CN1565582A - Chinese medicine formulation with functions of resisting senescence and preventing and treating posteoporosis and cerebral lesion - Google Patents

Abstract

The invention discloses an anti-senescence compound preparation which has the functions of preventing sclerotin loosening and brain damage caused by couverture hormones long term physiological dosage application. The preparation comprises red sage root, epimeddium, astragalus root, and bighead atractylodes rhizome.

Description

Have defying age, prevent and treat osteoporosis and encephaloclastic Chinese medicine preparation

The present invention relates to one group and a kind ofly had defying age, prevented and treated osteoporosis and encephaloclastic Chinese medicine preparation compound preparation and method for making thereof by what salviamiltiorrhizabung, Herba Epimedii, the Radix Astragali and Rhizoma Atractylodis Macrocephalae extract were formed, osteoporosis and cerebral lesion due to this compound preparation is had children outside the state plan after the reason dose application to glucocorticoid for a long time have the good preventing effect.

Glucocorticoid (glucocorticoid, GC) be the steroid hormone that has neural activity by the excretory class of adrenal cortex, its generation directly is subjected to hypothalmus-pituitary-adrenal axis (hypothalamic-pituitary-adrenocortical axis, HPAA) influence also can be carried out negative feedback inhibition to HPAA.Glucocorticoid is a class important substance of keeping the body normal activities, and extensively and complicated, too much or very few all can bringing to body seriously influences in its pharmacological action.Mainly influenced the metabolic process of basic living matter under normal condition by the glucocorticoid of the secreted physiological dose of adrenal cortex, the glucocorticoid of excusing from death reason dosage then also has effects such as antiinflammatory, anti-immunity, shock.Therefore, glucocorticoid is used very extensive clinically, as being used as the insufficient long-term alternative medicine of endocrine, treatment severe infections or inflammation, treatment autoimmunity or anaphylactic disease and hematopathy etc., evident in efficacy, but long-term heavy dose of use can cause a series of serious adverse effects, as hemo-hyperadrenocorticism syndrome, secondary infection, ulcer disease, cardiovascular disease, malnutrition, intellectual deterioration psychological problem even brain atrophy dementia, osteoporosis etc.Especially the generation of brain atrophy, psychosis and osteoporosis often is difficult to be aware, in case it is big hazardness to occur, has therefore become one of drug withdrawal indication of glucocorticoid treatment.Modern study shows, excusing from death reason dosage glucocorticoid can be to the central nervous system, especially hippocampal neuron produces toxic action.Hippocampus be among the central nervous system with the closely-related important brain of learning and memory function district, also be to be subject to the target organ that glucocorticoid is attacked, the long-term excusing from death reason dosage glucocorticoid of accepting, the capital makes the neuron generation atrophy even the death in rat hippocampus brain district, simultaneously also with the decline of animal learning memory ability.Find also that clinically also there are certain cause effect relation in some psychotic disorders such as Cushing disease, alzheimer disease, depression and anxiety neurosis etc. with the excessive secretion of glucocorticoid.In the central nervous system, GC mainly be by and glucocorticoid receptor (GR) (glucocorticoid receptors, GR) and mineralcorticoid receptor (mineralocorticoid receptors is MR) in conjunction with playing a role.Hippocampus is and the closely-related position of cognitive behavior function, owing to be rich in adrenal cortical hormone receptor, and be the brain district that content is the highest among the central nervous system, therefore, when glucocorticoid concentration increased in the body, Hippocampus will be subjected to optionally attacking, thereby caused the infringement of maincenter.Glucocorticoid plays an important role in the rescue process of acute, seriousness disease, is the prerequisite of saving patient's life, but long-term heavy dose of glucocorticoid treatment that uses is especially outstanding to encephaloclastic effect.In nearest SARS epidemic, have to be reported among the SARS recovery from illness patient who accepts the steroid HAART, the early indication of brain and nerve injury has appearred in half.On the other hand, have report to show, in the patient who accepts the glucocorticoid chronic treatment, though any age 30%～50% the concurrent osteoporosis of probability is all arranged.The generation of this type of osteoporosis may cause CaE to increase with the glucocorticoid prolonged application and absorb reduce and glucocorticoid significantly to be suppressed to bone cell function relevant, it mainly shows as the bone amount and significantly reduces, the osseous tissue fine structure is degenerated, so that the fragility of bone increases, spontaneous fracture or bland necrosis often take place in the patient, bring great burden for patient family and society.Therefore, discussion glucocorticoid prolonged application produces the possible cause of maincenter toxicity and osteoporosis and develops the antagonism medicine has become the problem that presses for solution now.

For the glucocorticoid osteoporosis, have report to treat, but because estrogen has significant side effects, life-time service can increase the sickness rate of carcinoma of endometrium and breast carcinoma, so limited its use clinically with suppressing bone resorption medicine such as estrogen; Also occur clinically now many new drugs as: diphosphonic acid salt, calcitonin class and estrogenic agents etc. are also to preventing that bone loss from having certain effect, but life-time service suppresses bone formation on the contrary, cause that bone is aging, bone mass descends; At present, according to the characteristics of glucocorticoid osteoporosis, many clinically opinions are used calcium preparation, Vit D ₃With the assimilation parahormone it is treated, so the class medicine belongs to and promote bone formation, short bone mineralising medicine, main effect is to make 1,25 (OH) ₂D ₃Concentration maintain normal level, can regulate the factor of influence in the bone environment better, promote bone formation and suppress bone resorption, reach the purpose that increases the bone amount.But clinical research has also been reported life-time service assimilation parahormone and can have been produced tangible hepatotoxicity and water-sodium retention, and its absorption is irregular, so limited its use clinically, therefore, to the treatment of glucocorticoid osteoporosis, be with promoting bone formation, bone mineralising medicine or coming effect all undesirable with inhibition bone resorption medicine.On the other hand, for the brain damage that glucocorticoid osteoporosis prolonged application causes, now do not see yet that active drug occurs, so, generally can only adopt symptomatic treatment, but curative effect is unsatisfactory to occurring the patient of glucocorticoid brain injury symptom clinically.Therefore, task of the present invention is patient's prevention of the long-term heavy dose of use glucocorticoid of a kind of suitable needs of development and treats encephaloclastic compound preparation that this compound preparation is made up of Radix Salviae Miltiorrhizae, Herba Epimedii, the Radix Astragali and Rhizoma Atractylodis Macrocephalae extract.Studies show that this compound preparation can effectively improve the intelligence of prednisone rat, damage and the content of reduction and old and feeble relevant enzyme and the bone amount that effectively improves rat of the hippocampal neuron that effectively prevents and treats prednisone and caused.Therefore, can be used for improving the generation of body memory ability and slow down aging, especially can effectively prevent brain injury and osteoporosis after the dose application glucocorticoid is managed in long-term excusing from death.So needing can be used for the patient of prolonged application glucocorticoid treatment to make preventive usage, to reduce or to slow down the generation of glucocorticoid untoward reaction.

The approach that solves this task is to realize like this, get salviamiltiorrhizabung, Herba Epimedii, the Radix Astragali and the Rhizoma Atractylodis Macrocephalae, be made into tablet, electuary or capsule by following technology, for clinical practice, studies show that, this preparation has good preventive and therapeutic effect to prednisone rat experimental cerebral lesion and osteoporosis.This preparation can improve the intelligence of laboratory animal and effectively protect hippocampal neuron, reduces the generation that causes the content of old and feeble enzyme and alleviate the untoward reaction of other glucocorticoids in the brain homogenate.There is no report with this compound preparation treatment glucocorticoid cerebral lesion domestic and foreign literature.

