Application of traditional Chinese medicine composition in preparation of medicine for promoting fracture healing

Abstract

The invention relates to the technical field of traditional Chinese medicines, in particular to application of a traditional Chinese medicine composition in preparation of a medicine for promoting bone healing. Researches find that the traditional Chinese medicine composition prepared from rhizoma arisaematis, radix aconiti, clove, cinnamon, radix angelicae, asarum, ligusticum wallichii, cynanchum paniculatum, frankincense, myrrh, camphor and borneol can obviously improve fracture bone nonunion of rats and promote callus formation; the traditional Chinese medicine composition has the advantages that bone morphogenetic protein-2 (BMP-2) and insulin growth factors (IGF-1) in callus tissues can be obviously improved, the levels of transforming growth factors-beta (TGF-beta) and vascular growth factors (VEGF) in serum of rats can be obviously up-regulated, and the traditional Chinese medicine composition has a good treatment effect on promoting osteoblast differentiation and proliferation and promoting bone defect healing.

Description

Technical Field

[0001] The present invention relates to the technical field of traditional Chinese medicine, and particularly relates to the application of a traditional Chinese medicine composition in the preparation of a drug for promoting fracture healing. Background Art

[0002] Fracture is a common disease in clinical practice. The process of bone healing is very complex and is affected by many factors in the body. Clinically, about 5%-10% of fracture patients will have delayed bone healing or even nonunion, which seriously affects the quality of life of patients. A fracture means that the bone or the structure of the bone is completely or partially broken. It is more common in children and the elderly, and also occurs in young and middle-aged people from time to time. Fractures can be caused by trauma and bone diseases (such as osteomyelitis, bone tumors, osteoporosis, etc., which cause bone destruction, reduce the ability to resist external forces, and fractures can occur with slight external forces), and the latter is called pathological fracture due to pathogenesis. Clinically, the most common one is the former, and the causes are as follows: 1. Direct violence: The direct action of violence causes fractures at the injured site, often accompanied by varying degrees of soft tissue damage. 2. Indirect violence: The violence causes fractures at a distant part of the limb through conduction, lever, rotation and muscle contraction. 3. Cumulative strain: Long-term, repeated, and minor direct or indirect injuries can cause fractures at a specific part of the limb.

[0003] Nonunion of fractures, also known as fracture nonunion, is one of the difficult problems in the rehabilitation medicine of the locomotor system. At present, its definition is not yet clear, but it is generally considered to be a serious complication after fracture in which the fracture healing process terminates and bony union has not been achieved after nine months of normal treatment. Since the prone sites of nonunion of fractures are related to the number of nutrient blood vessels, the tibia with fewer blood vessels has the highest probability of nonunion of fractures. Although in recent years, with the progress of medical technology, the fracture healing rate has been greatly improved, due to some more complex traumas, clinical reports show that the incidence of nonunion of fractures is still 5%-10%. Its treatment is also a long process. Although surgical treatment is the gold standard for clinical treatment, it has the disadvantages of high cost, many risks, great pain, and 5% of patients still cannot achieve normal bony union after secondary or multiple surgeries.

[0004] In view of the above problems, the field of traditional Chinese medicine should actively intervene in the prevention and treatment of fractures and develop symptomatic prescriptions. Summary of the Invention

[0005] In view of this, the purpose of the present invention is to provide the application of a traditional Chinese medicine composition in the preparation of a drug for promoting fracture healing.

[0006] In order to achieve the above-mentioned invention purpose, the present invention provides the following technical solutions:

[0007] The application of a traditional Chinese medicine composition in the preparation of a drug for promoting fracture healing, the traditional Chinese medicine composition comprises the following raw materials in parts by weight, or is made of the following raw materials in parts by weight:

[0008] 70 - 110 parts of Arisaema heterophyllum Blume, 50 - 90 parts of Aconitum carmichaelii Debx., 40 - 80 parts of Syzygium aromaticum (L.) Merr. & Perry, 40 - 80 parts of Cinnamomum cassia Presl, 80 - 120 parts of Angelica dahurica (Fisch. ex Hoffm.) Benth. & Hook. f. ex Franch. & Sav., 20 - 70 parts of Asarum sieboldii Miq., 50 - 90 parts of Ligusticum chuanxiong Hort., 20 - 70 parts of Cynanchum paniculatum (Bunge) Kitagawa, 10 - 40 parts of Olibanum, 10 - 40 parts of Myrrha, 5 - 40 parts of Camphora, 5 - 40 parts of Borneol.

[0009] In the application of the present invention, the weight parts of each raw material in the traditional Chinese medicine composition are as follows:

[0010] 79 - 99 parts of Arisaema heterophyllum Blume, 57 - 77 parts of Aconitum carmichaelii Debx., 46 - 66 parts of Syzygium aromaticum (L.) Merr. & Perry, 46 - 66 parts of Cinnamomum cassia Presl, 90 - 110 parts of Angelica dahurica (Fisch. ex Hoffm.) Benth. & Hook. f. ex Franch. & Sav., 32 - 52 parts of Asarum sieboldii Miq., 57 - 77 parts of Ligusticum chuanxiong Hort., 32 - 52 parts of Cynanchum paniculatum (Bunge) Kitagawa, 18 - 28 parts of Olibanum, 18 - 28 parts of Myrrha, 11 - 31 parts of Camphora, 11 - 31 parts of Borneol.

