CN101849526A - Qi-deficiency fever animal model and preparation method and application thereof - Google Patents
Qi-deficiency fever animal model and preparation method and application thereof Download PDFInfo
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- CN101849526A CN101849526A CN201010186415A CN201010186415A CN101849526A CN 101849526 A CN101849526 A CN 101849526A CN 201010186415 A CN201010186415 A CN 201010186415A CN 201010186415 A CN201010186415 A CN 201010186415A CN 101849526 A CN101849526 A CN 101849526A
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Abstract
The invention discloses a Qi-deficiency fever animal model and a preparation method and application thereof. The Qi-deficiency fever animal model is a rat Qi-deficiency fever model obtained by a swimming strain method and a controlled feeding method. The method for preparing the Qi-deficiency fever animal model comprises the following steps of: controlling the feeding of a rat and making the rat exhausted after swimming, and injecting LPS into the abdominal cavity of the rat. The Qi-deficiency fever animal model can be used for Chinese medicinal clinical syndrome differentiation and treatment of Qi-deficiency fever diseases, and can provide a platform for researching the Chinese medicinal treatment of the Qi-deficiency fever diseases.
Description
Technical field
The present invention relates to the model field of traditional Chinese medical science disease research, be specifically related to a kind of qi-deficiency fever animal model that is used for the research of fever due to deficiency of vital energy illness and preparation method thereof.
Background technology
The modeling method of relevant various syndrome of qi deficiency was various informative in recent years, there is fatigue to add intemperance of taking food or rushes down down, partial eclipse bitter taste or tart flavour, under rushing down, consumption gas relieving stagnant Qi or bitter cold add (Li Junlan such as consumption gas relieving stagnant Qi, Fang Zhaoqin. syndrome of qi deficiency Establishing Animal Model of Syndrome [J]. Shanghai Univ. of Traditional Chinese Medicine's journal, 2004,18 (3): 56-60.).At present the deficiency of vital energy modeling method of relatively generally acknowledging mainly contains that simple tired method, fatigue add the intemperance of taking food method, fatigue adds purgative therapy, fumigation and operation (Wu Jie, Liu Qinshe, noble and unsullied, Deng. the influence [J] of woman's recovering particles agent of nourishing blood to experimental qi and blood asthenic symptoms animal. the Shaanxi traditional Chinese medical science, 2002,23 (5): 467-469.; Chen Jiaxu, Chen Yixin, Ji Shaoliang, etc. the variation [J] that rush down down, the overstrain factor causes mouse hypothalamus under the temper void, blood plasma NPY content. Beijing University of Chinese Medicine's journal, 2002,25 (6): 22-24.; The field Dow process, Tang Faqing, Zhou Xiaojun, etc. the Primary Study [J] of syndrome of qi deficiency rat model antisense DNA array A district's gene expression spectrum signature. Hunan College of Traditional Chinese Medicine's journal, 2001,21 (2): 1-3.; Gao Xue, Li Hao, song respect, Deng. tonifying lung soup is to the influence [J] of syndrome of deficiency of lung qi rat lower respiratory tract pathological change. traditional Chinese medicine information, 2000,17 (5): 55-56.), but, on the basis of fatigue, add with the limit food, rush down inferior method generally to adopt tired method.There are some researches show, use the tired animal model of qi-asthenia that adds the making of limit food method and more meet theory of traditional Chinese medical science that can make gas more not enough, deficiency of vital energy disease is more obvious.
The deficiency of vital energy is many by the native endowment deficiency, or the day after tomorrow lose to support, or internal lesion caused by overexertion excessively consumes (overstrain may lead to decline of vital energy), or prolonged illness is multiple, due to the biochemical deficiency of gas etc. (Zhang Zhenyu. to the understanding [J] of traditional Chinese medicine gas. the Zhejiang College Of Traditional Chinese Medicine journal, 2000,24 (1): 32-32.), can cause the generation of multiple disease clinically, heating is one of them, calorific potential or high or low, but caused by vital energy deficiency, so many outbreak or the aggravations after fatigue of heating.The experimental rat fever model is mainly with yeast, endotoxin (LPS) and 2 at present, and the 4-dinitrophenol dinitrophenolate is the most common, and its common characteristic is that modeling is convenient, and the success rate height is also lower to requirement for experiment condition.Wherein the LPS heating of inducing is similar to heating due to the traditional traditional Chinese medicine disease, it is classical fever model, therefore be usually used in screening the Chinese medicine antipyretic analgesics and inquire into heating mechanism (Tang Xiaofeng, Xue Manqing, Wang Hui. rat fever model and heating Study and advance on mechanism [J]. ACAD J GCP, 2009,25 (3): 327-331.).