This compound preparation can prepare by following SOME METHODS:

1, Radix Salviae Miltiorrhizae, Herba Epimedii, the Radix Astragali and the Rhizoma Atractylodis Macrocephalae are ground into powder, add 75～95% soak with ethanol 24 hours, filter, reclaim ethanol, it is standby to get ethanol extract, and medicinal residues decoct with water 3 times again, merge three times filtrate, filter, filtrate is concentrated in right amount, add aforesaid ethanol extract, cold drying is pulverized, add appropriate amount of auxiliary materials, make granule, tabletting, sugar coating or encapsulated, promptly.

2, Radix Salviae Miltiorrhizae, Herba Epimedii, the Radix Astragali and Rhizoma Atractylodis Macrocephalae raw medicinal herbs decoct with several times water logging bubble back, repeat 3 times, and merging filtrate concentrates, and adds 3 times of ethanol precipitations, behind the recovery ethanol, use ethyl acetate extraction, reclaim ethyl acetate and obtain the Chinese medicine total extract.The Chinese medicine total extract adds appropriate amount of auxiliary materials, makes granule, tabletting, and sugar coating or encapsulated, promptly.

The goods that adopt above 1 or 2 methods to make, prove through pharmacology, toxicologic study, this product to glucocorticoid have children outside the state plan for a long time the reason dose application after due to osteoporosis and cerebral lesion tangible prevention and therapeutical effect are arranged, acute toxicity test and long term toxicity test (comprising 3 months to 6 months) confirm that all this product toxicity is low, take for a long time all harmless effects of each vitals such as the heart, liver, kidney, lung, stomach, intestinal, spleen, thymus, lymph nodes.

For observing the pharmacodynamic action of forming compound preparation by Radix Salviae Miltiorrhizae, Herba Epimedii, the Radix Astragali and the Rhizoma Atractylodis Macrocephalae, carried out following experiment:

1 materials and methods

1.1 material

1.1.1 medicine and reagent prednisolone acetate: raw material, fairy house Pharmacy stock Co., Ltd produces.Radix Salviae Miltiorrhizae (Hebei), the Radix Astragali (Inner Mongol), Herba Epimedii (Jiangsu), the Rhizoma Atractylodis Macrocephalae (Zhejiang) four medicines are all purchased in Guangzhou medical material company, recognize definite through the operation pharmacist of Chinese medicine.Monoamine oxidase, MAO testing cassete, malonaldehyde mensuration test kit, acetylcholinesterase mensuration test kit, superoxide dismutase testing cassete, Coomassie brilliant blue protein determination kit build up bio-engineering corporation available from Nanjing and produce.

1.1.2 experiment Chinese medicine extract: get the technology extraction preparation that salviamiltiorrhizabung, Herba Epimedii, the Radix Astragali and the Rhizoma Atractylodis Macrocephalae are introduced by this patent.

1.1.3 instrument low speed saw (Buehler LTD.USA); Sclerous tissues's microtome (German Leica 2155); The firm cutter of tungsten carbide (German Leica company product); Osseous tissue morphometry Survey Software (KSSScientific Consultants, UT USA); Automated image digital assay instrument comprises light microscopic and fluorescence microscope (Nikon, Japan); UV-3010 ultraviolet spectrophotometer (day island proper Tianjin); AE240 electronic balance (Mettler-Toledo instr.shanghai Ltd); Cut tissue refiner in the ZS83-I type (the new machinery plant in Zhejiang makes); MG-2 type labyrinth stimulator (Zhangjagang City's Biomedical Instruments factory); ASD-3D Aiwa electronic weighing scale (Aiwa of Shenzhen weighing apparatus company limited product).

2.4 experimental technique

1.2 material

1.2.1 4 monthly ages, 40 of the Sprague-Dawley of copulation (SD) male rats are not selected in animal model and grouping for use, the SPF level, and average weight 228 ± 10g is provided by Guangdong Medical College's Experimental Animal Center.

The raising condition: raise indoor temperature and remain on 24 ℃～26 ℃, 50%～60% (being provided by the court's Experimental Animal Center) is provided humidity, and temperature and humidity condition is suitable, the next day change bedding and padding, special chamber sub-cage rearing, the special messenger is responsible for; Standard feed and the pure filtered water of use after strict sterilization fed (calcic 1.33%, phosphorus 0.95% are provided by the court's Experimental Animal Center), all laboratory animal sub-cage rearings, and guaranteeing has enough activity spaces, can freely ingest, take the photograph water.

The macroprogram Analys32.x11 that 48 SD male rats are used microsoft Excel band by body weight produces random number series, and utilizes the random number series that produces to carry out random packet, is divided into 5 groups altogether, 8 every group, is specially:

(1) NS group: the normal control group, give 0.9% sodium chloride solution (NS) every day and press 5ml/kg filling hello administration, repeat behind the 1h to give NS by the 5ml/kg filling.

(2) GC group: the prednisone model group, give prednisolone acetate 2.7mg/kg every day earlier and irritate stomach (irritate stomach with method to prednisone earlier for 3～6 groups, then give the relative medicine treatment), irritate behind the 1h and give NS 5ml/kg.

(3) CDH group: high dose compound Salviae Miltiorrhizae treatment group, irritate compound red sage root preparation 10g/kg/d (each 2.5g/kg/d of the Herba Epimedii+Radix Astragali+Rhizoma Atractylodis Macrocephalae+Radix Salviae Miltiorrhizae), 5ml/kg after giving prednisolone acetate 1h.

(4) CDM group: middle dosage compound Salviae Miltiorrhizae treatment group, irritate compound red sage root preparation 5g/kg/d (each 1.25g/kg/d of the Herba Epimedii+Radix Astragali+Rhizoma Atractylodis Macrocephalae+Radix Salviae Miltiorrhizae), 5ml/kg after giving prednisolone acetate 1h.

(5) CDL group: low dosage compound Salviae Miltiorrhizae treatment group, irritate compound red sage root preparation 2.5g/kg/d (each 0.625g/kg/d of the Herba Epimedii+Radix Astragali+Rhizoma Atractylodis Macrocephalae+Radix Salviae Miltiorrhizae), 5ml/kg after giving prednisolone acetate 1h.All animal grouping backs were observed one day, second day beginning gastric infusion, qd, continuous 12 weeks.

1.2.2 memory ability assay method

Learning memory is important Fundamentals in the intelligence, in the basic research often to the height of learning memory as the medicine nootropics or cause one of silly important indicator.Usually judge the influence of medicine in the pharmacology pharmacodynamic test to remembering with passive withdrawal reflex of animal and acquisition and the reproduction of initiatively escaping ability.Maze method is the detection animal memory method of using always just, by adopting electric labyrinth stimulus method to detect rat the memory ability in labyrinth is just judged the influence of life-time service glucocorticoid to rat intelligence in this experiment.Experiment finishes preceding 1 week, allows rat escape automatically in the stimulator of MG-2 type three arms electricity labyrinth earlier, the custom direction of observing its escape, and itemized record it, if zero difference after testing, then need not stipulate secure side to; If rat selects to the direction of escaping that there were significant differences, as common to the left, then be decided to be electric shock district (as the II district) with the left side to escape, the right side is place of safety (as the III district); When experiment formally begins, the I district is the starting area, wherein I district and II district are charged, and the III district is not charged, first animal training was set up mnemonic trace in the 1st day, every rat runs 20 times earlier and (is divided into 5 circulations, in order to avoid electricity swashs overlong time animal is damaged, stop at once and rat is taken out in case corona phenomenon occurs, single cage keeps flat, close observation) writes down its accuracy, repeat behind the 24h, but this time delayed time 5 seconds earlier before the electric shock, point out to red light simultaneously, observe rat and be excited response situation, repeat the correct number of times (ET of rat in the record experiment behind the 72h again, times of the exactitude) and errors number (MT, timesof the mistake), divided by total testing time, calculate labyrinth accuracy ER[ER=ET/ (ET+MT) with ET].