[0011] In the application of the present invention, the weight parts of each raw material in the traditional Chinese medicine composition are: 80.1 parts of Arisaema heterophyllum Blume, 60 parts of Aconitum carmichaelii Debx., 50.1 parts of Syzygium aromaticum (L.) Merr. & Perry, 50.1 parts of Cinnamomum cassia Presl, 90 parts of Angelica dahurica (Fisch. ex Hoffm.) Benth. & Hook. f. ex Franch. & Sav., 37.5 parts of Asarum sieboldii Miq., 60 parts of Ligusticum chuanxiong Hort., 37.5 parts of Cynanchum paniculatum (Bunge) Kitagawa, 24.9 parts of Olibanum, 24.9 parts of Myrrha, 18.9 parts of Camphora, 18.9 parts of Borneol.

[0012] In the application of the present invention, the promotion of fracture healing includes at least one of the following 1) to 3):

[0013] 1) Promote the formation of callus at the fracture site;

[0014] 2) Increase the expression level of bone morphogenetic protein (BMP - 2) and / or insulin - like growth factor - 1 (IGF - 1);

[0015] 3) Up - regulate the level of transforming growth factor - β and / or vascular endothelial growth factor.

[0016] In some specific embodiments, the bone morphogenetic protein (BMP - 2) and / or insulin - like growth factor - 1 are from callus tissue; the growth factor - β and vascular endothelial growth factor are from serum.

[0017] In the application of the present invention, the traditional Chinese medicine composition can directly grind each traditional Chinese medicine into powder, or can be an extract obtained by conventional means in the art and the like. The traditional Chinese medicine materials used in the composition of the present invention can be raw herbs directly ground into powder, raw herbs ground into powder after drying, or extracts of raw herbs. It can also be processed products of raw herbs, such as processed products after processing, etc. The specific forms include powder after grinding, powder after drying and grinding, or its extract. In order to retain the active ingredients in the herbs to a greater extent and avoid the destruction of active ingredients by high temperature, the drying is preferably low - temperature drying.

[0018] In some embodiments, the traditional Chinese medicine composition is used in the form of components ground after low-temperature drying, and the specific preparation method includes: taking Arisaema heterophyllum Blume, Radix Aconiti, Flos Caryophylli, Cortex Cinnamomi, Radix Angelicae Dahuricae, Herba Asari, Rhizoma Chuanxiong, Herba Cynanchi Paniculati, Olibanum, Myrrha, drying them at low temperature, pulverizing and sieving; then adding Camphora and Borneolum Syntheticum and mixing them.

[0019] In the application of the present invention, the effective dose of the traditional Chinese medicine composition is 2.765 g to 5.529 g of crude drug per day.

[0020] Research shows that, compared with the model group, the traditional Chinese medicine composition of the present invention can significantly promote the formation of callus at the fracture site, significantly increase the concentrations of bone morphogenetic protein (BMP-2) and insulin-like growth factor-1 (IGF-1) in the callus tissue of rats, and significantly up-regulate the levels of transforming growth factor-β (TGF-β) and vascular endothelial growth factor (VEGF) in the serum of rats. It is suggested that the traditional Chinese medicine composition of the present invention can effectively improve the quality of bone healing, promote the healing of bone defects, and has a good promoting effect on the formation of callus at the fracture site.

[0021] The present invention also provides a drug for promoting fracture healing, which comprises the following raw materials in parts by weight, or is made from the following raw materials in parts by weight:

[0022] 70 - 110 parts of Arisaema heterophyllum Blume, 50 - 90 parts of Radix Aconiti, 40 - 80 parts of Flos Caryophylli, 40 - 80 parts of Cortex Cinnamomi, 80 - 120 parts of Radix Angelicae Dahuricae, 20 - 70 parts of Herba Asari, 50 - 90 parts of Rhizoma Chuanxiong, 20 - 70 parts of Herba Cynanchi Paniculati, 10 - 40 parts of Olibanum, 10 - 40 parts of Myrrha, 5 - 40 parts of Camphora, 5 - 40 parts of Borneolum Syntheticum.

[0023] The drug provided by the present invention also includes pharmaceutically acceptable excipients, such as rosin, paraffin, petrolatum, liquid paraffin, etc. Those skilled in the art can select appropriate types of excipients according to needs and make the required traditional Chinese medicine dosage forms according to the conventional preparation methods in pharmacy. The dosage forms include but are not limited to decoction, granule, capsule, tablet, pill, oral liquid, tincture, syrup, suppository, gel, spray, injection, ointment, plaster, rubber plaster and cataplasm.

[0024] In the present invention, the drug is a plaster, and the preparation method of the plaster includes:

[0025] Taking Arisaema heterophyllum Blume, Radix Aconiti, Flos Caryophylli, Cortex Cinnamomi, Radix Angelicae Dahuricae, Herba Asari, Rhizoma Chuanxiong, Herba Cynanchi Paniculati, Olibanum, Myrrha, drying them at low temperature, pulverizing them into fine powder, sieving, and mixing to obtain the medicinal powder;

[0026] Take rosin, paraffin wax, petrolatum, and liquid paraffin, heat them until melted, and when the temperature drops below 65°C, successively add methyl salicylate, camphor, borneol, and the said medicinal powder and mix them evenly to prepare a coating; use the said coating to prepare a plaster.

