Construction method and application of damp-heat downward flow type erectile dysfunction animal model

Abstract

The invention relates to a construction method of a damp-heat downward flow type erectile dysfunction animal model, which comprises the following steps: S1, from the day of modeling, feeding mice with high-sugar and high-fat feed, and simultaneously providing mixed drinking water containing alcohol and honey, free diet and water; s2, from the eighth day of modeling, feeding the mice in a humid and hot environment; s3, on the 15th day, the 20th day and the 25th day of modeling, a prostate tissue emulsion is adopted to conduct subcutaneous injection on the mouse, a cyclophosphamide solution is synchronously injected into the intraperitoneal cavity, and the prostate tissue emulsion is obtained by emulsifying prostate tissue homogenate and a complete Freund's adjuvant according to the volume ratio of 1: 1; and S4, carrying out comprehensive evaluation on the damp-heat downward flow type erectile dysfunction animal model. Internal and external damp-heat factors are combined with drug induction, the pathogenesis of'damp-heat accumulation in lower jiao 'in traditional Chinese medicine is cooperatively simulated through multiple mechanisms, the manifestation symptom of the animal model is almost consistent with the disease symptom of a clinical damp-heat downward flow type impotence patient, and good scientificity and reliability are achieved.

Description

Technical Field

[0001] The present invention relates to the technical field of animal model construction, and particularly relates to a method for constructing an animal model of erectile dysfunction with damp-heat pouring downward and its application. Background Art

[0002] Erectile dysfunction (ED) is a common sexual dysfunction in men. Due to the inability to achieve or maintain an effective penile erection, patients are dissatisfied with their sexual life, which affects their quality of life and relationship with their partners. Traditional Chinese medicine (TCM) refers to ED as "impotence". In TCM, it can be caused by one or more factors such as emotional factors, excessive sexual indulgence, improper diet, falls and injuries, invasion of the six exogenous pathogenic factors, prolonged illness, congenital deficiency, and aging. According to the "Expert Consensus on the Integrated Traditional Chinese and Western Medicine Diagnosis and Treatment of Erectile Dysfunction", TCM divides erectile dysfunction (impotence) into six syndromes: liver qi stagnation, damp-heat pouring downward, blood stasis obstruction, heart-spleen deficiency, kidney yang deficiency, and kidney yin deficiency; the "Guiding Principles for Clinical Research of New Chinese Medicines" divides the syndrome differentiation of impotence into six types: kidney yin deficiency syndrome, liver qi stagnation syndrome, decline of life gate fire syndrome, heart-spleen deficiency syndrome, damp-heat pouring downward syndrome, and fright injuring the kidney syndrome. The damp-heat pouring downward syndrome belongs to one of the important syndromes in different classification criteria, and its clinical manifestations are mainly characterized by scrotal dampness, urgency and frequency of urination, slippery pulse, and yellow tongue coating.

[0003] Limited by ethics, laws, etc., research on ED still mainly relies on animal models. Therefore, establishing an animal model that conforms to the characteristics of the occurrence and development of ED is the key to deeply exploring its pathogenesis, treatment, and accelerating the transformation of basic research results into clinical applications. Existing ED animal models are mainly divided into two categories according to their causes: Western medicine animal models and TCM syndrome animal models. Among them, the animal models developed for TCM impotence mainly include: kidney yang deficiency type, liver depression type, blood stasis obstruction type (usually using the Western medicine arterial ED model - bilateral internal iliac artery ligation surgery method), but there is no report on the animal model for impotence of the damp-heat pouring downward TCM syndrome type. For the models of TCM damp-heat syndrome, internal damp-heat, external damp-heat, or internal and external damp-heat are usually used for creation, but these models mainly reflect symptoms such as spleen-stomach or lung heat, and cannot reflect the clinical symptoms of pouring damp-heat factors into the bladder, urethra, cervix, or intestine, and even less can simulate impotence of the corresponding syndrome type. Therefore, a simple damp-heat syndrome animal model is not suitable for the research of TCM damp-heat pouring downward impotence, nor is it conducive to the development of new Chinese medicines and the evaluation of drug efficacy for the corresponding syndrome type.