1.2.3 brain tissue slice sample preparations method:

Take off rat head under the freezing state, stenosis is taken out olfactory bulb between the promptly square at the moment foremost socket of the eye of brain, take out pinus at the lambdoid suture intersection, between the leading edge sutura of the inboard horizontal ridge of frontal bone and interparietal bone, open skull then and take out complete brain, the big brain weight of weighing at first, and observe outward appearance and have atypism to change, then with the right side brain with the liquid-solid routine paraffin wax embedded section that carries out surely of Podbielniak.Wait the rat brain stereotaxic atlas to repair piece of tissue according to the bag new people at distance bregma caudal ward 3mm to 4mm place, the crown section of row, open at the about 3.5mm of the distance bregma caudal ward serial section 5-8 of place, thick 4 μ m, after HE dyeing, calculate unit are neuron number (giant pyramidal cells counting) and measurement rat brain striate cortex 17 districts and 18 interval molecular layers thick and cortex gross thickness in Hippocampus CA2 and CA3 district intersection 362.5 mu m ranges respectively at tissues observed structural change under the high power light microscopic and with the semi-automatic image analyser.Its concrete assay method is as follows:

(1) Hippocampus unit are giant pyramidal cells number (HN) measuring method: Hippocampus is the important structure of limbic system, structure is more regular, and in memory, play extremely crucial effect, become at present a hot topic of Neuroscience Research, wherein the morphological change of Hippocampus in ageing process caused that academia extensively payes attention to.In this experiment, under 10 * 40 times of mirrors, observe the hippocampal cell morphosis by the semi-automatic image analyser earlier, and measuring and calculating Hippocampus CA2 and the interior neuron number of CA3 district intersection 362.5 mu m ranges (mainly being the giant pyramidal cells counting) and this district's area, and calculate neurocyte number in this district's unit are, to estimate the index of glucocorticoid prolonged application to rat hippocampus structure influence degree.

(2) ratio (M/C) assay method of rat brain molecular layers thick and cortex gross thickness:

The mammal neopallium generally is divided into six layers by the surface to white matter, it is respectively I: molecular layer (molecular layer), II: external granular layer (internal granular layer), III: external pyramidal layer (pyramidal layer), IV: internal granular layer (internal granular layer), V: internal pyramidal layer (internal pyramidal layer) and VI: multiform layer (polymorphic layer), striate cortex 17 districts and 18 districts belong to neopallium, structure is more clear, and in half brain section with the hippocampal formation close together, therefore identification easily selects to measure this district's cortex gross thickness and molecular layers thick as parameter index.Confirm and the observation cortex construction with the semi-automatic image analyser is preliminary under 10 * 40 times of mirrors earlier, under 10 * 20 times of mirrors, be determined at thickness (the wall of molecular layer of molecular layer in the same plane more respectively, MW) and cortex gross thickness (wall of cortex, CW), and with its ratio M/C as estimating the index of glucocorticoid to rat brain cortex structure influence degree.

1.2.4 brain homogenate biochemical indicator assay method

After the rat right ventricle is got blood, break end rapidly, on the ice platform, take out complete brain immediately, left side, the back brain of weighing prestores prior to-20 ℃, 10% brain homogenate is made in 0.86% saline grinding with 4 ℃ on the inherent ice bath of 48h, the cut tissue refiner is fully pulverized in the ZS83-I type again, and the centrifuging and taking supernatant is measured MAO, SOD, AchE, MDA and brain homogenate protein content.

1.2.5 the making of bone specimen

Rat is put to death with the heart blood drawing of pentobarbital sodium anesthesia back, take out the left side tibia, open pulp cavity with sawing at near-end at a slow speed as sagittal, 1/3 place walks crosswise sawed-off in the tibia epimere, get that the tibia epimere is fixed, dehydration, embedding and section, downcut 5 μ m and 8 μ m section respectively with sclerous tissues's microtome, 5 μ m bone thin slices adopt Masson-Goldner Trichrome colouring method, and the dyeing back is in order to observe bone trabecula surface osteoclast number.The direct mounting of 8 μ m bone sheets adopts semi-automatic image digitization analyser to measure osseous tissue morphometry static parameter and dynamic parameter.

1.2.6 proximal tibia cancellous bone tissue morphometry is measured

With KSS Image image analysis system,, measure the static parameter and the dynamic parameter of proximal tibia by the definition and the basis of calculation of international bone morphometry parameter measurement.Static parameter comprises the osseous tissue gross area (T.Ar, mm ²), bone trabecula area (Tb.Ar, mm ²), bone trabecula girth (Tb.Pm, mm) and calculating parameter [the bone trabecula area percent (%Tb.Ar, %), bone trabecula thickness (Tb.Th, μ m) and bone trabecula separating degree (Tb.Sp, μ m)], total osteoclast number (Oc.No) and index thereof (Oc.N/mm, #/mm).Dynamic parameter comprises single labelling girth (sL.Pm, mm), double labelling girth (dL.Pm, mm) and double labelling spacing (Ir.L.Wi, μ m) and calculating parameter [labelling girth percent (%L.Pm, %), bone mineralising deposition (MAR, μ m/d), bone formation rate BER/BS (expression bone surface formation rate, %yr), bone formation rate BER/BV (expression bone degree of conversion, %yr), bone formation rate BER/TV (expression osseous tissue formation rate, %yr)].

1.2.7 statistical method

All testing indexs all adopt the form of mean plus-minus standard deviation to represent (Mean ± SD), data statistics uses the variance analysis (ANOVA) in the SPSS software to test, earlier use Levene ' s test that data are carried out homogeneity test of variance, check with Bonferroni together as variance, then use Tamhane ' s T2 check as heterogeneity of variance, with p＜0.05, p＜0.01 expression has statistical significance.

2 experimental results

2.1 the prednisone prolonged application is to the influence of central nervous system in rat biochemical indicator

The prednisone prolonged application sees Table 2.1 to MAO (monoamine oxidase, MAO), SOD (superoxide dismutase), AchE (acetylcholinesterase), MDA (malonaldehyde) content influence in rat brain weight (CW) and the brain homogenate:

Table 2.1 prednisone prolonged application to the influence of rat cerebral even slurry biochemical indicator (x ± s, n=8)

Tab?2.1?Changes?of?chemistry?parameters?of?cerebrum?plasm?in?prednisone?rats( x±s，n＝8)

CW MAO SOD AChE MDA

Group

(g) (U/h/mgprot) (U/h/mgprot) (umol/ml) (nmol/mgprot)

NS 1.905±0.08 9.57±3.06 29.29±1.87 0.53±0.02 2.97±0.22

GC 1.874±0.05 22.12±1.11 ^** 18.53±2.95 ^** 0.36±0.05 ^** 2.49±0.51

CDH 1.959±0.133 19.06±3.06 ^* 25.28±3.97 ^△ 0.67±0.12 ^△△* 2.17±0.28 ^△

CDM 1.953±0.104 17.20±2.13 ^△* 26.77±4.11 ^△ 0.61±0.07 ^△△* 2.40±0.26

CDL 1.924±0.133 12.39±2.45 ^△△ 24.64±2.63 ^△ 0.85±0.08 ^△△* 2.77±0.31

Compared?with?NS ^*：P＜0.05， ^**：P＜0.01；Compared?with?GC △：P＜0.05，△△：P＜0.01。

Compare (table 2.1) with the NS group, GC group rat brain weight has downward trend, and enzyme SOD relevant with aging in the brain homogenate obviously descends, and the content of MAO significantly raise (P＜0.01), prompting life-time service prednisone can make the rat nervous system functional activity be restricted, and old and feeble sign occurs.GC group AchE content descends and MDA changes of contents little (P＞0.05), and prompting GC group rat may take place that cholinergic nerve function rate is moved back and the reduction that causes AchE content; With GC group relatively, CDL significantly increased SOD and AchE content and reduce the content (P＜0.01) of MAO, the central untoward reaction of pointing out CD can resist prednisone to a certain extent.