[0027] The present invention provides an application of a traditional Chinese medicine composition in the preparation of a drug for promoting fracture healing. The traditional Chinese medicine composition is prepared from Arisaema heterophyllum Blume, Aconitum carmichaelii Debx., Syzygium aromaticum (L.) Merr. & Perry, Cinnamomum cassia Presl, Angelica dahurica (Fisch. ex Hoffm.) Benth. & Hook. f. ex Franch. & Sav., Asarum sieboldii Miq., Ligusticum wallichii Franch., Cynanchum paniculatum (Bunge) Kitag., Olibanum, Myrrha, camphor, and borneol. The experimental results show that the traditional Chinese medicine composition can significantly promote the formation of callus at the fracture site, significantly increase the concentrations of bone morphogenetic protein (BMP-2) and insulin-like growth factor-1 (IGF-1) in the callus tissue of rats, and significantly up-regulate the levels of transforming growth factor-β (TGF-β) and vascular endothelial growth factor (VEGF) in the serum of rats. It is suggested that the traditional Chinese medicine composition of the present invention can effectively improve the quality of bone healing, promote the healing of bone defects, and has a good effect on the formation of callus at the fracture site. Detailed implementation manners

[0028] The present invention provides an application of a traditional Chinese medicine composition in the preparation of a drug for promoting fracture healing. Those skilled in the art can draw on the content of this article and appropriately modify the process parameters to achieve it. It should be particularly noted that all similar substitutions and modifications are obvious to those skilled in the art, and they are all regarded as included in the present invention. The methods and applications of the present invention have been described through preferred embodiments, and those related can obviously make changes or appropriate modifications and combinations to the methods and applications in this article without departing from the content, spirit, and scope of the present invention to implement and apply the technology of the present invention.

[0029] In this article, descriptions such as "comprising", "including", and "having" include closed technical solutions composed of the listed features, and also include open technical solutions containing the listed features.

[0030] In this article, the term "and / or" includes any and all combinations of one or more of the related listed items.

[0031] In this article, regarding numerical ranges, unless otherwise specified, the above numerical ranges are considered continuous, and include the minimum value and the maximum value of this range, as well as every point value between such minimum value and maximum value. Further, when the range refers to integers, it includes every integer between the minimum value and the maximum value of this range. In addition, when providing multiple ranges to describe features or characteristics, these ranges can be combined. Unless otherwise specified, all ranges disclosed in this article should be understood to include any and all sub-ranges subsumed therein.

[0032] As described above, the present invention provides an application of a traditional Chinese medicine composition in the preparation of a drug for promoting fracture healing;

[0033] The traditional Chinese medicine composition comprises raw materials in the following parts by weight, or is made from raw materials including the following parts by weight:

[0034] 70 - 110 parts of Arisaema heterophyllum Blume, 50 - 90 parts of Aconitum carmichaelii Debx., 40 - 80 parts of Flos Caryophylli, 40 - 80 parts of Cortex Cinnamomi, 80 - 120 parts of Radix Angelicae Dahuricae, 20 - 70 parts of Herba Asari, 50 - 90 parts of Rhizoma Chuanxiong, 20 - 70 parts of Herba Cynanchi Paniculati, 10 - 40 parts of Olibanum Praeparatum, 10 - 40 parts of Myrrha Praeparata, 5 - 40 parts of Borneolum Syntheticum, 5 - 40 parts of Borneol.

[0035] The present invention also provides a preparation method of the traditional Chinese medicine composition, comprising:

[0036] Take Arisaema heterophyllum Blume, Aconitum carmichaelii Debx., Flos Caryophylli, Cortex Cinnamomi, Radix Angelicae Dahuricae, Herba Asari, Rhizoma Chuanxiong, Herba Cynanchi Paniculati, Olibanum Praeparatum, Myrrha Praeparata, after drying at low temperature, pulverize and sieve, and mix with Borneolum Syntheticum and Borneol to obtain.

[0037] The present invention also provides a preparation method of the drug for promoting fracture healing. The dosage form of the drug is a plaster, and its preparation method comprises:

[0038] Take Arisaema heterophyllum Blume, Aconitum carmichaelii Debx., Flos Caryophylli, Cortex Cinnamomi, Radix Angelicae Dahuricae, Herba Asari, Rhizoma Chuanxiong, Herba Cynanchi Paniculati, Olibanum Praeparatum, Myrrha Praeparata, dry at low temperature, pulverize into fine powder, sieve, mix evenly to obtain a drug powder for standby; separately take colophony, paraffin, vaseline, liquid paraffin, heat to melting, when the temperature drops below 65°C, sequentially add methyl salicylate, Borneolum Syntheticum, Borneol and the above-mentioned drug powder, mix evenly to form a coating, and perform plaster coating, cutting, lining, slicing to make a plaster.