PGE be the maincenter heating medium of generally acknowledging (the inscription chief editor of Jinhui. Pathological Physiology. Beijing. the People's Health Publisher, the 5th edition, 2001:96.).Research data shows that pyrogen causes adstante febre, PGE in cerebrospinal fluid and the hypothalamus
2Content obviously increases.The animal telocoele injects the PGE of trace
2Can induce exothermic reaction, and PGE
2Can directly stimulate PO/AH district responsive to temperature neuron, change its discharge frequency, suppress PGE material synthetic and that discharge and can suppress exothermic reaction (Zhao Hongyan, the model book is selected, Sun Lihua etc. baicalin is to the influence [J] of heating rat hypothalamus PGE2 and cAMP content. Chinese applied physiology magazine, 2002,18 (2): 139-141.).According to data in the past, domestic most scholars advocate that cAMP also is one of maincenter medium of heating.When intravenous injection ET such as Hu Jingxin or EP cause the rabbit diphasic fever, the equal showed increased of cAMP content in cerebrospinal fluid and the hypothalamus, and be remarkable positive correlation (tensor with the body temperature rising, Tong Lei, Yang Yu etc. Magainin is to the influence [J] of IL-1 β pyrogenicity rat hypothalamus and plasma cAMP content during low temperature. Chinese clinical pharmacology and acology, 2005,10 (12): 1354-1357.).
Basic Theories of Chinese Medicine is thought (Zhang Huimin, Zheng keeps once. the progress of deficiency of vital energy physique [J]. and Beijing University of Chinese Medicine's journal, 2004,27 (1): 14-16.), essence, gas, blood, body fluid are formations and keep human life activity's base substance, the deficiency of vital energy then green blood deficiency, the deficiency of vital energy then hematogenous blockage, the deficiency of vital energy then stop in Tianjin, so the deficiency of vital energy often with the deficiency of blood, blood stasis, the phlegm wetting phase folder of holding concurrently mutually, form complicated physique.The perfectly sound speech of the Ming Dynasty: " ... as to be subjected to the gas of lung be fur, and insufficiency of lung-QI,deficiency of lung-QI, then skin is thin and brittle timid cold, and hair is not given birth to; The gas that is subjected to the heart is blood vessels, deficiency of heart-QI, and the not magnificent look of blood then, face does not have brilliance; The gas that is subjected to spleen is meat, the temper subsidy, and then muscle is not given birth to, and brothers are as cutting; The gas that is subjected to liver is muscle, deficiency of liver-QI, and then bone is soft." also there are some researches show, make disease in conjunction with animal model be research fever due to deficiency of vital energy disease the important channel (Luo Yaoyue. syndrome of blood stasis due to qi deficiency The Animal Model Study commentary [J]. Hunan College of Traditional Chinese Medicine's journal, 2004,24 (5): 57-58.).
Summary of the invention
The objective of the invention is to according to the research of existing qi-deficiency fever animal model lessly, and the fever due to deficiency of vital energy illness is one of common clinical disease, adds limit food method by swimming strain method, and a kind of qi-deficiency fever animal model is provided.
Another purpose of the present invention is to provide the preparation method of above-mentioned qi-deficiency fever animal model.
A further object of the invention is to provide the application of above-mentioned qi-deficiency fever animal model.
Above-mentioned purpose of the present invention is achieved by the following technical programs:
A kind of qi-deficiency fever animal model, this model are to adopt swimming strain method to add limit food method to obtain rat fever due to deficiency of vital energy model.
The preparation method of qi-deficiency fever animal model of the present invention comprises the steps: rat is kept on a diet and power exhausts swimming, lumbar injection LPS.