2.2 the prednisone prolonged application detects the influence of accuracy to rat three arm labyrinths

Compare (table 2.2) with the NS group, GC group activities in rats obviously reduces, its respond that stimulates to external world significantly descends, in 60 race labyrinth test experiments, correct (ET) only arranged 32 times, (ER ET/60*100%) only has 53.8% to accuracy, and the accuracy of NS group reaches 71.5%, the ER of prompting GC group obviously descends (P＜0.01) than the NS group, and prompting life-time service prednisone can make rat that hypomnesis takes place.Though CDH group rat labyrinth accuracy descends (P＜0.05) than the NS group, improves (P＜0.05) than the GC group, prompting CD can effectively improve rat brain memory ability.The prednisone prolonged application sees Table 2.2 to the influence of rat correct number of times in three arm labyrinths (ET) and accuracy (ER).

Table 2.2 prednisone prolonged application to the influence of rat labyrinth accuracy (x ± s, n=8)

Tab2.2?Changes?of?memory?ability?in?prednisone?rats( x±s，n＝8)

Group ET ER(％)

NS 42.9±4.5 71.5±7.6

GC 32.3±5.1 ^** 53.8±8.5 ^**

CDH 37.1±2.3 ^*△ 61.9±3.8 ^*△

CDM 36±2.7 ^** 60±4.6 ^**

CDL 36.2±3.5 ^** 60.3±5.9 ^**

Compared?with?NS ^*：P＜0.05， ^**：P＜0.01；Compared?with?GC?△：p＜0.05，△△：P＜0.01。

2.3 the prednisone prolonged application is to rat cerebral tissue's structure influence

The prolonged application prednisone sees Table 2.3 to the influence of the ratio (M/C) of the gross thickness of rat hippocampus unit are giant pyramidal cells number (HN), cerebral cortex thickness (CW) and molecular layers thick and cortex.

Table 2.3 prolonged application prednisone to rat cerebral tissue's structure influence (x ± s, n=8)

Tab2.3?Changes?of?structure?of?cerebrum?in?prednisone?rats( x±s，n＝8)

HN CW M/C

Group

(N/mm ²) (mm) (mm/mm)

NS 1901±378 1.244±0.11 0.111±0.03

GC 1737±533 1.198±0.19 0.124±0.01

CDH 1927±558 1.290±0.10 0.105±0.02

CDM 1996±270 1.265±0.17 0.099±0.02△

CDL 1874±369 1.372±0.21 0.098±0.01△

Compared?with?NS ^*：P＜0.05， ^**：P＜0.01；Compared?with?GC?△：P＜0.05，△△：P＜0.01。

Each organizes the rat brain outward appearance does not have significant difference, is tip wedge forward, and smooth surface lacks ditch returns, and belongs to flat brain type (lissencephalic brain), does not see tangible swelling, atrophy or change color.Rat striate cortex 17 districts and increase trend is arranged 18 interval molecular layers thick and cortex gross thickness (CW) has minimizing trend behind the visible prolonged application prednisone under the light microscopic; Though the cortical cell layering is still clear, the cell arrangement disorder, obvious edema takes place in the part cell, as seen has cell that obvious degeneration necrosis takes place.On the other hand, visible NS group hippocampal neuron distribution rule under the light microscopic, size is consistent, and rat hippocampus district neuron is arranged dispersion behind the life-time service prednisone, cell space swelling or shrinkage, wherein CA2 and CA3 intersection have more neurocyte generation karyopycnosis even dissolving death.Organize than (table 2.3) with NS, the gross thickness ratio (M/C) of rat molecular layers thick and cortex rises behind the prolonged application prednisone, unit are giant pyramidal cells number (HN) has downward trend in Hippocampus CA2 and the CA3 district intersection 362.5um length range, and the long-term excusing from death reason of prompting dosage uses prednisone can cause the Hippocampal Neurons damage.And the M/C of two groups of CDM and CDL significantly descends (P＜0.05) than the GC group, and three groups of HN of CD are all on the rise, and are wherein obvious with CDM, and the neural cell injury that prompting CD causes prednisone has certain preventive effect.

2.4 the prednisone prolonged application influences rat tibia epimere spongy bone morphometry

The prednisone prolonged application sees Table 2.4 to the influence of static parameter %Tb.Ar in the rat bone morphometry (bone trabecula area percent), Tb.N (bone trabecula quantity), Tb.Th (bone trabecula thickness), Tb.SP (bone trabecula separating degree) and bone resorption marker Oc.N/BV (unit bone trabecula girth amount of osteoclast), %Oc.Pm (bone resorption girth percent):

Table 2.4 prednisone to the influence of rat bone morphometry static parameter (x ± s, n=8)

Tab?2.4?Changes?of?the?static?histomorphometry?parmeters?in?prednisone?rats( x±s，n＝8)

Tb.Ar TbN TbTh TbSp Oc.N/BV ％Oc.Pm

Group (％) (mm ^-1） (um) (um) (mm ^-2) (％)

NS 15.07±1.70 2.92±0.23 51.86±6.48 292.88±25.52 11.42±1.46 0.873±0.09

GC 8.83±0.98 ^** 1.98±0.20 ^**?44.47±2.43 ^* 463.53±54.54 ^** 38.38±8.65 ^** 254±0.34 ^**

N-％ -41.3 -31.9 -14.2 58.3 236.1 191.4

CS 13.93±2.48 ^△△2.83±0.50 ^△△49.02±1.92 ^△△?311.03±56.31 ^△△?16.14±3.33 ^*△△ 1.15±0.26 ^*△△

G-％ 57.7 42.6 10.3 -32.9 -57.9 -54.7

CDH 14.14±2.80 ^△△2.82±0.50 ^△△50.08±4.52 ^△△?312.55±55.14 ^△△?15.35±5.37 ^*△△ 1.27±0.46 ^*△△

G-％ 60.1 42.1 12.6 -32.6 -60 -50.3

CDM 14.00±2.93 ^△△2.75±0.56 ^△ 50.96±5.33 ^△ 326.14±76.62 ^△△?18.49±4.42 ^*△△ 1.46±0.32 ^*△△

G-％ 58.1 35.5 14.6 -29.6 -51.8 -42.7

CDL 13.65±2.43 ^△△2.88±0.51 ^△ 47.57±4.53 309.87±67.98 ^△△?16.21±4.23 ^*△△ 1.28±0.26 ^*△△

G-％ 54.4 44.9 7 -33.2 -57.8 -49.7

Compared?with?NS? ^*：P＜0.05， ^**：P＜0.01；Compared?with?GC?△：P＜0.05，△△：P＜0.01。

Compare (table 2.4) with the NS group, %Tb.Ar, Tb.N, Tb.Th all descend (P＜0.01) in the GC group rat bone static parameter, and prompting GC group rat spongy bone has obviously to be lost; And GC organizes Tb.SP, Oc.N, %Or.Pm obviously increases (P＜0.01), points out this group osteoclast to increase, and bone resorption increases.With GC group relatively, three groups of %Tb.Ar, Tb.N and Tb.Th that also can significantly improve the prednisone rat of CD, and reduce Tb.SP, Oc.N, %Oc.Pm, remarkably productive with CDH.