[0039] Specifically, the preparation method of the above plaster comprises:

[0040] Take 267 g of Arisaema heterophyllum Blume, 200 g of Aconitum carmichaelii Debx., 167 g of Flos Caryophylli, 167 g of Cortex Cinnamomi, 300 g of Radix Angelicae Dahuricae, 125 g of Herba Asari, 200 g of Rhizoma Chuanxiong, 125 g of Herba Cynanchi Paniculati, 83 g of Olibanum Praeparatum, 83 g of Myrrha Praeparata, dry at low temperature, pulverize into fine powder, sieve, mix evenly to obtain a drug powder for standby; separately take 267 g of colophony, 1330 g of paraffin, 800 g of vaseline, 233 g of liquid paraffin, heat to about 100°C, melt, when the temperature drops to 100°C, sequentially add 160 g of methyl salicylate, 63 g of Borneolum Syntheticum, 63 g of Borneol and the above-mentioned drug powder, mix evenly to form a coating, and perform plaster coating, cutting, lining, slicing to make a plaster.

[0041] In the present invention, the traditional Chinese medicine composition and the medicine can significantly promote the formation of callus at the fracture site, significantly increase the concentrations of bone morphogenetic protein (BMP-2) and insulin-like growth factor-1 (IGF-1) in the callus tissue of rats, and significantly up-regulate the levels of transforming growth factor-β (TGF-β) and vascular endothelial growth factor (VEGF) in the serum of rats. It is suggested that the traditional Chinese medicine composition of the present invention can effectively improve the quality of bone healing, promote the healing of bone defects, and has a good effect on the formation of callus at the fracture site.

[0042] The test materials used in the present invention are all ordinary commercially available products and can be purchased on the market.

[0043] The present invention will be further described below in conjunction with examples:

[0044] Example 1 Preparation of the plaster of the composition of the present invention

[0045] Formulation: 267 g of raw Arisaema heterophyllum Blume, 200 g of raw Radix Aconiti Carmichaeli Debx., 167 g of Flos Caryophylli, 167 g of Cortex Cinnamomi, 300 g of Radix Angelicae Dahuricae, 125 g of Herba Asari, 200 g of Rhizoma Chuanxiong, 125 g of Herba Cynanchi Paniculati, 83 g of processed Olibanum, 83 g of processed Myrrha, 63 g of Camphora, 63 g of Borneolum Syntheticum.

[0046] The preparation method of the traditional Chinese medicine composition plaster is as follows:

[0047] Take the above twelve raw medicinal materials. Except for Camphora and Borneolum Syntheticum, take the ten raw medicinal materials such as Arisaema heterophyllum Blume, etc., dry them at low temperature, crush them into fine powder, sieve them, mix them evenly, and set aside. Take another 267 g of Colophonium, 1330 g of Paraffin, 800 g of Vaseline, and 233 g of Liquid Paraffin, heat them to about 100 °C until melted. When the temperature drops to 60 °C, add 160 g of Methyl Salicylate, Camphora, Borneolum Syntheticum and the above-mentioned medicinal powder in sequence, mix them evenly to make a coating, and then carry out plaster coating, cutting, lining, and slicing to make a plaster with a size of 10 cm * 13 cm.

[0048] Example 2 Therapeutic effect of the traditional Chinese medicine composition on the rat tibial nonunion model

[0049] 1. Experimental materials

[0050] 1.1 Animals

[0051] SD rats, male, SPF grade, 72 rats, weighing 160 - 180 g, purchased from Spf (Suzhou) Biotechnology Co., Ltd., animal certificate number: NO.202364746.

[0052] Feeding environment: room temperature 20 - 26 °C, relative humidity controlled at 40 - 70%, feeding density 6 rats / cage.

[0053] 1.2 Medicines

[0054] Traditional Chinese medicine composition, prepared by the method of Example 1, Jiangsu Kanion Pharmaceutical Co., Ltd.; Indometacin Cataplasm (Wuhan Bingbing Pharmaceutical Co., Ltd.); Chloral Hydrate; Penicillin; Normal Saline (Chenxin Pharmaceutical Co., Ltd., batch number: 2101282721), VEGF / TGF-β Enzyme-Linked Immunosorbent Assay (Elisa) Kit.

[0055] 1.3 Instruments

[0056] Electronic balance (Sartorius Scientific Instruments Company, model BS224S); Low-temperature centrifuge (Eppendorf Company, Germany, model 5804R); Purified water instrument (Millipore Company, Germany, model: Milli-Q Plus); Animal weighing scale (Shanghai Xiangchuan Electronic Weighing Instrument Co., Ltd., model: CX-SC-DA); Microplate reader (Molecular Devices, USA, model Flexstation3).

[0057] 2. Dosage Design

[0058] The daily dosage of the traditional Chinese medicine composition is 5.529 g of crude drug / day. According to the body surface area conversion method, the equivalent dose for rats is 5.529 g / 60 kg * 6.2 = 0.571 g of crude drug / kg. The maximum daily dosage for rats is 0.571 * 200 / 1000 g = 0.11 g / rat. Human single use: 1.843 g / 10 * 13 cm 2 = 0.014 g / cm 2 The single use patch area for rats is 7.9 cm 2 The size of the cataplasm is 3 * 3 cm 2 . According to the 1 / 4:1 / 2:1 dosing method, the low dose is 0.15 g of crude drug / kg / d, the medium dose is 0.30 g of crude drug / kg / d, and the high dose is 0.60 g of crude drug / kg / d. Once every other day.

[0059] The daily dosage of the indometacin cataplasm is 45.5 mg / day. According to the body surface area conversion method, the equivalent dose for rats is 45.5 mg / 60 kg * 6.2 = 4.70 mg / kg. The maximum daily dosage for rats is: 4.70 * 200 / 1000 g = 0.94 mg / rat. Human single use: 45.5 mg / 10 * 14 cm 2 = 0.325 mg / cm 2 The single use patch area for rats is 2.89 cm 2 The size of the cataplasm is 2 * 2 cm 2 . Once every other day.

