Allicin bladder cancer in mice in vivo and in vitro anti-tumor
effects of experimental study
【Abstract】 Objective To understand the allicin on mouse bladder tumor anti-tumor effect and mechanisms. Methods The MTT test of allicin directly assessed the cytotoxic activity, animal experiments showed that allicin has significant in vivo anti-tumor effect, and measured using LDH release assay cytotoxicity. The results of allicin directly to the bladder tumor cytotoxicity. High-dose treatment group compared with
control group, tumor growth rate was obviously inhibited (P <0.01), after treatment of allicin produced lymphocytes against B16 tumor cells. Conclusion of allicin on bladder cancer has obvious anti-tumor effect. This effect may be
related to direct cytotoxicity and immune responses. Allicin may be an effective treatment for superficial bladder cancer cavity.
Key words garlic; anti-tumor; bladder cancer
The Experimental Research on the Antitumor Effect of Allicin on Murine Bladder
Abstract: ObjectiveTo understand anti tumor effect and
mechanism of allicin on murine bladder.MethodsThe direct inhibition effects of allicin in vitro was evaluated by MTT Assay.Anti tumor effect of allicin in vivro was determined by animal experiment. CTL activity of lymphocyte was measured
by the release of endogenous lactate dehydrogenase (LDH). Results Allicin had significant direct cytotoxicity to bladder cancer cells. In high dose group, it showed a marked delay in the appearance and growth of tumors after sc injection
compared with control group (P <0.01 ). The body generated the cytotoxic lymphocyte responses to B16 tumor cells after treated with allicin.ConclusionAllicin has a marked anti
tumor effect on bladder tumor.This effect may be related to
direct cytotoxicity and immune response.It may be an effective intravesical treatment agent for superficial bladder cancer.
Key words: Allicin; Anti tumor effect; Bladder tumor
Liliaceae Allium garlic herb for two years, effective
ingredient allicin, folk medicine in China has 3000 years of history. Modern research suggests that garlic has a good anti-
tumor cancer, anti-cancer effects. Reports have suggested that garlic extract significantly inhibited the growth of bladder
cancer, but some experimental animals died from treatment-
related toxicity .
B16 tumor cells are the University of Tennessee (Tennessee University) MSSoloway use of oral N-formamide
(FANFT) from C3H mice induced from . This poorly differentiated transitional cell carcinoma demonstrated by the transfer of properties and human transitional cell carcinoma is similar. In order to understand allicin treatment of bladder tumors and mechanism, we measured allicin in vitro growth of B16 tumor cells and to assess its in vivo anti-tumor
effect and the impact on local immunity.
1 Materials and methods
1.1 Animals 30 6 ~ 8-week-old female C3H/He mice (weight
g) from Beijing Tong Lihua Victoria Experimental Technology Co., Ltd. purchased. All experiments are in line with the guidelines for animal experiments, Guangdong Medical College.
1.2 allicin B16 tumor cells and tumor cells by the Japanese Cancer Resource Library (JCRB) cell library provides (cell number: IFO50041). Cultured with 10% fetal calf serum
(FBS) in DMEM (GIBCO BRL, Life Technologies, MD, USA) and 37 ? in a 5% CO2. 10000 Allicin by the Guangdong-based
Pharmaceutical Ltd. (Approval number: State medicine accurate H44023561, production batch number 20,030,509, size 25 mg / extension; formula C6H10S3; molecular weight 178.33).
2.1 MTT experiment to establish cell culture 96-well
plates. In each hole containing 5 × 103B16 tumor cells, each
for a group of five holes. For the determination of allicin in vitro on tumor cell growth rate of the impact of different
doses of allicin (2.5, 0.5,0.1,0.02 mg * ml-1) and B16 cells
were cultured 48 h. Add MTT reagent train last 4 h. Wavelength of 560 nm on a microplate reader measured OD values.