2.5 the prednisone prolonged application is to the influence of rat spongy bone morphometry dynamic parameter

The prednisone prolonged application sees Table 2.5 to the influence of rat spongy bone morphometry dynamic parameter %L.Pm (fluorescent perimeter percent), MAR (bone mineralising deposition) and three kinds of bone formation rate index BFR/BS, BFR/BV and BFR/TV:

Table 2.5 prednisone to the influence of rat spongy bone morphometry dynamic parameter (x ± s, n=8)

Tab?2.5?Changes?of?dynamic?histomorphometry?parameters?in?prednisone?rats( x±s，n＝8)

Group ％L.Pm(％) MAR(um/d) BFR/BS(％/year) BFR/BV(％/year) BFR/TV(％/year)

NS 31.88±8.19 0.85±0.16 10.28±3.98 336.6±140.8 49.3±17.7

GC 29.81±9.02 0.81±0.25 8.91±3.97 332.8±142.5 29.6±13.2 ^*

CS 31.67±5.41 0.92±0.11 10.81±2.85 366.1±89.0 51.3±15.6 ^△

CDH 31.34±0.76 0.90±0.19 10.68±5.17 351.4±146.7 49.5±24.5 ^△

CDM 30.78±11.3 0.85±0.29 10.00±4.67 322.3±146.6 43.3±19.7 ^△

CDL 28.82±8.78 0.85±0.16 9.31±4.81 326.3±162.5 41.7±12.0 ^△

Compared?with?NS ^*：P＜0.05， ^**：P＜0.01；Compared?with?GC △：P＜0.05，△△：P＜0.01。

Compare (table 2.5) with the NS group, the variation little (P＞0.05) of %L.Pm and MAR, BFR/BS, BFR/BV in the GC group rat bone dynamic parameter, but BFR/TV obviously reduces (P＜0.05), and prompting excusing from death reason for a long time dosage uses prednisone can make the rat Grafting Cancellous Bone Bolt form the total amount minimizing; With GC group ratio, the bone mineralising deposition and the bone formation rate of three groups of rat spongy bone of CD are all on the rise, wherein raise the most obvious with BFR/TV, CDH, CDM and CDL organize 67%, 46% and 41% (P＜0.05) of having risen than GC respectively for three groups, compare zero difference (P＞0.05) with CS, prompting CD can significantly promote rat bone to form.

3 discuss

3.1 having children outside the state plan the reason dose application for a long time, glucocorticoid can cause maincenter toxicity clinical data to show to rat, the glucocorticoid level is far above adult and adolescence in the old human blood, therefore the patient of life-time service glucocorticoid can think that glucocorticoid and brain aging and central nervous system's degeneration is in close relations often with memory dysfunction.The prednisone that gives rat excusing from death reason dosage in this experiment is irritated to feed and is observed the rat hypomnesis after 3 months, cerebral cortex changes with hippocampal formation and the enzyme content relevant with aging changes, with observed phenomenon is consistent clinically, the prompting glucocorticoid is to neural people and the rat exhibit comparable on one's body that acts on.Prednisone causes central nervous system's toxicity concrete manifestation as follows to rat after having children outside the state plan the reason dose application for a long time:

3.1.1 having children outside the state plan the reason dose application for a long time, glucocorticoid can make rat three arms electricity labyrinth accuracy (ER) obviously descend GC group rat three arms electricity labyrinth accuracy than NS group (P＜0.01) (the seeing Table 2.2) that obviously descend.In general, intelligence test should comprise the test of several respects such as learning memory, thinking ability, judgment, faculty of understanding and creativity, they all lead between mutually and and influence, be difficult for absolutely separating, learning memory is one of them important Fundamentals, therefore more to its research, method is also more.Three arms electricity maze method is the test animal learning memory method of using always just, this method mainly is to judge the influence of medicine to learning memory by observing the passive and active withdrawal reflex capacitation and the reproduction that are produced when animal is faced danger, giving prompting and do not giving in the test of prompting, if medication animal accuracy obviously improves than non-medication animal, then point out medicine to have significant nootropic effect, vice versa.Rat three arm labyrinth accuracy significantly descend after finding the life-time service prednisone in this experiment, and the prompting prednisone may suppress the central nervous system grows, and influences the Animal Intelligence development, causes intellectual deterioration even old and feeble occurring ahead of time.

Can make that the content of MAO significantly rises in the rat cerebral tissue 3.1.2 glucocorticoid is had children outside the state plan the reason dose application for a long time, the content of SOD and TchE descends obviously in the GC group rat cerebral even slurry that the content of MAO significantly rises and the content of SOD and TCHE obviously descends (P＜0.01) (seeing Table 2.1).This is three kinds of requisite enzyme in brain neurological motion, its in vivo balance whether can directly influence the body movable function decline whether, so relation is very close between these three kinds of enzymes and the organism aging.Mechanism about aging has a variety of sayings, and wherein free radical theory was at first proposed by D.Harman in 1956, and it thinks that the excessive meeting of the free radical that produces causes DNA and the damage of other macromole in metabolic processes.Free radical mainly is by causing that unsaturated lipids generation peroxidating causes body generation degeneration in the various biomembranes, even causes that organism aging death, this kind reaction mainly exist in the active histoorgan of function, as brain, the heart, liver, kidney etc.The mechanism of body antagonism radical damage mainly contains two kinds, and the one, to damaging macromolecular repair mechanism, the 2nd, eliminate free radical mechanism.In vivo, the most effective free radical scavenger is exactly SOD, and its content reduces prompting body endoperoxides significant reaction, and catabiosis appears in body.In the GC group, the SOD content in the rat cerebral even slurry significantly descends, and the long-term excusing from death reason of prompting dosage uses prednisone can obviously increase the weight of brain inner lipid peroxidating process, causes the free radical bulk deposition, has damaged the activity and the function of brain.On the other hand, the changes of Catecholamine Content in the MAO degradable brain reduces cerebration, so MAO and old and feeble in close relations.The increase with the age after 45 years old of MAO-B content in the human brain increases severely, so MAO is called aging relevant enzyme again, is considered to aged sign.MAO content in the GC group rat cerebral even slurry significantly rises, prompting life-time service glucocorticoid can quicken the degraded of catecholamine in the brain, cause 5-hydroxy tryptamine and metabolite 5-hydroxyindoleacetic acid content thereof to increase relatively, make the physiological activity imbalance of brain, cause old and feeble generation.Acetylcholine (Ach) is the most ancient mediator, and its major function is the chemical mediator as nerve conduction, and main deactivation mode in vivo is the effect by acetylcholinesterase (AchE).The content of AchE descends in the GC group rat cerebral even slurry, vigor reduces, can be observed GC group rat cerebral tissue structure disturbance simultaneously, neurocyte is badly damaged, therefore can infer the prednisone prolonged application cause in the brain homogenate AchE vigor descend may with the cholinergic nerve degeneration, cause Ach generate to reduce and cause due to the slump in demand (being the secondary reactions that Ach lacks) to AchE, this with study recently consistent.Bibliographical information is also arranged in the Alzheimer patient in addition, find cholinergic neuron generation selectivity degeneration in the main basal forebrain relevant with identification and memory function, AchE is active in cortex and the cerebrospinal fluid reduces; Other observes, and the AchE activity also is starkly lower than the normal person in the senile dementia patient frontal cortex, similar to the Alzheimer patient, therefore can infer, but prolonged application glucocorticoid injured neuron, thereby cause the synthetic and disorderly appearance in advance that causes the organism aging of release function of neurotransmitter in the brain.