[0004] In view of this, the present invention is specifically proposed. Summary of the Invention

[0005] In order to solve the above technical problems, the present invention provides a method for constructing an animal model of erectile dysfunction with damp-heat pouring downward and its application.

[0006] The technical solution adopted by the present invention is as follows:

[0007] The present invention provides a method for constructing an animal model of erectile dysfunction due to damp-heat pouring downward, comprising the following steps:

[0008] S1. From the day of model establishment, feed the mice with a high-sugar and high-fat diet, and at the same time provide a mixed drinking water containing alcohol and honey, allowing free access to food and water;

[0009] S2. From the 8th day of model establishment, raise the mice in a damp-heat environment;

[0010] S3. On the 15th, 20th, and 25th days of model establishment, subcutaneously inject the mice with a prostate tissue emulsion, and simultaneously intraperitoneally inject a cyclophosphamide solution. Among them, the prostate tissue emulsion is obtained by emulsifying a prostate tissue homogenate and complete Freund's adjuvant in a volume ratio of 1:1;

[0011] S4. Conduct a comprehensive evaluation of the animal model of erectile dysfunction due to damp-heat pouring downward.

[0012] The method for constructing an animal model of erectile dysfunction due to damp-heat pouring downward proposed by the present invention combines internal and external damp-heat factors with drug induction (complete Freund's adjuvant induces prostatitis, and at the same time cyclophosphamide is used to inhibit immune function, induce oxidative stress and reproductive system damage). Multiple mechanisms synergistically simulate the pathogenesis of "damp-heat accumulating in the lower energizer" in traditional Chinese medicine. Moreover, the symptoms shown by the animal model are almost the same as those of clinical patients with impotence due to damp-heat pouring downward, having good scientificity and reliability. The construction method of the present invention fills the blank of the animal model of erectile dysfunction due to damp-heat pouring downward, provides reliable technical support for exploring the mechanism of treating erectile dysfunction due to damp-heat pouring downward, and compared with other traditional modeling methods using lipopolysaccharide or pathogens to cause diseases, it also has characteristics such as a high animal modeling rate and a low animal mortality rate.

[0013] Preferably, in step S1, the high-sugar and high-fat diet is obtained by mixing basal diet, lard, cholesterol, and white sugar in a mass ratio of 70:15:2:13;

[0014] The volume fraction of alcohol and the mass fraction of honey in the mixed drinking water are both 5%.

[0015] Preferably, in step S1, before the model establishment of the mice, pre-screening of the mice is further included, which specifically comprises the following steps: subcutaneously inject the mice with an apomorphine solution at 1 - 2 mg / kg, and screen for qualified mice that show >1 penile erection within 30 minutes after drug administration, and then carry out model establishment;

[0016] The penile head congestion, penile body elongation, and the appearance of the end of the penile body are recorded as one penile erection.

[0017] Preferably, in step S2, the feeding in the damp-heat environment is specifically as follows: The mice are placed in an environment with a temperature of 34-36°C and a relative humidity of 75-90% for 4-6 hours every day.

[0018] Preferably, in step S3, the injection doses of the prostate tissue emulsion and the cyclophosphamide solution are each 10 mL·kg -1 of the mouse body weight, wherein the concentration of the prostate tissue emulsion is 10 mg·mL -1 and the concentration of the cyclophosphamide solution is 8 mg·mL -1 .

[0019] Preferably, in step S3, before mixing and emulsifying the prostate tissue homogenate with the complete Freund's adjuvant, it further includes pre-diluting the prostate tissue homogenate with 1% carrageenan.

[0020] Preferably, in step S4, the comprehensive evaluation indexes of the animal model include traditional Chinese medicine syndrome observation, APO erection test, urination test and mating test.

[0021] More preferably, the test indexes of the traditional Chinese medicine syndrome observation include: appearance, urine and feces, tongue coating;

[0022] The test indexes of the APO erection test include: number of erections within 30 minutes, erection latency;

[0023] The test indexes of the urination test include: total urine volume in 3 hours, urination frequency within 3 hours, average single urine volume;

[0024] The test indexes of the mating test include: licking and sniffing chasing latency, licking and sniffing chasing times, mounting latency, mounting times, ejaculation latency, ejaculation times.