[0060] 3. Experimental Method

[0061] 3.1 Model Preparation

[0062] Seventy-two SD rats were randomly divided into cages, with 6 rats in each cage. They were adaptively fed with regular feed in a laboratory at an environmental temperature of 24°C and a humidity of 40% - 50%. When their body weights all reached 300 ± 30 g, modeling was started. They were randomly grouped into a sham operation group, a model group, a positive drug indomethacin cataplasm group, and high, medium, and low dose groups of the traditional Chinese medicine composition, with 12 rats in each group. All rats were intraperitoneally injected with anesthesia before surgery using 10% chloral hydrate by volume at a dose of 0.03 mL / kg. -1 After anesthesia, the rats were fixed in the supine position on the operating table. The right tibia of the rats was exposed, the hair was shaved off and disinfected with an electric clipper. A 2-cm incision was made in the middle of the ventral tibia, and the calf muscles were dissected to fully expose the right tibia. After the calf muscles were dissected in the sham operation group, it was directly sutured. In the modeling group, after a 4-mm bone defect was created in the tibia with a bone clamp, the two broken ends of the fracture were cauterized with an oral filler after being heated red by an alcohol lamp. Internal fixation was performed with a 0.8-mm Kirschner wire using a hand drill. It entered from the bone marrow cavity at the proximal end of the fracture and then passed through the knee joint. Then, the Kirschner wire was driven into the lower end marrow cavity to the ankle in a retrograde manner, and the excess steel needles at both ends were cut off. The needle tips were left subcutaneously. Finally, the wound was cleaned and sutured in two layers. During the operation, every effort was made to avoid breaking the fibula or causing bone cortex necrosis by cauterization. After the operation, 80,000 units of penicillin were routinely given by intramuscular injection once a day for 3 consecutive days.

[0063] 3.2 Animal grouping and drug administration

[0064] Twelve rats were randomly selected from the model group 8 weeks after modeling. After gross observation and X-ray examination, it was shown that the fracture ends were atrophied and sclerosed, the bone marrow cavity was closed, and there was no obvious callus growth, confirming that the non-union fracture model was successfully established. Then, the remaining 48 successfully modeled rats were randomly divided into 4 groups, namely the positive drug indomethacin cataplasm group, and high, medium, and low dose groups of the traditional Chinese medicine composition cataplasm, with 12 rats in each group.

[0065] (1) Sham operation group (n = 12): Normal feeding, without any intervention;

[0066] (2) Model group (n = 12): Normal feeding, without any intervention;

[0067] (3) Indomethacin cataplasm group (n = 12): Normal feeding, and cataplasms of corresponding sizes were applied to the fracture site. The medicine was changed every other day, and it was applied 3 times in total.

[0068] (4) High, medium, and low dose groups of the traditional Chinese medicine composition cataplasm (n = 12): Normal feeding, and cataplasms of corresponding sizes were applied to the fracture site. The medicine was changed every other day, and it was applied 3 times in total.

[0069] At the 4th and 8th weekends after drug administration, 6 experimental animals were selected from each group. They were anesthetized using 10% chloral hydrate by volume at a dose of 0.03 mL / kg. -1Anesthetize the rats by intraperitoneal injection before surgery at a certain dose. Take blood from the abdominal aorta, centrifuge to obtain serum, and use it for detecting the contents of VEGF / TGF-β with a kit.

[0070] 4. Statistical methods

[0071] The results were statistically analyzed by t-test.

[0072] 5. Experimental results

[0073] 5.1 Effects on the content of vascular endothelial growth factor (VEGF) in the serum of the rat tibial nonunion model After 4 weeks of drug administration, compared with the sham operation control group, the VEGF level in the model group was significantly decreased (P < 0.01); compared with the model group, the VEGF levels in the positive drug indomethacin group and each group of the traditional Chinese medicine composition were significantly increased (P < 0.01); compared with the positive drug indomethacin group, the VEGF content in the high-dose group of the traditional Chinese medicine composition was significantly increased (P < 0.05), and there was no significant difference between the medium-dose and low-dose groups of the traditional Chinese medicine composition (P > 0.05). After 8 weeks of drug administration, compared with the sham operation control group, the VEGF level in the model group was significantly decreased (P < 0.01); compared with the model group, the VEGF levels in the positive drug indomethacin group and each group of the traditional Chinese medicine composition were significantly increased (P < 0.01); compared with the positive drug indomethacin group, the VEGF contents in the high-dose and medium-dose groups of the traditional Chinese medicine composition were significantly increased (P < 0.01). The results are shown in Table 1.

[0074] Table 1 Comparison of the VEGF protein levels in the serum of each group ( n = 6)

[0075]

[0076] Note: Compared with the sham operation group, & P < 0.05, && P < 0.01; compared with the model group, # P < 0.05, ## P < 0.01; compared with the positive drug indomethacin control group, *P < 0.05, **P < 0.01.

