| Objective: Astragalus membranaceus(AS) is one of the most commonly used traditional Chinese herbal medicine, which possesses "Buqishengyang, Yiweigubiao, Tudushengji, Lishuituizhong" function. Modern pharmacological studies showed AS contains glycoside, polysaccharide, chromocor, amino acid, and microelement. Clinical usage suggested that AS is effective in many fields such as enhancement of immunological function, protection of hemopoiesis, etc.Recently, many studies on the anti-cancer effects of AS have been carried out. The results were quite controversial. Some studies showed that AS could inhibit the growth of tumors. Yet, there were some studies showed that AS may accelerate the growth of tumors. Up to now, the studies were carried out either on the improvement of immuno-function in vivo, or on the effects of tumor cells in vitro, or on the weight and volume of tumors in experimental animals. Few studies involving both tumor inhibiting and immunological function has been seen. There is no report of works on the systemic evaluation of effects of AS on tumor apoptosis, growth and immune system of the animal with tumor in whole animal level. To explore the systemic effects of ASA on H22 in mice and to objectively evaluate the putative advantages and disadvantage of AS on the animals with tumor, the following studies were carried out.Methods: 1 Animals and treatment:Fifty male Kunming mice, 18-20g in body weight were first inoculated with H22. The mice were then randomly divided into control and four ASA treatment groups. The mice in ASA treatment groups were treated by gavage with ASA at the dosage of 0.031 mg/g, 0.125mg/g, 0.5 mg/g and 2mg/g body weight respectively. The mice in control group were treated accordingly with normal saline. The experiment was repeated once. 2 Evaluation of tumor inhibiting effects:The mice were killed after 10 days of treatment. The mean weight and volume of the tumors were first measured and then the tumor tissues were divided into two parts. One part was fixed in 10% formaldehyde for the preparation for morphological observation and immunohistochemical staining of PCNA. The other was fixed in 70% ethanol for preparation of single cell suspension for flow cytometric analysis of apoptosis, proliferation and the expression of Bcl-2 and Bax. 3 Splenic CD4+ and CD8+ lymphocyte analysis:At the same time, splenic tissues from experimental mice were fixed in 70% ethanol and prepared for single cell suspension for the analysis of the fraction and ratio of CD4+ and CD8+ lymphocytes. 4 Peritoneal macrophages phagocytosis assay:Peritoneal macrophages phagocytosis of chicken RBC were carried out and the rate of phagocytosis and the index of phagocytosis were calculated under light microscope with Giemsa stain.Results: 1 The results of tumor inhibiting effects:In the first experiment, the mean weight and volume of the tumor in experimental groups were slightly lower than that in control group, but no statistic significance could be found(P?0.05). In the repeat experiment, there were no difference in both tumor weight and volume among the groups(P?0.05).2 Morphological observation: Areas of necrosis and hemorrhage and infiltration of lymphocytes could be seen in all the tumor tissue sections. The number of infiltrated lymphocytes in the 0.5mg/g and 2mg/g groups were 51.76±0.43, 73.94±10.20,which were significantly higher than that in control group (35.65±5.37, P<0.05).The HPIA-1000 analysis system revealed that there were no difference in the ratio of areas of necrosis among different groups (P?0.05). 3 The effects of AS on the proliferation: Positive PCNA nuclear staining cells could be seen in all groups by immunohistochemical staining. No difference in the positive expression rate of PCNA among the experimental groups and control group were found. FCM analysis showed that no significant differences in PI were seen between experimental groups and control group(P?0.05).4 The effects of AS on the apoptosis: FCM analysis revealed that the apoptosis rat... |
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