2.2 The animal experiments for the determination of
allicin treatment of bladder cancer will C3H/He mice were divided into three groups, each containing 10 mice. 1 group was injected with normal saline 1 ml (control group), 2 group was injected with 12.5 mg of allicin (low-dose group), group 3
was injected 25 mg of Allicin (high-dose group). Planted tumor
cells were cultured with cell shovel from the bottle to collect, wash two times with normal saline. Its suspended in saline. The concentration adjusted to 1 × 106 cells / ml.
Cultivation of tumor cells into C3H mice after the
subcutaneous flanks, a dose of 0.1 ml. The next day will be injected into the tumor in the vicinity of allicin subcutaneous dose were 12.5 mg and 25 mg of allicin. Gauges measured twice a week with two tumor diameter to observe the
growth of tumors .
2.3 LDH release assay measuring cytotoxicity C3H/He mice were injected subcutaneously 25 mg of allicin once immunization, injected with normal saline as a negative control. Spleen cells were collected after 1 week. 96 Effect of culture medium plates with spleen cells from B16 will be target cells mixed in different proportions (mixing ratio from 12.5:1 to 100:1). Each group of three holes. Before the experiment, B16 target cells with mitomycin C injection (100 μg / ml) cultured for 1 h. 96-well culture plates in 37 ?
CO2 incubator 24 h. By LDH cytotoxicity kit provides methods and 492 nm detection wavelength. Figure 1.
Cytotoxic activity (%) = 100% × experimental group, LDH-
natural release of LDH release LDH-largest natural release of
2.4 Statistical analysis of all the results of
statistical analysis, analysis software for SPSS11, analytical methods for the analysis of variance after the calculation of P values of pairwise comparison. Reposted elsewhere in the
paper for free download http://
3.1 Effect of Allicin on the impact of tumor cells in vitro MTT test showed that when allicin dose 0.1 mg / ml, that is when there is moderate cytotoxicity, tumor inhibition rate could reach 55.7%, a dose of 2.5 mg / ml of bladder cancer cells have significant direct cytotoxicity (tumor inhibition rate reached 98.6%). The results in Table 1.
Figure 1 LDH release assay measuring cytotoxic activity (abbreviated)
Table 1 Effect of Allicin on B16 tumor cells in vitro
growth inhibition of (omitted)
3.2 allicin in vivo anti-tumor effect with the control
group compared allicin dose treatment group was significantly delayed tumor development. Injection 19 d after the control group tumor incidence reaches as high as 80%; while the low-
dose treatment group (12.5 mg) of tumor incidence was 50%;, but high-dose treatment group (25 mg) of tumor incidence was only 20% (Figure 2). In the first 19 days, saline control group, low-dose treatment group and high-dose treatment group,
tumor size, respectively (0.86 ? 1.20), (0.73 ? 1.06) and
(0.22 ? 0.54). High-dose treatment group compared with the
saline control group, tumor growth rate was obviously inhibited (P <0.01). The results in Table 2.
Allicin 3.3 after treatment, resulting in B16 tumor cells against cytotoxic T lymphocytes
To study whether this treatment can promote the in vivo lymphocyte cytotoxic activity of tumor cells, the C3H mice were injected 25 mg of allicin immunization, injected with
normal saline as negative control. Shown in Figure 2, produced after immunization of allicin against B16 tumor cells, cytotoxic lymphocytes , while the normal saline does not have this effect.
Garlic (Allium sativum)-diene-C chemical called allyl
trisulfide, is the main component of garlic. 1997 DRRiggs such as found in the old garlic extract concentration of 31.3 mg / ml, the inhibition of B16 tumor cells rate was 29.9%; when the
concentration of 62.5 mg / ml or higher than at this time, the old garlic extract on bladder cancer cells obviously a direct cytotoxic activity (inhibition rate 98%) .
Table 2 garlic treated tumor area (omitted)
High dose of allicin compared with the control group, * P
Figure 2 garlic tumor incidence after treatment (abbreviated)
We observed in our study of allicin on tumor cells has a direct cytotoxic activity. MTT results show that allicin concentration of 20 μg / ml that contained moderate direct cytotoxicity, inhibition rate could reach 41%; when the concentration of 2.5 mg / ml you can see a direct and significant cytotoxic activity (tumor suppressor 98.6 %). Garlic extract in comparison with the older, the biological
activity of allicin is 25 times stronger. This may be because the main component of the two different, although they are derived from garlic, garlic extract, but the old main ingredient is S-allyl cysteine.