[0077] 5.2 Effects on the content of transforming growth factor-β (TGF-β) in the serum of the rat tibial nonunion model

[0078] After 4 weeks of administration, the TGF-β protein level in the model group was significantly lower than that in the sham operation control group (P < 0.01); compared with the model group, there was no significant difference in the sham operation group, and the TGF-β content in the serum of the positive drug indomethacin group and the high and medium dose groups of the traditional Chinese medicine composition was significantly increased (P < 0.01). After 8 weeks of administration, compared with the model group, the TGF-β content in the serum of the positive drug indomethacin group and the high and medium dose groups of the traditional Chinese medicine composition was significantly increased (P < 0.01); compared with the positive drug indomethacin group, the TGF-β content in the serum of the high dose group of the traditional Chinese medicine composition was significantly increased (P < 0.05). The results are shown in Table 2.

[0079] Table 2 Comparison of TGF-β protein levels in the sera of each group ( n = 6)

[0080]

[0081] Note: Compared with the sham operation group, & P < 0.05, && P < 0.01; compared with the model group, # P < 0.05, ## P < 0.01; compared with the positive drug indomethacin control group, *P < 0.05, **P < 0.01.

[0082] 6. Experimental conclusions

[0083] The traditional Chinese medicine composition at 0.6 and 0.3 g crude drug / kg can significantly up-regulate the levels of transforming growth factor-β (TGF-β) and vascular endothelial growth factor (VEGF) in the serum of rats and promote fracture healing. It is suggested that the traditional Chinese medicine composition has a certain therapeutic effect on nonunion of rat tibia.

[0084] Example 3 Therapeutic effect of traditional Chinese medicine composition on nonunion model of rat femur

[0085] 1. Experimental materials

[0086] 1.1 Animals

[0087] SD rats, male, SPF grade, 72 rats, weighing 240 - 260 g, purchased from Beijing SPF Biotechnology Co., Ltd., animal certificate number: NO.110324231106389814.

[0088] Breeding environment: room temperature 20 - 26 °C, relative humidity controlled at 40 - 70%, breeding density 6 rats / cage.

[0089] 1.2 Drugs

[0090] Traditional Chinese medicine composition, prepared by the method of Example 1, Jiangsu Kanion Pharmaceutical Co., Ltd.; Indometacin Cataplasm (Wuhan Bingbing Pharmaceutical Co., Ltd.); Chloral Hydrate; Penicillin; Normal Saline (Chenxin Pharmaceutical Co., Ltd., batch number: 2101282721), BMP-2 / IGF-1 Enzyme-Linked Immunosorbent Assay (Elisa) Kit.

[0091] 1.3 Instruments

[0092] Electronic balance (Sartorius Scientific Instruments Company, model BS224S); Low-temperature centrifuge (Eppendorf Company, Germany, model 5804R); Purified water instrument (Millipore Company, Germany, model: Milli-Q Plus); Animal weighing scale (Shanghai Xiangchuan Electronic Weighing Apparatus Co., Ltd., model: CX-SC-DA); Microplate reader (Molecular Devices, USA, model Flexstation3).

[0093] 2. Dosage Design

[0094] The daily dosage of the traditional Chinese medicine composition is 5.529 g of crude drug / day. According to the body surface area conversion method, the equivalent dose for rats is 5.529 g / 60 kg * 6.2 = 0.571 g of crude drug / kg. The maximum daily dosage for rats is 0.571 * 200 / 1000 g = 0.11 g / rat. Human single use: 1.843 g / 10 * 13 cm 2 = 0.014 g / cm 2 , The single use patch area for rats is 7.9 cm 2 , The size of the cataplasm is 3 * 3 cm 2 . According to the 1 / 4:1 / 2:1 dosing method, the low dose is 0.15 g of crude drug / kg / d, the medium dose is 0.30 g of crude drug / kg / d, and the high dose is 0.60 g of crude drug / kg / d. Once every other day.

[0095] The daily dosage of the indometacin cataplasm is 45.5 mg / day. According to the body surface area conversion method, the equivalent dose for rats is 45.5 mg / 60 kg * 6.2 = 4.70 mg / kg. The maximum daily dosage for rats is: 4.70 * 200 / 1000 g = 0.94 mg / rat. Human single use: 45.5 mg / 10 * 14 cm 2 = 0.325 mg / cm 2 , The single use patch area for rats is 2.89 cm 2 , The size of the cataplasm is 2 * 2 cm 2 . Once every other day.

[0096] 3. Experimental Method

[0097] 3.1 Model Preparation

[0098] Seventy-two SD rats were randomly caged, six rats in each cage, and adaptively fed with conventional feed in a laboratory at an environmental temperature of 24 °C and a humidity of 40% - 50%. When the body weights of all rats reached 300 ± 30 g, modeling was started. The rats were randomly divided into a sham operation group, a model group, a positive drug indomethacin cataplasm group, and high, medium, and low dose groups of the traditional Chinese medicine composition, with 12 rats in each group. All rats were intraperitoneally injected with anesthesia before surgery at a dose of 10% chloral hydrate by volume at 0.03 mL / kg -1 After anesthesia, the rats were fixed in the supine position on the operating table. The right hind limb was routinely disinfected and covered with a sterile towel. A longitudinal incision about 1 cm long was made on the outside of the right knee. The distal femoral intercondylar fossa was exposed by traction. A Kirschner wire with a diameter of 1.2 mm was retrogradely inserted from the center point of the femoral intercondylar fossa and slowly passed through the medullary cavity with a low-speed electric drill. Finally, the Kirschner wire retrogradely pierced out about 5 mm from the greater trochanter. The distal end of the Kirschner wire was buried under the femoral condylar cortical bone and cut off. After irrigation, the incision was sutured with 5-0 absorbable sutures. Then, an incision about 2 cm long was made on the outside of the middle segment of the femur. The femur was exposed along the muscle space. The femur was transected at intervals of 3 mm by about 50% of the cross-section with an electric oscillating saw. Then, the bone tissue between the two cross-sections was clamped off and removed with a small hemostat to create a bone defect of about 3 mm. Finally, the outer periosteum of both cut ends was circularly cauterized with a high-frequency electric knife in the electrocoagulation mode (30 watts) for 2 mm. During the operation, the surrounding muscle tissues were protected. It was seen that the fracture alignment was good, without rotation, and hemostasis was complete. Each layer was sutured in turn. After surgery, penicillin 80,000 units was routinely given by intramuscular injection once a day for 3 consecutive days.