3.1.3 glucocorticoid is had children outside the state plan the reason dose application for a long time rat hippocampus structure genetic morphology is changed, unit are neural pyramidal cell number in specific region falls now, obvious karyopycnosis necrotic brain tissues cut sections for microscopic examination take place and find in giant pyramidal cells, NS group rat hippocampus CA2 and CA3 district intersection neuron size distribution rule, cellularity and clear layer are the C shape and arrange.Compare (seeing Table 2.3) with the NS group, GC group rat hippocampus giant pyramidal cells quantity descends, and wherein the neural pyramidal cell number of unit are descends 9% in CA2 and the CA3 district intersection 362.5um length range; Hippocampal cell structure disturbance, arrangement disperse, cell space swelling or shrinkage, and as seen more glial cell appears in the visible karyopycnosis engrain of part, even visible neuronal necrosis in the zone of large pyramids.Generally speaking, neuronal cell karyopycnosis phenomenon is seldom seen, and neuronal cell nuclear generation pyknosis is the performance before the cell death, GC group rat hippocampus cell generation obvious variation prompting prednisone prolonged application can cause hippocampal neurons death or accelerator nerve unit to degenerate, and body aging, hypomnesis that prompting glucocorticoid prolonged application causes may be relevant with the apoptosis of hippocampal cell.Hippocampal formation is developed by archicortex (archipallium) on evolving, main neuron is a giant pyramidal cells, it is the significant points of limbic system, relevant with senior neural activities such as affective behavior, learning and memories, be again simultaneously to regulate the high-order maincenter of hypothalamo-pituitary-adrenal axis (hpa axis) and the main target area of stress damage.Studies show that, damage of irritability Hippocampus and hpa axis hyperfunctioning reciprocal causation relation, both have caused the vicious cycle of chronic stress Hippocampus damage, and its mechanism understands as yet, and the organelle degeneration that may cause with the glucocorticoid prolonged application is relevant with a large amount of consumption of energy.Observe in this experiment that the neuronal quantity of rat hippocampus CA2 and CA3 district intersection descends behind the life-time service prednisone, glial cell increases, the prompting prednisone is had children outside the state plan the reason dose application for a long time and can be caused rat hippocampus neurocyte composition and structure to change, and causes the neural activity ability drop relevant with intelligence.Hippocampus is the pith that participates in memory in the brain, its structure changes and points out memory function to descend, amnesia is that an old and feeble early symptom appears in organism, so the morphological change of Hippocampus in aging course caused the great attention of academia.The mechanism of bibliographical information glucocorticoid damage Hippocampus may consume with ATP in the accelerator nerve unit, disturbs due to the neurocyte metabolism; Also have the scholar to think that the hippocampal pyramidal cell infringement is the important pathologic basis of Alzheimer patient memory dysfunction, main cause is to have caused due to the cholinergic neuron degeneration; But this experimental result has also shown the respond of life-time service prednisone appreciable impact animal, causes the rat hypophrenia and old and feeble or even dull-witted occurring ahead of time.

3.1.4 having children outside the state plan the reason dose application for a long time, glucocorticoid can make rat layer cell arrangement disorder, part cell generation hydropic degeneration, visible NS group rat layer clear in structure under molecular layers thick and the skin thickness ratio rising light microscopic, the cell marshalling, and GC group rat layer structural stratification is still clear, but the cell arrangement disorder, pyramidal cell reduces.Compare with the NS group, GC group rat cerebral cortex thickness reduces 4%, and the ratio of the gross thickness of striate cortex 17 districts and 18 interval molecular layers thick and cortex rising 12% (seeing Table 2.3), the prompting glucocorticoid is had children outside the state plan the reason dose application for a long time can destroy and change the rat cerebral cortex structure.Generally speaking, the molecular layer neuron is less, mainly be horizontal cell, star-like cell and the nerve fiber surperficial parallel with cortex, atrophy may take place in the thickness that the ratio of this layer in the cortex gross thickness increases other cellular layer of prompting, especially external pyramidal layer that thickness is bigger and internal pyramidal layer, this is two-layer to be the main aggregate site of Gorky I type neuron, atrophy may take place in thickness attenuation prompting cortex pyramidal cell, the formed quantity of information unofficial biography in indication cortex zone weaken, and are the main causes of intellectual impairment.

3.2 having children outside the state plan the reason dose application for a long time, glucocorticoid can cause osteoporosis rat

3.2.1 glucocorticoid is had children outside the state plan the reason dose application for a long time rat bone girder area percentage rate is reduced, bone density obviously reduces, separating degree significantly increases in this experiment GC group rat bone morphometry static parameter and shows that GC group rat tibia epimere spongy bone obviously loses (P＜0.01) (seeing Table 2.4), compare with the NS group, bone trabecula area percent (%Tb.Ar) has reduced by 41.3% (P＜0.01), bone trabecula number (Tb.N) reduces by 31.9% (P＜0.01), bone trabecula thickness (Tb.Th) has reduced by 14.2% (P＜0.05), bone trabecula separating degree (Tb.SP) has increased by 58.3% (P＜0.01), illustrate that GC organizes the rat bone girder and obviously loses, density obviously reduces, and separating degree significantly increases; %Tb.Ar is the important indicator of reflection bone quantitative changeization, and Tb.N, Tb.Th and Tb.SP are the microstructural aggregative indicatores of reflection bone trabecula, more than changes to have shown GC group rat spongy bone structure generation significant change, presents typical osteoporosis state.

3.2.2 glucocorticoid is had children outside the state plan the reason dose application for a long time the rat osteoclast activity is increased, bone resorption is active in this experiment bone resorption parameter, GC group rat unit bone trabecula area osteoclast number (Oc.N/BV) increases by 236.1% (P＜0.01), bone resorption girth percentage rate (%Oc.Pm) increases by 191.4% (P＜0.01), the rat osteoclast activity increases behind the long-term excusing from death reason of the prompting dose application prednisone, bone resorption is active, quickened osteolysis, this effect may suppress the intestinal calcium absorption with glucocorticoid, increasing urine calcium output causes blood calcium to descend and reflexive causes that PTH (parathyroid hormone) secretion increases, and stimulates the osteoclast differentiation to enliven relevant.