As a kind of preferred version, among the above-mentioned preparation method, described keeping on a diet is that feeding amount with rat is reduced to 1/2～1/3 of regular diet amount.
As a kind of preferred version, among the above-mentioned preparation method, when described rat power exhausts swimming, can not have a rest in the bottom with shoe, continue the sinking of swimming, till still can not returning the water surface behind the 10s, swim every day 1～2 time, continued for 3～4 weeks.
As a kind of preferred version, among the above-mentioned preparation method, the water temperature when described rat power exhausts swimming is 18～25 ℃.
As a kind of preferred version, among the above-mentioned preparation method, the amount of described lumbar injection LPS is 60～100 μ g/kg, most preferably is 80 μ g/kg.
Find by experiment of the present invention, the rat of deficiency of vital energy disease by hair sparsely easily come off, dull uninteresting, carry out lung pathology after the experiment and learn and check and find in its lung tissue a small amount of inflammatory cell is arranged, epithelial cell is impaired to present cilium lodging, adhesion; Normal rat bronchial mucosa structure and lung tissue structure are clear, pathological changes such as no epithelium necrosis comes off, cell infiltration, thus confirm to have certain relations of dependence really between lung and the fur.And carry out lienopathla section of science also prompting morphological change is arranged, some can judge tentatively that the inventive method can successfully make the fever due to deficiency of vital energy rat model from this.
Qi-deficiency fever animal model of the present invention can be used for the tcm clinical practice diagnosis and treatment of fever due to deficiency of vital energy illness, and the research that also can be Chinese traditional treatment fever due to deficiency of vital energy illness provides platform.
Compared with prior art, the present invention has following beneficial effect:
The present invention also has lumbar injection LPS by rat being kept on a diet and its swimming strain power being exhausted, and has obtained qi-deficiency fever animal model.Data shows that little the opening of the big rathole of qi-deficiency fever animal model of the present invention do not have god, and autonomic activities is few, reacts the section of eating to external world, and escape, attack obviously reduce; Body temperature is higher, can be up to 39.4 ℃ and hot journey longer; The histopathology sections observation lung partial groups of fever due to deficiency of vital energy model as can be known is woven with inflammatory cell infiltration, and it is fuzzy that institutional framework appears in part, alveolar septum attenuation or fracture.So the rat fever due to deficiency of vital energy model that the inventive method obtains is compared with existing fever due to deficiency of vital energy model, gas is more not enough, deficiency of vital energy disease is more obvious, and it is similar to heating due to the traditional Chinese medicine disease by the heating paresthesia that LPS induces, it is classical fever model, therefore the fever due to deficiency of vital energy symptom of rat fever due to deficiency of vital energy model of the present invention is more remarkable, for Chinese traditional treatment fever due to deficiency of vital energy illness provides good research platform.
Description of drawings
Fig. 1 is that the behaviouristics of normal rats is observed;
Fig. 2 is that the behaviouristics of deficiency of vital energy group rat is observed;
Fig. 3 is that the behaviouristics of fever due to deficiency of vital energy group rat is observed;
Fig. 4 be behind the lumbar injection LPS rat put question to raise (Δ T) influence (℃);
Fig. 5 is the pathological section figure of normal rats lungs;
Fig. 6 is the pathological section figure of deficiency of vital energy group rat lungs;
Fig. 7 is the pathological section figure of fever due to deficiency of vital energy group rat lungs;
Fig. 8 is the pathological section figure of normal rats spleen;
Fig. 9 is the pathological section figure of deficiency of vital energy group Rats Spleen;
Figure 10 is the pathological section figure of fever due to deficiency of vital energy group Rats Spleen.
Embodiment
Further explain the present invention below in conjunction with embodiment, but embodiment does not do any type of qualification to the present invention.
The preparation of embodiment 1 qi-deficiency fever animal model
1. laboratory animal SD rat, 220～250g, SPF level, male and female half and half, Traditional Chinese Medicine University Of Guangzhou's Experimental Animal Center provide [quality certification number: SCXK Guangdong 20080020].
2. (Escherichcacoliell-dotoxin 055:B5 L2880) purchases the company in Sigma for medicine and reagent lipopolysaccharides LPS; PGE
2, cAMP puts and exempts from medicine box: the good reagent of Guangzhou prestige Co., Ltd; Computer numeral thermometer MC-3B type is produced by OMROM Omron (Dalian) Co., Ltd.
