the present study was designed to evaluate its preventive effects against diethylnitrosamine (NDEA) induced hepatotoxicity in male Balb/c mice. Exposure of NDEA caused a significant increase in micronucleated cell score, lipid peroxidation levels (LPO) and activity of lactate dehydrogenase (LDH). A significant decrease in reduced glutathione (GSH) contents and activity of glutathione-S-transferase (GST) was also observed upon NDEA treatment, whereas their activities of cytochrome P450 and cytochrome b5 showed non-significant alterations. Aqueous A. indica leaf extract (AAILE) pretreatment showed protective effects against NDEA induced toxicity by decreasing the frequency of micronucleated cell, levels of LPO and LDH activity. Also, a decreased activity of GST, cytochrome P450 and an increased activity of cytochrome b5, GSH contents was observed when AAILE pretreated mice were injected with NDEA. Only AAILE treatment caused a noticeable decrease in the frequency of micronuclei, activity of cytochrome P450 and cytochrome b5, but a significant increase in the activity of GST and GSH contents, whereas, non significant alterations were observed in the activity of LDH and levels of LPO. Significance of these observations with respect to hepatoprotective efficacy of A. indica has been discussed in the present manuscript

<b>Pitfalls in Journey from Traditional to Modern medicine</b>

6-13For thousands of years, man has sought healing powers from the natural world especially from plants. With the advent of modern medicine humans started isolating the active components and used them as such or made more effective analogs for therapy. Crude extract(s) based traditional medicine, on the other hand has better compatibility with the human body with minimal side effects. It is evident that crude extract has lesser side effects because of the modulatory activity of some components present in it along with the active component, which neutralizes the side effects and even synergies the medicinal effects of the later. In this article authors have analyzed the advantages of use of crude extract than their active component or synthetic counterpart based on information in literature

Modulatory effects of different doses of alpha-tocopherol on benzo(a)pyrene-DNA adduct formation in the pulmonary tissue of cigarette smoke inhaling mice

1139-1143Cigarette smoke (CS) has been established as one of the major risk factors for many pathologies including lung cancer in humans and experimental animals. In view of the discrepancy about the role of alpha-tocopherol (AT) in carcinogenesis, the present study was designed to investigate the effects of different doses of AT on benzo(a)pyrene-DNA [B(a)P-DNA] adduct formation in lungs of CS inhaling mice. Extent of carcinogen-DNA adduct formation has been considered as an index for carcinogenesis. Feeding of 35 IU AT/kg body weight increased B(a)P-DNA adducts formation significantly whereas feeding of 5 IU AT/kg body weight did not altered much the B(a)p-DNA adduct levels when both were compared to the control counterparts. With CS inhalation, the B(a)P-DNA adducts formation increased in all the groups when compared to their respective sham counterparts. Interestingly, in CS exposed groups, there was least increase in B(a)P-DNA adducts formation in 5 IU AT/kg fed animals followed by the control and 35 IU AT/kg body weight fed groups respectively. The results suggest that higher doses of AT accentuate DNA adduct formation in CS inhaling mice

Differential effect of flavonolignans on human PCA DU145 cells and endothelial HUVEC in cell culture assays.

<p>(<b>A</b>) Effect of flavonolignans (at 90 µM dose) on the three dimensional growth of DU145 cells was studied as detailed in ‘Materials and Methods’. Representative spheroids photomicrographs are shown at 100x and 400x. (<b>B</b>) In Transwell invasion assay, either DU145 cells (plated in the lower chamber) or HUVEC (plated in the upper chamber) were treated with individual diastereoisomers (at 30 µM dose), and HUVEC invasiveness was measured. (<b>C</b>) DU145 cells were treated with individual diastereoisomers (at 30 µM dose) and conditioned media was collected. HUVEC were plated on matrigel along with 0.5% FBS or conditioned media from different treatment groups; and tube formation was analyzed. Representative photomicrographs of tubular network are shown at 100x. Abbreviations: Sil A: Silybin A; Sil B: Silybin B; Iso A: Isosilybin A, Iso B: Isosilybin B; CM: Conditioned media; *, p ≤ 0.001; #, p ≤ 0.01.</p

Flavonolignans inhibit VEGF-induced proliferation and invasion in HUVEC.

<p>(<b>A</b>) HUVEC were induced with VEGF and treated with each flavonolignan in 0.5% serum media, and total cell number was analyzed after 24 h. (<b>B</b>) HUVEC were plated in the upper chamber with DMSO or individual diastereoisomer, while VEGF was added in the lower chamber and HUVEC invasion was studied. Abbreviations: Sil A: Silybin A; Sil B: Silybin B; Iso A: Isosilybin A, Iso B: Isosilybin B; *, p ≤ 0.001; #, p ≤ 0.01; $, p ≤ 0.05.</p