[0025] Preferably, the mice are SPF grade and weigh 20-50 g.

[0026] On the other hand, the present invention provides an application of a damp-heat type erectile dysfunction animal model obtained by the construction method described in any one of the above technical solutions in the efficacy evaluation of traditional Chinese medicines for clearing heat and promoting diuresis.

[0027] The damp-heat type erectile dysfunction animal model established by the method of the present invention conforms to the research of traditional Chinese medicine clinical practice for damp-heat type impotence, and lays a foundation for the research and application in aspects such as the screening of new traditional Chinese medicines and the evaluation of drug efficacy. Brief Description of the Drawings

[0028] Figure 1 It is a comparison chart of the urination test results of each group of mice in Example 1 of the present invention. Among them, A is the total urine volume in 3 hours, B is the urination frequency in 3 hours, and C is the average single urine volume;

[0029] Figure 2 This is a comparison chart of the APO erection test results of mice in each group in Example 1 of the present invention. Among them, A is the number of erections within 30 minutes, and B is the erection latency;

[0030] Figure 3 This is a comparison chart of the mating test results of mice in each group in Example 1 of the present invention. Among them, A is the licking, sniffing, and chasing latency, B is the mounting latency, C is the ejaculation latency, D is the number of licking, sniffing, and chasing times, E is the number of mounting times, and F is the number of ejaculation times. Detailed implementation manners

[0031] In order to more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the following will briefly introduce the drawings required to be used in the embodiments. Obviously, the drawings in the following description are only some embodiments of the present invention. For those of ordinary skill in the art, without creative efforts, other drawings can be obtained based on these drawings.

[0032] One aspect of the present invention provides a method for constructing an animal model of erectile dysfunction due to damp-heat pouring downward, including the following steps:

[0033] S1. Since the day of model establishment, feed mice with a high-sugar and high-fat diet, and at the same time provide a mixed drinking water containing alcohol and honey, allowing free access to food and water;

[0034] S2. Starting from the 8th day of model establishment, raise the mice in a damp-heat environment;

[0035] S3. On the 15th, 20th, and 25th days of model establishment, subcutaneously inject mice with prostate tissue emulsion, and simultaneously intraperitoneally inject cyclophosphamide solution. Among them, the prostate tissue emulsion is obtained by emulsifying a prostate tissue homogenate and complete Freund's adjuvant at a volume ratio of 1:1;

[0036] S4. Conduct a comprehensive evaluation of the animal model of erectile dysfunction due to damp-heat pouring downward.

[0037] In a preferred embodiment of the present invention, in step S1, the high-sugar and high-fat diet is prepared by mixing basal diet, lard, cholesterol, and white sugar at a mass ratio of 70:15:2:13;

[0038] The volume fraction of alcohol and the mass fraction of honey in the mixed drinking water are both 5%.

[0039] In a preferred embodiment of the present invention, in step S1, before establishing the model for mice, it also includes pre-screening the mice, specifically including the following steps: subcutaneously inject mice with 1 - 2 mg / kg of apomorphine solution, and screen for qualified mice that show > 1 penile erection within 30 minutes after drug administration, and then carry out model establishment;

[0040] One penile erection is recorded when the glans penis becomes congested, the penile shaft elongates, and the distal penile shaft appears.

[0041] In a preferred embodiment of the present invention, in step S2, the breeding in a hot and humid environment is specifically as follows: The mice are placed in an environment with a temperature of 34 - 36°C and a relative humidity of 75 - 90% for 4 - 6 hours every day.

[0042] In a preferred embodiment of the present invention, in step S3, the injection doses of the prostate tissue emulsion and the cyclophosphamide solution are 10 mL·kg -1 of the mouse body weight respectively, wherein the concentration of the prostate tissue emulsion is 10 mg·mL -1 and the concentration of the cyclophosphamide solution is 8 mg·mL -1 .