3. experimental technique is divided into normal group, deficiency of vital energy group, fever due to deficiency of vital energy group at random with 30 SD rats, 10 every group.Normal group is conventional raises, free diet, and keeping on a diet (feeding amount at ordinary times 1/2) and carry out power for all the other two groups exhausts swimming, and water temperature is controlled at 25 ℃, contains in the plastics bucket.During rats'swimming, can not at the bottom of the pond, have a rest with shoe, 1 time/day, continue 21 days, (this experiment exhausts criterion (field Dow process with reference to the power that MCARDLE recommends to set up deficiency of vital energy animal model, Tang Faqing, Zhou Xiaojun, Deng. the Primary Study [J] of syndrome of qi deficiency rat model antisense DNA array A district's gene expression spectrum signature. Hunan College of Traditional Chinese Medicine's journal, 2001,21 (2): 1-3.), the last sinking of i.e. swimming, still can not return the water surface behind 10s is that power exhausts), fever due to deficiency of vital energy group rats by intraperitoneal injection LPS (LPS 80 μ g/kg), normal group, deficiency of vital energy group do not process.Write down rat body weight in the experimentation every other day, and observe to rat behavior; Get the rat hypothalamus tissue, adopt the ELISA standard measure to measure PGE
2, cAMP content; Taking-up spleen, lung carry out pathological examination.
The behaviouristics of embodiment 2 qi-deficiency fever animal models is observed
To the state of mind of each group rat, the reaction agility, the faecal volume size, daily routines such as defecation frequency are observed.
The result: the normal rats performance is flexible, and eye has god, is swift in response, and by the pure white gloss of hair, faecal volume is big, and defecation frequency is few.The gentle big rathole of thermal model group of shooting without hitting the target of the deficiency of vital energy is little opens does not have refreshingly, and autonomic activities is few, and is slow in reacting to external world, escape, attack obviously reduces, by hair sparsely easily come off, dull uninteresting, faecal volume is little and rare, defecation frequency few (Fig. 1～3).
The body weight change of embodiment 3 qi-deficiency fever animal models
Write down rat body weight in the experimentation every other day.
The result: the normal rats body weight increases gradually, and the deficiency of vital energy is gentle, and the thermal model group rat body weight of shooting without hitting the target significantly descends along with the prolongation of modeling time, has compared statistical significance (P＜0.05 or P＜0.01) with normal group.The results are shown in Table 1.
The variation of table 1 rat body weight (g) (n=10,
)
Annotate: compare with normal group: * P＜0.05, * * P＜0.01;
The body temperature of embodiment 4 qi-deficiency fever animal models changes
2d before the modeling measures sooner or later and respectively organize rat anus temperature every day, chooses body temperature at 36.6～38.0 ℃, and body temperature changes＜0.3 ℃ rat continuously, gets its mean value as arm's length basis body temperature (T
0); After the modeling, measure anus temperature (T every 30min
i), continuous measurement 8h calculates body temperature changing value Δ T (Δ T=T
i-T
0).
As can be seen from Figure 4, compare with normal group, deficiency of vital energy group and the fever due to deficiency of vital energy group rat 0.5h after injection LPS (80 μ g/kg) each point of observation Δ T in the 8h obviously raises, and significance poor (P＜0.05) is arranged statistically; The basal body temperature of deficiency of vital energy group in whole experiment is all higher, and reaches 38.5 ℃ of maximum temperatures about 1.5h, fluctuation among a small circle after this; The fever due to deficiency of vital energy group is behind injection LPS 2.5h, and body temperature has significantly rising, has compared significant difference (P＜0.05) with deficiency of vital energy group, and 5h left and right sides body temperature peaks 39.4 ℃, after this descend gradually, but decline scope is not obvious, and hot journey is longer, reaches more than the 8h; Normal rats body temperature is relatively stable, remains at fluctuation in the normal body temperature.
