Sentences with word «kahweol»

Different in vitro assays were carried out in order to test the specific effects of kahweol treatment on several key steps of the angiogenic process in both endothelial and tumor cells.

Figure 6 shows the effects of 75 µM kahweol on endothelial cell migration, as determined by the «wound healing» assay, after 8 and 24 h of treatment.

A second experimental approach used to test the effects of kahweol on in vivo angiogenesis was the use of a model of transgenic zebrafish.

A) Gelatin zymography of MMP - 2 in conditioned media of HUVEC after treatment with different kahweol concentrations.

The minimal inhibitory concentration for kahweol in this assay of «tubule - like» structures formation on Matrigel was 25 µM, in the range of concentrations at which other known antiangiogenic compounds produce this kind of effect [13], [27].

Figure 9 (A and B) shows that kahweol inhibits in a dose dependent manner the expression of COX - 2 protein by HUVEC.

To get new, additional insights on the features of kahweol as an anti-angiogenic compound, we carried out a complete set of in vitro assays previously used by us to characterize the anti-angiogenic effects of other compounds from natural sources, including aeroplysinin - 1, homocysteine, ursolic acid, puupehenone, hypericin, hyperforin and aloe - emodin, among others [11], [12], [13], [19], [20], [21].

The global morphological features (including centrifugal growth of the peripheral vessels - relative to the position of the disc -, avoiding the treated area, with an overall decrease in the vascular density) elicited by kahweol treatment are also in agreement with those previously observed for other anti-angiogenic compounds.

Figure 2 (middle line) shows that 50 nmol kahweol induced apoptosis in a small percentage of cells but this effect did not seem to be endothelial cell specific, since only few apoptotic nuclei corresponded to endothelial cells.

On the other hand, the anti-oxidant nature of kahweol also points to its potential anti-inflammatory capabilities.

Taken together, our data indicate that, indeed, kahweol behaves as an anti-inflammatory and anti-angiogenic compound with potential use in antitumoral therapies.

The CAM and zebrafish in vivo assays and the ex vivo mouse aortic ring assay clearly identify kahweol as a new anti-angiogenic compound, but gives no information on which specific steps of angiogenesis are targeted by this compound.

Medicinal compounds detected in coffee include lignans, quinides, antioxidants, dipertenes like kahweol and cafestol...

In fact, the rate of invasion was inhibited a 23, 33 and 52 % (as compared to the rate of invasion for control, untreated cells) by 25, 50 and 75 µM kahweol treatments, respectively.

Our results indicate that kahweol inhibits two key inflammatory mediators, COX - 2 and MCP - 1, which are also related with angiogenesis.

Therefore, it would be advisable to test the potential effects of kahweol on endothelial cell apoptosis.

Figure 2 (upper lane) shows that kahweol treatment induced disorganization and inhibition of the ingrowth of new vessels in the area covered by the discs containing the compound.

Survival curves of proliferative (squares) and non-proliferative (circles) HUVEC endothelial cells treated with kahweol.

On the other hand, these data suggest that the potential effects of kahweol on apoptosis could exhibit certain cell specificity.

Subsequently, additional in vitro angiogenesis assays were carried out both in the absence and presence of kahweol in the range of concentrations of its IC50 value in the MTT assay.

First of all, the MTT cell proliferation assay was carried out to determine the long term (after three days of incubation) cytotoxicity of kahweol and to evaluate its IC50 value for endothelial cells, as described by us elsewhere [10].

Figure 8 (A) shows that kahweol inhibits HUVEC MMP - 2 expression, with a clear dose response effect and complete inhibition by 50 µM kahweol.

On the other hand, the present research work shows a confirmatory evidence of the potential of kahweol to inhibit in vivo angiogenesis, by using another completely independent model system, namely, that of genetically modified zebrafish.

Figure 8 (C) shows that kahweol - treatment induces a dose - dependent decrease in the levels of urokinase in HUVEC conditioned media, with an almost complete inhibition at 50 µM kahweol.

Since invasion is dependent on extracellular matrix remodeling capabilities, this inhibitory effect strongly suggested that the two key extracellular membrane remodeling enzymes expressed by endothelial cells, namely, MMP - 2 and uPA could be other main key targets of the pharmacological action of kahweol on endothelial cells.

Furthermore, a clear dose - response inhibition of microvessel formation was observed and quantified in treatments with kahweol.

In contrast, Video S2 shows that larvae treated with 50 µM kahweol for 24 h exhibited no blood flow along intersegmental vessels.

A quantitative analysis of these effects revealed that 85 % of 75 µM kahweol - treated and 75 % of 25 µM kahweol - treated larvae exhibited inhibited angiogenesis (results not shown).

The results shown in the present study clearly indicate that kahweol is another natural anti-angiogenic compound with a wide spectrum of targets [10], [11], [12], [19], [20].

In the CAM assay, the inhibitory doses exhibited by kahweol are similar to those of other anti-angiogenic compounds found by us to inhibit angiogenesis in the CAM assay [10], [19], and much lower than those of other anti-angiogenic compounds [21].

The results obtained in the «wound healing» assay clearly show that 75 µM kahweol is able to inhibit endothelial cell migration.

Six eggs were used for each tested dose of kahweol.

Our results reinforce the potential pharmacological interest of kahweol, as suggested by its anti-angiogenic and anti-inflammatory effects.

Invading controls and 25 and 75 µM kahweol - treated HUVEC cell count values are represented by using diamonds, squares and triangles, respectively.

These data may contribute to the explanation of the reported antitumoral effects of kahweol, including the recent epidemiological meta - analysis showing that drinking coffee could decrease the risk of certain cancers.

Altogether, our results demonstrate that kahweol is a potent anti-angiogenic compound both in vitro and in vivo, targeting some key steps shared with tumor progression, key molecules involved in ECM remodeling (MMP - 2 and uPA), and key molecules involved in inflammation (COX - 2 and MCP - 1).