[0099] 3.2 Animal grouping and administration

[0100] Twelve rats were randomly selected from the model group 8 weeks after modeling. After gross observation and X-ray examination, it was shown that the fracture ends were atrophied and sclerosed, the medullary cavity was closed, and there was no obvious callus growth, confirming the successful establishment of the non-union fracture model. Then, the remaining 48 rats were randomly divided into 4 groups, namely the positive drug indomethacin cataplasm group, and high, medium, and low dose groups of the traditional Chinese medicine composition cataplasm, with 12 rats in each group.

[0101] (1) Sham operation group (n = 12): Normal feeding, without any intervention;

[0102] (2) Model group (n = 12): Normal feeding, without any intervention;

[0103] (3) Indomethacin cataplasm group (n = 12): Normal feeding, apply the cataplasm of corresponding size at the fracture site, change the dressing every other day, and apply it 3 times in total.

[0104] (4) High, medium, and low dose groups of the traditional Chinese medicine composition cataplasm (n = 12): Normal feeding, apply the cataplasm of corresponding size at the fracture site, change the dressing every other day, and apply it 3 times in total.

[0105] At the 3rd and 8th weeks after administration respectively, 6 rats were selected from each group of experimental animals and sacrificed. The periosteum and inner muscle tissues about 1 cm above and below the fracture site of the rats were dissected; in the sham operation group, the periosteum and inner muscle tissues about 1 cm in the middle of the femur were dissected and stored in a -80 °C refrigerator for later testing. Before testing, the samples were first thawed, homogenized, and the supernatant was taken to prepare samples for detecting the contents of BMP-2 and IGF-1 using the test kit.

[0106] 4. Statistical method

[0107] The results were statistically processed using the t-test.

[0108] 5. Experimental results

[0109] 5.1 Effects on the content of bone morphogenetic protein-2 (BMP-2) in the callus tissue of the rat femoral nonunion model

[0110] Three weeks after administration, compared with the model group, the contents of BMP-2 in the callus of the positive drug indomethacin, the high and medium dose groups of the traditional Chinese medicine composition were significantly increased (P < 0.01); compared with the sham operation group, the content of BMP-2 in the callus of the positive drug indomethacin group was significantly increased (P < 0.01), and the contents of BMP-2 in the high and medium dose groups of the traditional Chinese medicine composition were significantly increased (P < 0.05). Eight weeks after administration, the content of BMP-2 in the administration group was lower than that at the 3rd week, but there was no significant difference. The results are shown in Table 3.

[0111] Table 3 Concentrations of bone morphogenetic protein-2 (BMP-2) in the callus tissue of each group n = 6

[0112]

[0113] Note: Compared with the model group, # P < 0.05, ## P < 0.01; compared with the sham operation group, *P < 0.05, **P < 0.01.

[0114] 5.2 Effects on the content of insulin-like growth factor (IGF-1) in the callus tissue of the rat tibial nonunion model

[0115] Three weeks after administration, compared with the sham operation group, the contents of IGF-1 in the callus of the positive drug indomethacin group and the three dose groups of the traditional Chinese medicine composition were significantly increased (P < 0.01), and the content in the medium dose group of the traditional Chinese medicine composition was significantly increased (P < 0.05); compared with the model group, the contents of IGF-1 in the callus of the positive drug indomethacin group and the three dose groups of the traditional Chinese medicine composition were significantly increased (P < 0.01). Eight weeks after administration, the content of IGF-1 in the administration group was lower than that at the 3rd week, but there was no significant difference. The results are shown in Table 4.

[0116] Table 4 Content of insulin growth factor (IGF-1) in callus tissues of each group ( n = 6)

[0117]

[0118] Note: Compared with the model group, # P < 0.05, ## P < 0.01; compared with the sham operation control group, *P < 0.05, **P < 0.01.

[0119] 6. Test conclusion

[0120] The traditional Chinese medicine composition at 0.6 and 0.3 g crude drug / kg can significantly up-regulate the levels of bone morphogenetic protein-2 (BMP-2) and insulin growth factor (IGF-1), promote callus growth and fracture healing. It is indicated that the traditional Chinese medicine composition has a certain therapeutic effect on femoral nonunion in rats.