PGE in the inferior colliculus cerebral tissue of embodiment 5 qi-deficiency fever animal models
2Detection with the cAMP content
As can be seen from Table 2, cAMP, PGE in the fever due to deficiency of vital energy group rat hypothalamus
2Content has the significant difference (P＜0.01) of highly significant apparently higher than normal group.The result shows that cAMP content raises in the inferior colliculus cerebral tissue, and inferring may be because Ca
2+Concentration decline (Na
+/ Ca
2+Rise) institute causes that the heating medium increases and causes, and therefore measures the interior Na of hypothalamus
+/ Ca
2+Value will be for the stronger evidence of providing of its mechanism be provided.This experimental result also shows when lumbar injection LPS causes cooling, PGE in the hypothalamus
2Content reduces, and points out it may pass through to suppress PGE
2Secretion process reduce body temperature.
PGE in table 2 inferior colliculus cerebral tissue
2With cAMP content (n=10,
)
Annotate: compare * p＜0.01 with deficiency of vital energy group.
Embodiment 6 qi-deficiency fever animal model histopathology sections observation
The histopathology cut sections for microscopic examination: experiment finishes the back sacrificed by decapitation, gets spleen, that lung is put into 10 volume % formalin solutions is fixing, and 24h is after the dehydration of ethanol gradient, and dimethylbenzene is transparent, puts under the Electronic Speculum after waxdip, section, the HE dyeing and observes.
Fig. 5～7 are induced lung popular name for reason slice map, as can be seen, normal rats lung tissue structure is normal, lung tissue structure is clear, alveolar and lung there is no obvious morphological change at interval, the deficiency of vital energy and fever due to deficiency of vital energy group induced lung partial groups are woven with cell infiltration, and it is fuzzy that institutional framework appears in part, alveolar septum attenuation or fracture.
Fig. 8～10 are Rats Spleen pathology slice maps, from figure as can be seen, compare with normal group, and the deficiency of vital energy and fever due to deficiency of vital energy group Rats Spleen partial groups are woven with cell infiltration, and it is fuzzy that institutional framework appears in part.
Claims (8)
1. a qi-deficiency fever animal model is characterized in that it being to adopt swimming strain method to add limit food method to obtain rat fever due to deficiency of vital energy model.
2. the preparation method of the described qi-deficiency fever animal model of claim 1 is characterized in that comprising the steps: rat is kept on a diet and power exhausts swimming, lumbar injection LPS.
3. according to the preparation method of the described qi-deficiency fever animal model of claim 2, it is characterized in that described keeping on a diet is that feeding amount with rat is reduced to 1/2～1/3 of regular diet amount.
4. according to the preparation method of the described qi-deficiency fever animal model of claim 2, when it is characterized in that described rat power exhausts swimming, can not have a rest in the bottom with shoe, continue the sinking of swimming, till still can not returning the water surface behind the 10s, swim every day 1～2 time, continued for 3～4 weeks.
5. according to the preparation method of the described qi-deficiency fever animal model of claim 4, it is characterized in that the water temperature when described rat power exhausts swimming is 18～25 ℃.
6. according to the preparation method of the described qi-deficiency fever animal model of claim 2, the amount that it is characterized in that described lumbar injection LPS is 60～100 μ g/kg.
7. according to the preparation method of the described qi-deficiency fever animal model of claim 6, the amount that it is characterized in that described lumbar injection LPS is 80 μ g/kg.
8. the application of the described qi-deficiency fever animal model of claim 1 is characterized in that described qi-deficiency fever animal model provides platform for the research of Chinese traditional treatment fever due to deficiency of vital energy illness.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108553473A (en) * | 2018-07-17 | 2018-09-21 | 中国药科大学 | A kind of method for building up of liver-kidney deficiency card animal model |
| CN109006689A (en) * | 2018-09-13 | 2018-12-18 | 中国辐射防护研究院 | A kind of method for building up of the rat animal model of physical fatigue |
-
2010
- 2010-05-21 CN CN201010186415A patent/CN101849526A/en active Pending
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108553473A (en) * | 2018-07-17 | 2018-09-21 | 中国药科大学 | A kind of method for building up of liver-kidney deficiency card animal model |
| CN109006689A (en) * | 2018-09-13 | 2018-12-18 | 中国辐射防护研究院 | A kind of method for building up of the rat animal model of physical fatigue |
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Application publication date: 20101006 |