Effect of Allicin on the mechanism of direct inhibition
of tumor cells is still unclear. The cytotoxic effect of allicin may be by lowering the level of active oxygen species
free radicals affect NFκB activity in tumor cells, so that NFκB-DNA binding activity to reduce, NFκB pathway is
inhibited, leading to some changes in gene expression levels ; or through the reduce the aromatic amines N
acetyltransferase, and 2 aminofluorene activity, and thus
inhibit tumor cell growth . In addition, the anti-tumor
effect of allicin Another important mechanism is to stimulate the immune system.
Animal experiments showed that allicin has significant in
vivo anti-tumor effects. This effect is dose-related. In the
high-dose treatment group, both the incidence of cancer or tumor growth rate were significantly lower than the low-dose
LDH release assay measuring cytotoxicity experiment
results also show that, after injection of allicin, which can produce against the B16 tumor cells, cytotoxic T lymphocytes. Allicin garlic stimulates the immune mechanism and is similar. Some studies suggest that garlic can cause IL 2, TNF α
and IFN γ release, and enhance phagocytosis . Other activities include the promotion of garlic killer cell activity and mitogen-stimulated lymphocyte proliferative
response of the immune . These effects, especially in cytokine release pattern, indicating that allicin activate the Th1 cellular immune response, this response is an effective anti-tumor-specific immune therapy.
In conclusion, allicin pairs of bladder tumors has obvious anti-tumor effects. This effect may be related to
direct cytotoxicity and immune responses. This study describes the allicin on the biological activity of bladder cancer and support such a view that allicin is an effective
chemopreventive or chemical treatments. Allicin may be an effective treatment for superficial bladder cancer, cavity, and the price low, adverse reactions is small, worthy of clinical use.
 Lamm DL, Riggs DR.The potential application of Allium sativum (garlic) for the treatment of bladder cancer
[J]. Urol Clin North Am 2000,27 (2): 157.
 MickeyDD, Mickey GH, Murphy WM et al.In virto characterzation of four N-[4 (5 nitro 2 furyl)
2 thiazoly] formamide (FANFT) induced mouse bladder tumors. [J]. Urol, 1982, Jun, 127 (6): 1233.
 Research since.1998 Epidemiology, Shanghai Cancer Institute, Cancer incidence in Shanghai urban area [J]. Cancer, 2001,21 (4): 312.
 Wang Jian, Tan tomorrow, Wu Hong, et al. Urological cancer inpatients survey [J]. Guangdong Medicine, 2001,22
 Riggs DR, DeHaven JI and Lamm DL.Allium sativum (garlic) treatment for murine transitional cell carcinoma [J]. Cancer 1997,79 (10): 1987.
 Dirsch VM, Gerbes AL and Vollmar AM.Ajoene, a compound of garlic, indeces apoptosis in human
promyeloleukemic cells, accompaied by generation ofreactive oxygen species and activation of nuclear factor happaB. [J]. Mol Pharmacol .1998,53 ( 3): 402.
 Lin JG, Chen GW, Su CC, et al.Effects of garlic
components dially sulfide and diallyl disulfide on arylamine N-acetyltransferase activity and 2-aminofluorene-DNA adducts
in human promyelocytic leukemia cells.Am [J]. Chin Med .2002,30 (2 ~ 3): 315.
 Dausch JG, Nixon DW.Garlic: A review of its
relationship to malignant disease [J]. Prev Med 1990,19 (3): 346.
 Kyo E. Suzuki A. Kakimoto M, etal.Immunomodulation and antitumor activities of age garlic extract [J]. Phyto medicine 1998,5 (4): 259. Reposted elsewhere in the paper for
free download http://www. hi138.com