3.2.3 glucocorticoid is had children outside the state plan the reason dose application for a long time the bone formation rate of rat unit osseous tissue area is reduced from bone morphometry dynamic parameter result observation (seeing Table 2.5), the dynamic indicator of reflection bone conversion comprises L.Pm% (fluorescent labeling girth percent), MAR (bone mineralising deposition), BFR/BS (bone formation rate on the unit length bone trabecula surface), BFR/BV (unit bone trabecula area bone formation rate), BFR/TV (unit osseous tissue area bone formation rate), these indexs all can reflect that osteoplastic vigor [48] BFR/BV then reflects the height of bone degree of conversion.GC group rat BFA/TV has reduced by 40% (P＜0.05) in this experiment, and the bone formation of prompting unit osseous tissue area reduces, and %L.Pm, MAR, BFR/BS, BFR/BV also have minimizing in various degree, but not statistically significant (P＞0.05).Bibliographical information uses prednisone 4.5mg/Kg, 3 days once, can significantly suppress %L.Pm, MAR, BFR/BS, BFR/BV and BFA/TV in the rat tibia bone formation index in continuous 2 months, the prednisone prolonged application does not have obvious inhibition to %L.Pm, MAR, BFR/BS and the BFR/BV of rat tibia in this experiment, may be different with medication relevant with the consumption of prednisone in the experiment, this phenomenon is worth further inquiring into.In this experiment, what adopt is the method for prednisone successive administration every day, totally 12 weeks, this usage can make the BFA/TV of GC group rat raise and amount of osteoclast increases, and may point out continuous use may cause that the PTH secretion increases and causes osteoclast activity to increase, feedback stimulating osteoblast activity, cause the osteoblast osteogenesis function to strengthen, promote the formation of bone, this may be to cause that other dynamic parameters change unconspicuous reason in this experiment, and it is still needed further discussion.

3.3 compound red sage root preparation composition principle and foundation

Herba Epimedii, the Radix Astragali, the Rhizoma Atractylodis Macrocephalae and Radix Salviae Miltiorrhizae all are the Chinese medicine with anti-aging effects, and osteoporosis is old and feeble a kind of important behaviour, and therefore four flavor Chinese medicines may have the effect of osteoporosis.This seminar once carried out the animal experiment study of osteoporosis disease to the preparation of being made by Herba Epimedii, the Radix Astragali, the Rhizoma Atractylodis Macrocephalae three flavor Chinese medicines, found that its each single or compound recipe all have drug resistance rerum natura osteoporosis or the osteoporotic effect of castration, its mechanism may be to increase the bone amount by strengthening the bone reconstruction.The bibliographical information Herba Epimedii can make the bone metabolism of body be in the positive balance state by approach such as protection gonad, inhibition bone resorption and promotion bone formation, experiment in vitro has proved that also Herba Epimedii has the osteoblastic proliferation of promotion and differentiation, can coordinate the balance of body intracellular metabolic by endocrine regulation; It is outstanding to study the effect of thinking Radix Astragali defying age, enhancing immunity and improving blood circulation protection hepatic and renal function, can not suppress the bone loss that bone formation is used for resisting castrated rats by suppressing bone resorption; The traditional Chinese medical science think the Rhizoma Atractylodis Macrocephalae have the spleen reinforcing stomach reinforcing and in effect, studies show that the Rhizoma Atractylodis Macrocephalae has and promote proliferation of bone marrow cells effect and the effect of IL-1 sample that stronger promote osteogenesis effect is arranged.The glucocorticoid prolonged application is except that can causing untoward reaction such as osteoporosis, immunosuppressant, cardiovascular disease and digestive system damage, toxic action to the central nervous system is outstanding, can cause some patient's Mental Subnormality behind the life-time service glucocorticoid that also appears in the newspapers clinically.Consider that glucocorticoid is obvious to the toxic action of maincenter, also in order to improve the antiinflammatory action of glucocorticoid, form compound Salviae Miltiorrhizae (CD) Chinese medicine preparation simultaneously so on original basis, add Radix Salviae Miltiorrhizae.Radix Salviae Miltiorrhizae has the Oesteoblast growth of promotion, improves the nervus centralis function, promotes immunity and antiinflammatory action, be desirably in the preparation of being formed, the calmness of Radix Salviae Miltiorrhizae is allayed excitement and promoted to remember can be to central nervous system's toxic action of Antiglucocorticoid, and the gonad inhibitory action that the gonadotropic Effect of Herba Epimedii can cause Antiglucocorticoid; The Radix Astragali improves circulatory function can be to the cardiovascular system infringement of Antiglucocorticoid; The Rhizoma Atractylodis Macrocephalae can react the digestive system that the Antiglucocorticoid prolonged application is caused.So the principles of formulating prescriptions of CD are in order to reduce the untoward reaction that the glucocorticoid prolonged application causes to greatest extent.

3.3.1 compound red sage root preparation can effectively prevent rats with osteoporosis

Experimental result prompting compound red sage root preparation can effectively prevent the bone loss of rat, wherein the effect with high dose is comparatively obvious, its mechanism of action may can obviously suppress the increase that rat bone that the prednisone prolonged application causes absorbs with the CD preparation, has certain facilitation relevant to bone formation simultaneously.The effect of this remarkable increase bone amount of compound red sage root preparation may be relevant with the combined effect of Radix Salviae Miltiorrhizae, the Radix Astragali, Herba Epimedii and four kinds of Chinese medicines of the Rhizoma Atractylodis Macrocephalae, bibliographical information 1. Radix Salviae Miltiorrhizae can improve the blood circulation of the inner and whole body of bone environment, the nutrition supply that helps bone, Radix Salviae Miltiorrhizae also can increase the content of Zn in serum and reduce the content of copper, can further increase the formation of spongy bone and has reduced copper to osteoplastic inhibitory action; 2. the Radix Astragali can promote collagen protein synthesis, promotes osteoblast secretion osteoid, the function of osteoblast inhibitory action that can effectively cause the Antiglucocorticoid prolonged application.3. Herba Epimedii is traditional kidney-nourishing yang-strengthening medicine, also has strong effect, clinical report, give the patients with osteoporosis made a definite diagnosis with the Herba Epimedii compound treatment after 3 months symptom obviously improve, also can obviously accelerate the healing of fracture patient callus and improve the arrangement and the density of callus, experiment in vitro has also pointed out Herba Epimedii water extract can suppress the generation of osteoclast and the formation that reduces bone resorption and promoted bone by the differentiation of stimulating osteoblast.4. the Rhizoma Atractylodis Macrocephalae is the Qi-tonifying drug of using always, and experimentation shows that the Rhizoma Atractylodis Macrocephalae can promote proliferation of bone marrow cells and have the effect of IL-1, can further promote osteoblastic conversion; Also there is report to gavage the Rhizoma Atractylodis Macrocephalae decocting liquid 6g/kg every day for mice, continuous 1 month, can significantly promote body weight gain of mice and muscle to strengthen, the strong effect of the Rhizoma Atractylodis Macrocephalae is obvious.Therefore, the CD preparation to the effect of prednisone rat tibia epimere spongy bone may be four kinds of Chinese medicines mutual coordination, mutually promote and have play a role separately due to.

3.3.2 compound red sage root preparation can effectively prevent rat maincenter toxic action

The CD preparation is observed in this research also has the part antagonism to other untoward reaction of rat that the prednisone prolonged application is caused, and has following several performance:

(1) compound red sage root preparation can effectively prevent glucocorticoid to have children outside the state plan the phenomenon experimental result that the rat three arms electricity labyrinth accuracy (ER) of reason due to the dose application descend for a long time to show, compound red sage root preparation can effectively be prevented and treated the brain function decline that the life-time service prednisone is caused, compare with the GC group, CDH, the ER of CDM and three groups of rats of CDL raises 15% respectively, 12% and 12%, obviously increase (P＞0.05) than the GC group, the prompting compound red sage root preparation can effectively be had children outside the state plan the reason intellectual deterioration that dose application caused to Antiglucocorticoid for a long time, improves the ability of learning and memory of prednisone rat.