[0043] In a preferred embodiment of the present invention, in step S3, before mixing and emulsifying the prostate tissue homogenate with complete Freund's adjuvant, it further includes pre - diluting the prostate tissue homogenate with 1% carrageenan.

[0044] In a preferred embodiment of the present invention, in step S4, the comprehensive evaluation indexes of the animal model include traditional Chinese medicine syndrome observation, APO erection test, urination test, and mating test.

[0045] In a more preferred embodiment of the present invention, the test indexes of traditional Chinese medicine syndrome observation include: appearance, defecation and urination, tongue coating;

[0046] The test indexes of the APO erection test include: number of erections within 30 minutes, erection latency;

[0047] The test indexes of the urination test include: total urine volume within 3 hours, urination frequency within 3 hours, average single - urination volume;

[0048] The test indexes of the mating test include: licking - sniffing - chasing latency, licking - sniffing - chasing times, mounting - straddling latency, mounting - straddling times, ejaculation latency, ejaculation times.

[0049] In a preferred embodiment of the present invention, the mice are SPF - grade and weigh 20 - 50 g.

[0050] On the other hand, the present invention provides an application of an animal model of erectile dysfunction with damp - heat pouring downward type obtained by the construction method of any one of the above - mentioned technical solutions in the efficacy evaluation of traditional Chinese medicines for clearing heat and promoting diuresis.

[0051] Example 1

[0052] 1. Animal screening

[0053] Ninety SPF male ICR mice (20 - 50 g) were adaptively fed for 1 week, fasted for 12 h, weighed and recorded. A subcutaneous injection of 1.5 mg / kg apomorphine solution (APO) was given to the loose skin on the back of the mouse's neck. The penile erection was recorded when the glans penis became congested, the penile body grew, and the distal penile body appeared. Eligible mice with more than 1 penile erection within 30 min after drug administration were screened for model establishment.

[0054] 2. Animal model establishment

[0055] The eligible test mice were randomly divided into 2 major groups: 8 in the blank control group and 70 in the damp-heat pouring downward type erectile dysfunction model group (abbreviated as the model group). The model group was divided into the damp-heat castration group (denoted as SZ+QS), the damp-heat + complete Freund's adjuvant group (denoted as SZ+Freund's), the damp-heat + complete Freund's adjuvant + cyclophosphamide group (denoted as SZ+Freund's+CP), the damp-heat + complete Freund's adjuvant + prostate tissue homogenate group (denoted as SZ+Freund's+QLX), and the damp-heat + complete Freund's adjuvant + prostate tissue homogenate + cyclophosphamide group (denoted as SZ+Freund's+QLX+CP).

[0056] The model group was given a high-fat and high-sugar diet (by mass percentage, consisting of 70% basal diet, 15% lard, 2% cholesterol, and 13% white sugar), and at the same time, a mixed drinking water containing 5% vol alcohol and 5 wt% honey was provided; the blank control group was given an equal amount of basal diet and pure water.

[0057] After the model group was fed for 1 week, starting from the 8th day of model establishment, they were placed in an external damp-heat environment at a temperature of 35 °C and a relative humidity of 85% for 4 h once a day until the end of the experiment.

[0058] On the 15th, 20th, and 25th days of model establishment, a subcutaneous injection of prostate tissue emulsion with a concentration of 10 mg·mL -1 was given to the mice, and a simultaneous intraperitoneal injection of cyclophosphamide solution with a concentration of 8 mg·mL -1 was given, and the injection dose was 10 mL·kg -1 per mouse body weight.

[0059] The prostate tissue emulsion is obtained by emulsifying the prostate tissue homogenate with complete Freund's adjuvant in a volume ratio of 1:1, and its preparation specifically includes the following steps: taking a male rat, disinfecting the local skin, making a midline incision in the lower abdomen, cutting the rat's abdominal skin and muscles, peeling off the prostate tissue, washing it with physiological saline, and placing it at -80°C for use. The prostate tissue is placed in a centrifuge tube, 20 times the weight of 1% carrageenan is added, and a high-speed tissue crusher (6000rpm, 1min) is used to make a homogenate, and then the prostate tissue homogenate is placed in a high-speed centrifuge for centrifugation (5000rpm, 10min), the supernatant is taken, and 1% carrageenan is added to dilute the mass concentration of the prostate tissue homogenate to 20mg·mL -1， Complete Freund's adjuvant was then added at a volume ratio of 1:1 and emulsified in a tissue homogenizer at 1000 rpm for 20 seconds.