[0121] The above embodiments of the present invention are only for clearly illustrating the examples made by the present invention, rather than limiting the implementation manners of the present invention. For those of ordinary skill in the art, other different forms of changes or variations can be made based on the above description. It is not necessary and impossible to list all the implementation manners here. Any modifications, equivalent replacements, and improvements made within the spirit and principle of the present invention shall be included in the protection scope of the claims of the present invention.

Claims

1. Use of a traditional Chinese medicine composition in the preparation of a medicament for promoting fracture healing, characterized in that, The traditional Chinese medicine composition comprises raw materials in the following parts by weight, or is made from raw materials in the following parts by weight: Arisaema heterophyllum Blume 70 - 110 parts, Radix Aconiti 50 - 90 parts, Flos Caryophylli 40 - 80 parts, Cortex Cinnamomi 40 - 80 parts, Radix Angelicae Dahuricae 80 - 120 parts, Herba Asari 20 - 70 parts, Rhizoma Chuanxiong 50 - 90 parts, Herba Cynanchi Paniculati 20 - 70 parts, Olibanum 10 - 40 parts, Myrrha 10 - 40 parts, Camphora 5 - 40 parts, Borneolum Syntheticum 5 - 40 parts.

2. The application according to claim 1, characterized in that, The parts by weight of each raw material are: Arisaema heterophyllum Blume 79 - 99 parts, Radix Aconiti 57 - 77 parts, Flos Caryophylli 46 - 66 parts, Cortex Cinnamomi 46 - 66 parts, Radix Angelicae Dahuricae 90 - 110 parts, Herba Asari 32 - 52 parts, Rhizoma Chuanxiong 57 - 77 parts, Herba Cynanchi Paniculati 32 - 52 parts, Olibanum 18 - 28 parts, Myrrha 18 - 28 parts, Camphora 11 - 31 parts, Borneolum Syntheticum 11 - 31 parts.

3. The application according to claim 2, wherein The parts by weight of each raw material are: Arisaema heterophyllum Blume 80.1 parts, Radix Aconiti 60 parts, Flos Caryophylli 50.1 parts, Cortex Cinnamomi 50.1 parts, Radix Angelicae Dahuricae 90 parts, Herba Asari 37.5 parts, Rhizoma Chuanxiong 60 parts, Herba Cynanchi Paniculati 37.5 parts, Olibanum 24.9 parts, Myrrha 24.9 parts, Camphora 18.9 parts, Borneolum Syntheticum 18.9 parts.

4. The application according to any one of claims 1 to 3, characterized in that, The promotion of fracture healing includes at least one of the following 1) - 3): 1) Promote the formation of callus at the fracture site; 2) Increase the expression levels of bone morphogenetic protein (BMP - 2) and / or insulin - like growth factor - 1 (IGF - 1); 3) Up - regulate the levels of transforming growth factor - β and / or vascular endothelial growth factor.

5. The application according to claim 4, characterized in that, The bone morphogenetic protein (BMP - 2) and / or insulin - like growth factor - 1 are from callus tissue; the transforming growth factor - β and vascular endothelial growth factor are from serum.

6. The application according to any one of claims 1 to 5, characterized in that The preparation method of the traditional Chinese medicine composition includes: taking Arisaema heterophyllum Blume, Radix Aconiti, Flos Caryophylli, Cortex Cinnamomi, Radix Angelicae Dahuricae, Herba Asari, Rhizoma Chuanxiong, Herba Cynanchi Paniculati, Olibanum, Myrrha, drying at low temperature, pulverizing and sieving; then adding Camphora and Borneolum Syntheticum and mixing.

7. The application according to any one of claims 1 to 6, characterized in that, The effective dose of the traditional Chinese medicine composition is 2.765 g - 5.529 g of crude drug per day.

8. A drug for promoting fracture healing, characterized in that, Comprises raw materials in the following parts by weight, or is made from raw materials in the following parts by weight: Arisaema heterophyllum Blume 70 - 110 parts, Radix Aconiti 50 - 90 parts, Flos Caryophylli 40 - 80 parts, Cortex Cinnamomi 40 - 80 parts, Radix Angelicae Dahuricae 80 - 120 parts, Herba Asari 20 - 70 parts, Rhizoma Chuanxiong 50 - 90 parts, Herba Cynanchi Paniculati 20 - 70 parts, Olibanum 10 - 40 parts, Myrrha 10 - 40 parts, Camphora 5 - 40 parts, Borneolum Syntheticum 5 - 40 parts.

9. The medicament according to claim 8, wherein Its dosage forms include decoction, granule, capsule, tablet, pill, oral liquid, tincture, syrup, suppository, gel, spray, injection, ointment, plaster, rubber plaster or cataplasm.

10. The drug according to claim 9, characterized in that, The dosage form of the drug is plaster, and the preparation method of the plaster includes: Taking Arisaema heterophyllum Blume, Radix Aconiti, Flos Caryophylli, Cortex Cinnamomi, Radix Angelicae Dahuricae, Herba Asari, Rhizoma Chuanxiong, Herba Cynanchi Paniculati, Olibanum, Myrrha, drying at low temperature, pulverizing into fine powder, sieving, mixing evenly to obtain medicinal powder; Taking colophony, paraffin, vaseline, liquid paraffin, heating to melting, when the temperature drops below 65 °C, successively adding methyl salicylate, Camphora, Borneolum Syntheticum and the medicinal powder and mixing evenly to make a coating; using the coating to make a plaster.

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