(2) compound red sage root preparation can be effectively has children outside the state plan for a long time to Antiglucocorticoid that MAO content raises and the decline of SOD content in the rat cerebral tissue due to the reason dose application, but the content of the TchE that also significantly raises and GC group be (seeing Table 3.6) relatively, the MAO of CDH, CDM and three groups of rats of CDL descends 14%, 22% and 44% respectively, (P＜0.05), SOD content 36%, 44% and 33% (the seeing Table 3.6) that raise respectively, (P＜0.05) takes place obviously to change in enzyme content in the homogenate, and the prompting compound red sage root preparation can prevent effectively that the cerebral tissue zymetology due to the prednisone from changing.The TchE content of CDH, CDM and three groups of rats of CDL all significantly raises (P＜0.05) than the NS group, and is especially obvious with CDL, and its effect needs further to inquire into.

(3) can effectively to prevent glucocorticoid to have children outside the state plan the unit are hippocampal neuron countings of the rat hippocampus cell injury compound red sage root preparation three group rats of reason due to the dose application for a long time all on the rise for compound red sage root preparation; CDH; the neural pyramidal cell number of CDM and three groups of rat hippocampus specific regions of CDL unit are rises 11% than the GC group respectively; 15% and 9%; visible giant pyramidal cells queueing discipline under the mirror; degeneration is few; obviously improve than the GC group; the prompting compound red sage root preparation can effectively be protected the hippocampal neurons injury that is caused by the glucocorticoid prolonged application, and this may can effectively improve the neurocyte degeneration that the glucocorticoid prolonged application causes and reduce prednisone rat brain energy expenditure relevant with it.

(4) compound red sage root preparation can effectively prevent glucocorticoid have children outside the state plan for a long time reason due to the dose application the infringement of rat brain cortex neurocyte and the structural change mirror under visible CDH, CDM and three groups of rat cerebral cortex cellularity of CDL clear, the ratio of molecular layers thick/total skin thickness descends, the odds ratio GC group of the molecular layers thick of two groups of CDM and CDL/total skin thickness significantly descend (P＜0.05) wherein, the infringement that the prompting compound red sage root preparation is had children outside the state plan the rat cerebral cortex of reason due to the dose application for a long time to glucocorticoid has certain preventive effect, but dose-effect relationship is not obvious.

(5) the compound red sage root preparation possible cause experimental result that can effectively prevent glucocorticoid to have children outside the state plan the rat maincenter toxic action of reason due to the dose application for a long time shows, compound red sage root preparation has significant preventive effect to the rat nervous system damage that the prednisone prolonged application causes, compare with the GC group, the nervous function of three groups of rats of compound red sage root preparation is obviously improved.The possible cause that compound red sage root preparation produces this kind effect is: 1. Radix Salviae Miltiorrhizae can obviously improve the metabolism disorder of monoamines material in the brain after brain blood supply and the cerebral ischemia, neurotransmitter and vaso-active substance also there are good regulating action, can significantly improve the content of neuropeptide and excitatory amino acid and the content of eliminating oxygen-derived free radicals in the brain.2. the Radix Astragali has life lengthening, direct anti-aging effects, can increase in the blood and brain in the content of SOD, and obviously reduce the generation of interior free yl.The prednisone rat presents old and feeble sign in this experiment, and the direct anti-aging effects of the Radix Astragali should have certain preventive effect to it.3. the Herba Epimedii and the Rhizoma Atractylodis Macrocephalae ^[93]All are medicines, can improve the content of SOD in vivo, effectively remove the free radical that metabolism produces, body is had the certain protection effect with anti-aging effects; On the other hand, but Herba Epimedii cerebral blood flow increasing amount, reduce cerebral vascular resistance and the Rhizoma Atractylodis Macrocephalae ^[98]Vegetative nerve function is had dual regulation, and both all have the certain protection function to nervous system.Long-term excusing from death reason dosage uses glucocorticoid can cause that neurotransmitter content changes in the brain, cause hippocampal neuron degeneration necrosis and cerebral cortex structure disturbance, cause rat labyrinth accuracy obviously to descend, and compound red sage root preparation has significant preventive effect to this, so be applicable to the prevention of the nervous function disorder due to the prolonged application glucocorticoid.

Conclusion:

4 monthly age SD rats are irritated and feed prednisone after 12 weeks, its untoward reaction mainly shows as and loses weight, osteoporosis, central nervous system function goes down and immune system and liver function damage, testis, thymus and atrophy of adrenal gland, blood-fat and blood sugar raises and HDL content reduction etc.

Compound red sage root preparation can effectively resist prednisone and have children outside the state plan the rat body weight of reason due to the dose application for a long time and alleviate; Can effectively resist rat bone growth inhibited and osteoporosis due to the prednisone, effect strengthens with the increase of drug level, and its effect may be relevant with osteoclast activity with its coordination osteoblast; The central nervous system in rat untoward reaction that the reason dose application of the antagonism prednisone having children outside the state plan for a long time causes and high CH, high Glu and low HDL mass formed by blood stasis and adrenal gland, thymus and atrophy of testis have certain antagonism, have children outside the state plan the untoward reaction of reason due to the dose application for a long time because of it can resist prednisone from many aspects, and toxicity is lower.

Studies show that this preparation can improve the intelligence of prednisone rat, the damage of the hippocampal neuron that effectively prevents and treats prednisone and caused and the content of reduction and old and feeble relevant enzyme also have the good preventing effect to osteoporosis.Therefore, this preparation can be used for improving the body memory ability, and the generation of prevention intellectual deterioration and slow down aging especially can effectively prevent brain injury and osteoporosis after the dose application glucocorticoid is managed in long-term excusing from death.

Claims (5)

1, a kind of have a defying age, prevents and treats osteoporosis and encephaloclastic compound preparation, it is characterized in that this compound preparation is made up of salviamiltiorrhizabung, Herba Epimedii, the Radix Astragali and Rhizoma Atractylodis Macrocephalae extract.

2, as claimed in claim 1 have a defying age, prevent and treat osteoporosis and encephaloclastic compound preparation, Chinese medicine extract in this compound preparation is meant the extract by following prepared: Radix Salviae Miltiorrhizae, Herba Epimedii, the Radix Astragali and the Rhizoma Atractylodis Macrocephalae are ground into powder, add 75～95% soak with ethanol 24 hours, filter, reclaim ethanol, it is standby to get ethanol extract, and medicinal residues decoct with water 3 times again, merge three times filtrate, filter, filtrate is concentrated in right amount, adds aforesaid ethanol extract, cold drying, pulverize, add appropriate amount of auxiliary materials, make granule, tabletting, sugar coating or encapsulated, promptly.

3, as claimed in claim 1 have a defying age, prevent and treat osteoporosis and encephaloclastic compound preparation, Chinese medicine extract in this compound preparation is meant the extract by following prepared: Radix Salviae Miltiorrhizae, Herba Epimedii, the Radix Astragali and Rhizoma Atractylodis Macrocephalae raw medicinal herbs decoct with several times water logging bubble back, repeat 3 times, merging filtrate concentrates, and adds 3 times of ethanol precipitations, behind the recovery ethanol, use ethyl acetate extraction, reclaim ethyl acetate and obtain the Chinese medicine total extract.The Chinese medicine total extract adds appropriate amount of auxiliary materials, makes granule, tabletting, and sugar coating or encapsulated, promptly.

4, as claimed in claim 1 have a defying age, prevents and treats osteoporosis and encephaloclastic compound preparation, it is characterized in that this compound preparation contains danshensu and TANSHINONES.

5, as claimed in claim 1 have a defying age, prevents and treats osteoporosis and encephaloclastic compound preparation, it is characterized in that this compound preparation contains Herba Epimedii total flavones.