[0060] The preparation of cyclophosphamide solution specifically includes the following steps: taking cyclophosphamide test drug, taking injection saline to prepare a concentration of 8 mg mL -1 Solution.

[0061] 3. Index detection and methods

[0062] 3.1 Observation of TCM Syndrome

[0063] According to the TCM syndrome classification of erectile dysfunction in the Guidelines for the Multidisciplinary Diagnosis and Treatment of Erectile Dysfunction with Integrated Traditional Chinese and Western Medicine (2022 Edition), damp-heat in the lower abdomen belongs to the Zongjin damp-heat syndrome, and the main clinical symptoms are: weak erection, damp scrotum, drowsiness, heavy body, bitter and sticky mouth, burning and short red urine, occasional white drops from the urethra, sticky and uncomfortable stool, red tongue, yellow and greasy fur, and slippery pulse. According to relevant literature, the relevant symptoms of damp-heat syndrome in humans and animals were converted, and the general condition of mice, including appearance, urine, feces, tongue coating, etc., were observed and recorded daily to determine whether the damp-heat syndrome mouse modeling was successful or not, and the degree of symptoms was recorded as 3, 2, 1, and 0, respectively. The symptom scores of mice were recorded at the end of modeling and experiment. (Reference: Li Weiying, Zhang Huiyong, Yan Xiaorui, et al. Effects of Gegenqinlian Decoction on Inflammatory Factors and Intestinal Mucosal Barrier Function in Rats with Damp-Heat Syndrome [J]. Lishizhen Traditional Chinese Medicine, 2023, 34(12): 2856-2860.)

[0064] 3.2 Urinary test

[0065] Refer to the experimental methods of Zhang Xiaowen et al. and Yuan Wenpeng et al., and adopt the urine spot test of water load. Mice were fasted for 12 h without water restriction before the experiment. One hour before the experiment, each group was respectively given normal saline, the water decoction of Liuniao Qiyang, and the aqueous solution of the positive drug by gavage. After 35 min, the abdomen was gently squeezed to drain the remaining urine. First, each group of mice was given 2.0 mL of normal saline (containing 1 mg / mL neutral red dye) by gavage as the water load; 5 min later, the mice were placed in the corresponding breeding cages with 4 layers of clean filter paper laid at the bottom. The weights of the filter paper before and after 3 h were recorded, and the weight increase of the paper was calculated. The mean and standard deviation were calculated, and the P value was calculated by t-test to determine the results. (References: Zhang Xiaowen. Study on the Diuretic Effect of Tongjingcao on Mice [J]. Henan Science and Technology, 2014(24): 73-74. And Yuan Wenpeng, Zhang Shuofeng, Shen Xin, et al. Inhibitory Effect of Piniaoxiaokang on Urination in Rats and Mice and Pharmacokinetic Experiments [J]. Chinese Journal of Experimental Traditional Medical Formulae, 2000(06): 18-20.)

[0066] 3.3 APO Erection Test

[0067] The test mice were placed in a transparent box of 45×45×45 cm and adapted for 10 min. Then, 1.5 mg / kg of apomorphine (APO) was subcutaneously injected at the soft skin on the back of the mouse's neck. The number of penile erections and the erection latency within 30 min were observed and recorded by using a camera (a penile erection was recorded as one time when the glans penis was congested, the penile body grew, and the distal penile body appeared).

