| As one of the biggest threats to human health, influenza is an acute respiratory infection caused by influenza virus with characteristics of high morbidity and high mortality.According to some data, more than50million people around the world would die of this desease each year. The morbidity of influenza keeps10%-30%these years.Since1957, there have been three worldwide influenzas originated from China, and China has become a country with high morbidity of influenza. A survey has shown that the influenza mortality was as high as0.6%~1.4%in the1950s. H5N1avian flu has broken out frequently in recent years. The mortality has been up to a new high of66%-80%since1997. H7N9, a new type of bird flu firstly seen in Shanghai and Anhui, is characterized with speed infection, serious symptoms and high death rate of35%.The main causes of death in lethal influenza are respiratory complications, especially severe pneumonia (including viral pneumonia and mixed infection of viruses and bacteria) and respiratory distress syndrome (ARDS). Thus, an effective treatment is urgently needed for severe flu, but at present, it still remains a great clinical difficulty.Influenza is defined as exogenous fever by traditional Chinese medicine (TCM). It is proved that the TCM dialectical therapy for H1N1flu had been applauded both at home and abroad. H1N1is mainly featured with fever. It is with the nature of warm disease. It is manifested as high fever with heat toxin, wasting qi and damaging fluid, and bleeding caused by heat. The early stage of the disease tends to be excess pattern, and the main pathogenesis is toxic-heat, stasis, and qi yin difficiency.Based on the theory of TCM, there are two reasons accounting for influenza. One is that the rising heat inside the patients results in the great consumption of body fluid.In this case, the patient suffers from troubles in breathing, unconsciousness and even death; the other is the increasing heat leads to stagnating and bleeding.As a result of the above pathogenesis, two therapeutic treatments with each own emphasis are given by TCM. One treatment is to drive the heat inside patients out while the other is to enhance the patients’immunity by tonifing qi and nourishing yin. In view of this controversy, this thesis responds to these clinical problems and different point of views concerning influenza on the basis of animal experiment.PURPOSEThere were three kinds of injections,including Reduning Injection with function of clearing heat and resolving toxin, Xuebijing Injection with function of cooling blood and removing stasis, and Shenmai Injection with function of tonifying qi and nourishing yin.Our study was to observe the curative effects of any two injections combination and explore the optimal compatibility of Chinese Medicines and its mechanism of action.METHODExperiment OneTo research which kind of mouse, C57BL/6or Balb/C mouse,was susceptive to BJ501strain by survival rate and wet-to-dry ratio.And finally the model mouse and virus load were confirmed by lung pathology.Experiment Two175mice of SPF C57BL/6,4-6weeks,were randomly divided into seven groups,including normal group,model group,Re plus Xue group, Re plus Shen group, Xue plus Shen group, Re plus Shen(48h) group, Xue plus Shen(48h) group.Among the treatment groups,every injection was given immediately after infection,while Shen(48h) was given48h post-infetion.The treatment dosage of each injection was equal to their maximum usage in clinical. All above injections were given continuously5days. Death rate, weight change(n=10), lung wet-to-dry ratio(n=5),lung pathology (n=5),the level of bronchoalveolar lavage fluid(BALF) cytokines,including IL-6, IL-10, MCP-1, IFN-r and TNF-r(n=5) were the measuRements.RESULTS1. Survival Rate,Weight Change and General Condition1) Survival rateAfter one infection cycle,all mice in model group survived.There were three mice left in model group,with the survival rate30%. The survival rate in Re plus Xue group was50%,in both Re plus Shen group and Xue plus Shen group was40%. But all mice in Re plus Shen group died. The survival rate in Xue plus Shen group was the same as that of model group.2) Weight changeAll infected mice began to lose weight after two days infection.The weight lose of mouse in Re plus Xue group decreased slowly while Re plus Shen(48h) group the most quickly.The trend of mouse weight in Xue plus Shen(48h) was similar to that of model group.The weight change in all treatment groups in which TCM injections was given immediately after infection were superior to that in model group.3) General situationMice in Re plus Xue group were in good condition both physically and mentally. In contrast, the situation in other treatment group had poor spirit,difficult breath, and cold body which was similar to that of model group.2.Lung Wet-to-dry RatioAt5days post-infetion, wet-to-dry ratio in all treatment groups was measuRed(n=5).Lung wet-to-dry ratio in Re plus Shen group and Xue plus Shen group were both lower than that of model group whereas there were no statistically significant difference (P>0.05). The wet-to-dry ratio of Re pius Xue group was significantly less than that of model group (P<0.05). The wet-to-dry ratio of Xue plus Shen(48h) group was similar to the model group while the wet-to-dry ratio of Re plus Shen(48h) group was over that of model group,with no statistically significant diffeRence (P>0.05).3. Lung PathologyModel group, inflammation activity(+++),representative pathological manifestation were diffuse inflammatory cell infiltration, shedding and necrosis of alveolar epithelial cell, and bronchial epithelial cell degeneration accompanied by edema, congestion, hemorrhage. Re plus Xue group, inflammation activity(++),it was also diffuse inflammatory cell infiltration, shedding and necrosis of alveolar epithelial cell and alveolar space became thicker. Re plus Shen group, inflammation activity(+++~++++),there were diffuse inflammatory cell infiltration, shedding and necrosis of alveolar epithelial cell and fibrous exudation to accompany. Xue plus Shen group, inflammation activity(++),there were shedding and necrosis of alveolar epithelial cell with inflammatory cell infiltration in alveolar septum and mild fibrous exudation. Re plus Shen(48h), inflammation activity(++++),it was lung abscess, lung consolidation, diffuse inflammatory cell infiltration,bronchial epithelial hyperplasia that was similar to Xue plus Shen(48h) group,with inflammation activity(++++).4. BALF Cytokines AssaysAfter48h post-infetion, the infected mouse BALF level of IL-6IL-10MCP-1, IFN-y, TNF-a etc had risen markedly,especially IL-6, IL-10MCP-1both at48h and96h. But at96h, the level of MCP-1had gradrually descended and the level of TNF-a had been back to normal. All treatment group could Reduce the level of IL-6, IL-10IFN-y to varying degRees at48h. After96h post-infetion,the level of IL-6in Re plus Xue group,Re plus Shen group and Xue plus Shen group were down, well into the normal range wheReas the level of IL-6in Re plus Shen(48h) group and Xue plus Shen(48h) group were lower than that of normal group. All treatment group could reduce the level of both IL-10and IFN-y, especially Re plus Shen(48h) group.CONCLUSION Compared with Balb/C mice, C57BL/6mice were susceptible to influenza BJ501strain.4-6weeks old C57BL/6mice infected by BJ501strain could successfully induced mouse model of seveRe influenza pneumonia.According to the trial, any two injections including Reduning Injection with function of clearing heat and resolving toxin, Xuebijing Injection with function of cooling blood and removing stasis, and Shenmai Injection with function of tonifying qi and nourishing yin, can be combined to tReat the infected rats in incubation periods of influenza. And the treatments are effective. The combination of Reduning Injection and Xuebijing Injection are the most effective, which suggests that the early intervene with Chinese Medicine Injection is very important for the prognosis.Shenmai Injection with function of tonifying qi and nourishing yin is ineffective to mice with early stage of severe pneumonia at48h post-infetion, or even leading the death of all rats. The reason may be that the adaptive immune response starts when medicating, while Shenmai Injection can make the immune response inhibited. It suggests that the use of medicine with function of tonifying qi and nourishing yin should be cautious at the early stage of warm disease. |
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