[0068] 3.4 Mating Test

[0069] All male mice were given mating training before the mating test. All female mice needed to be prepared for the estrus period. Estradiol was injected at 2 mL·kg -1 3 days before mating, and progesterone was injected at 2 mL·kg -1 4 h before mating. During the test, the male mice were placed in a transparent box of 45 cm×45 cm×45 cm and observed in a quiet and dimly lit room. Starting from 7 pm at night, after each male mouse was placed in the observation cage, it was adapted to the environment for 10 min, and then 1 female mouse was placed in the same cage with it, and the timing was started. The time (latency) and frequency of the male mouse's first licking of the female mouse's genital area within 30 min, the time (i.e., latency) and frequency of the male mouse's first mounting and straddling the female mouse, and the time (latency) and frequency of the male mouse's first ejaculation were recorded. (References: Gan Xiuguo, An Ruihua, Wang Yongquan, et al. Effects of Long-term and Excessive Alcohol Consumption on Sexual Function in Rats (English) [J]. China Journal of Modern Medicine, 2006(19): 2881-2883+2887.)

[0070] 3.5 Scoring Criteria for Each Index

[0071] According to the above primary and secondary clinical symptoms, a scoring standard table for the animal model of erectile dysfunction with damp-heat pouring downward type was formulated and recorded in Table 1.

[0072] Table 1

[0073]

[0074]

[0075] 4. Results Analysis and Evaluation

[0076] 4.1 Observation of Traditional Chinese Medicine Syndromes

[0077] In the mouse model (SZ + Freund's + QLX + CP group) constructed by mixing and injecting the emulsified solution of prostate tissue with damp-heat inside and outside in the embodiment of the present invention, the mice showed symptoms such as listlessness, squinting, dull hair, yellow and greasy tongue coating, loose stools from time to time, and yellow and stinky urine, which conformed to the evaluation criteria of traditional Chinese medicine for the damp-heat syndrome model.

[0078] 4.2 Urination Test

[0079] As can be seen from Figure 1 compared with the blank control group, the urine output of each group of mice within 3 hours decreased significantly, and the difference was extremely significant (P < 0.01). Among them, the SZ + Freund's + QLX + CP group had the best modeling effect and the most significant decrease in urine volume. In the comparison of single-urination volume, the SZ + Freund's + QLX + CP group, SZ + Freund's + QLX group, and SZ + Freund's + CP group had better modeling effects. Compared with the blank control group, the single-urination volume decreased significantly, and the difference was significant (P < 0.01, or t-test, P < 0.05). Among them, the SZ + Freund's + QLX + CP group had the best modeling effect.

[0080] 4.3 APO Erection Test

[0081] As can be seen from Figure 2 compared with the blank control group, the number of erections of each group of mice within 30 minutes decreased significantly, and the difference was extremely significant (P < 0.01). Among them, the SZ + Freund's + QLX + CP group, SZ + Freund's + CP group, and SZ + CP group had better effects. The comparison results of the erection latency showed that the SZ + Freund's + QLX + CP group, SZ + CP group, and SZ + Freund's + CP group had better modeling effects, and the difference was significant compared with the blank control group (P < 0.01).

[0082] 4.4 Mating Test

[0083] As can be seen from Figure 3It can be seen that compared with the blank control group, each modeling method had a significant impact on the licking-sniffing-chasing latency, mounting latency, and number of mountings in mice. In particular, there were extremely significant differences in the number of mountings in each group compared with the blank control group, p < 0.01. In the investigation of the licking-sniffing-chasing latency and mounting latency, the latency in the SZ + Freund's + QLX + CP group was prolonged the longest.

[0084] 4.5 Comparison of the comprehensive scores of each modeling method

[0085] According to the aforementioned test results and scoring criteria, the modeling methods for different types of erectile dysfunction with damp-heat pouring downward were scored and compared. The scores of each group are shown in Table 2 below.

[0086] Table 2

[0087]

[0088] As can be seen from Table 2, the modeling method of the present invention by mixing and injecting the prostate tissue emulsion combined with cyclophosphamide solution according to internal and external damp-heat (SZ + Freund's + QLX + CP group) had the highest score. This method can cause damp-heat to pour downward into the bladder in mice, resulting in erectile dysfunction in mice, successfully obtaining a mouse model of erectile dysfunction with damp-heat pouring downward, and effectively reflecting the clinical characteristics of traditional Chinese medicine erectile dysfunction with damp-heat pouring downward.

[0089] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, not to limit them; although the present invention has been described in detail with reference to the foregoing embodiments, those of ordinary skill in the art should understand that they can still modify the technical solutions described in the foregoing embodiments, or perform equivalent replacements for some or all of the technical features; and these modifications or replacements do not make the essence of the corresponding technical solutions deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. A method for constructing an animal model of erectile dysfunction caused by damp heat, characterized in that: The following steps are involved: S1. From the day of modeling, mice were fed with a high-sugar and high-fat diet and provided with mixed drinking water containing alcohol and honey, and free access to food and water; S2. From the 8th day of modeling, the mice were kept in a hot and humid environment; S3. On the 15th, 20th and 25th days of modeling, mice were subcutaneously injected with prostate tissue emulsion and simultaneously intraperitoneally injected with cyclophosphamide solution, wherein the prostate tissue emulsion was obtained by emulsifying prostate tissue homogenate and complete Freund's adjuvant in a volume ratio of 1:1; S4. Conduct a comprehensive evaluation of the animal model of damp-heat-induced erectile dysfunction.

2. The method for constructing an animal model of damp-heat injection type erectile dysfunction as claimed in claim 1, characterized in that: In step S1, the high-sugar and high-fat feed is obtained by mixing basic feed, lard, cholesterol and white sugar in a mass ratio of 70:15:2:13; The volume fraction of alcohol and the mass fraction of honey in the mixed drinking water are both 5%.

3. The method for constructing an animal model of erectile dysfunction caused by damp heat injection as claimed in claim 1, characterized in that: In step S1, before the mouse model is established, the mice are pre-screened, specifically including the following steps: subcutaneously injecting 1-2 mg / kg apomorphine solution into the mice, screening qualified mice that have >1 penile erection within 30 minutes after administration, and conducting model establishment; The engorgement of the glans penis, the growth of the penis shaft and the appearance of the terminal penis shaft were recorded as one penile erection.

4. The method for constructing an animal model of erectile dysfunction caused by damp heat injection as claimed in claim 1, characterized in that: In step S2, the feeding in the humid and hot environment is specifically: exposing the mice to an environment with a temperature of 34-36° C. and a relative humidity of 75-90% for 4-6 hours every day.

5. The method for constructing an animal model of erectile dysfunction caused by damp heat injection as claimed in claim 1, characterized in that: In step S3, the injection doses of the prostate tissue emulsion and the cyclophosphamide solution are 10 mL·kg each time. -1 Mouse weight, wherein the concentration of the prostate tissue emulsion is 10 mg·mL -1 The concentration of the cyclophosphamide solution is 8 mg mL -1 .

6. The method for constructing an animal model of erectile dysfunction caused by damp heat injection as claimed in claim 1, characterized in that: In step S3, before the prostate tissue homogenate is mixed and emulsified with the complete Freund's adjuvant, the prostate tissue homogenate is diluted with 1% carrageenan in advance.

7. The method for constructing an animal model of erectile dysfunction caused by damp heat injection as claimed in claim 1, characterized in that: In step S4, the comprehensive evaluation indexes of the animal model include TCM syndrome observation, APO erection test, urination test and mating test.

8. The method for constructing an animal model of erectile dysfunction caused by damp heat injection as claimed in claim 7, characterized in that: The test indicators for TCM syndrome observation include: appearance, stool and urine, and tongue coating; The test indicators of the APO erection test include: the number of erections within 30 minutes, the erection latency period; The test indicators of the urination test include: total urination volume within 3 hours, urination frequency within 3 hours, and average single urination volume; The inspection indicators of the mating test include: licking and sniffing pursuit latency, licking and sniffing pursuit times, back climbing and mounting latency, back climbing and mounting times, ejaculation latency, and ejaculation times.

9. The method for constructing an animal model of erectile dysfunction caused by damp heat injection as claimed in claim 1, characterized in that: The mice were of SPF grade and weighed 20-50 g.

10. Use of an animal model of erectile dysfunction of damp-heat type obtained by the construction method according to any one of claims 1 to 9 in the efficacy evaluation of traditional Chinese medicines for clearing away heat and dampness.

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