CN112546118B - A kind of traditional Chinese medicine composition for treating the disease of repeated embryo implantation failure and its application - Google Patents

Abstract

The invention relates to a traditional Chinese medicine composition for treating embryo repeated planting failure diseases, which is prepared from the following raw material medicines in parts by weight: 29-31 parts of amethyst, 14-16 parts of semen cuscutae, 14-16 parts of prepared rehmannia root, 11-13 parts of caulis spatholobi, 14-16 parts of parasitic loranthus, 8-10 parts of red paeony root, 14-16 parts of angelica sinensis, 14-16 parts of radix paeoniae alba, 8-10 parts of ligusticum wallichii, 14-16 parts of bighead atractylodes rhizome, 14-16 parts of codonopsis pilosula, 14-16 parts of poria cocos, 11-13 parts of eucommia ulmoides, 11-13 parts of teasel root and 8-10 parts of dried orange peel. The invention also provides the application of the traditional Chinese medicine composition. The traditional Chinese medicine composition has the advantages of quick curative effect, short treatment course and obvious effect on diseases caused by repeated embryo planting failure; in addition, has no toxic and side effects on human bodies.

Description

A kind of traditional Chinese medicine composition for treating the disease of repeated embryo implantation failure and its application

technical field

The invention relates to the technical field of hospitals, in particular, to a traditional Chinese medicine composition for treating the disease of repeated embryo implantation failure and its application.

Background technique

Repeated transfer failure (RIF) refers to at least 3 consecutive in vitro fertilization-embryo transfer (IVF-ET) cycles of embryo implantation failure, and there are 1-2 high-quality embryos in each cycle. RIF has gradually become a major problem affecting IVF pregnancy rates. Endometrial receptivity means that the endometrium allows embryos to implant only during a certain period of time, which is called the "implantation window". A receptive endometrial environment is to ensure embryo implantation, growth and development. important factor. Embryo quality and endometrial receptivity are two important factors that affect the outcome of transplantation, and 2/3 of repeated transplantation failures are related to low endometrial receptivity. At present, there is no effective targeted treatment for RIF. Freezing-thawed embryo transfer after whole embryo freezing can artificially control the timing of the opening of the endometrial “implantation window”, so that the development of the endometrium and the embryo can be synchronized. It is a common treatment for RIF.

Chinese patent document CN110585369A discloses a traditional Chinese medicine composition for improving endometrial receptivity and its application. Including 15-25 parts of raw astragalus, 25-35 parts of dodder seeds, 15-25 parts of antler cream, 5-15 parts of Cistanche deserticola, 5-15 parts of Morinda officinalis, 10-20 parts of Poria, 5-15 parts of fried Atractylodes, 3 parts of Atractylodes -8 servings, 15-25 servings of fried yiren, 15-25 servings of psyllium, 5-15 servings of dried tangerine peel, 5-15 servings of lotus root, 5-15 servings of Huoxiang, and 3-8 servings of Treats. Chinese patent document CN109954049A discloses a traditional Chinese medicine composition for repairing endometrium and assisting pregnancy. It is made of Shudi, Salvia miltiorrhiza, Dodder, Xianlingpi, Morinda officinalis, Baked Turtle, Poria, Fried Chibaishao, Angelica sinensis, and Phyllostachys chinensis. But about curative effect is quick, course for the treatment of is short, effect is remarkable; The traditional Chinese medicine composition of the treatment embryo repeated implantation failure disease without toxic and side effects to human body has not yet seen report at present.

SUMMARY OF THE INVENTION

The first object of the present invention is to, for the deficiencies in the prior art, provide a kind of Chinese medicine composition for the treatment of repeated embryo implantation failure disease.

The second object of the present invention is to provide the use of the traditional Chinese medicine composition.

In order to realize the above-mentioned first purpose, the technical scheme adopted in the present invention is: a kind of Chinese medicine composition for the treatment of the disease of repeated implantation failure of embryos, and the described Chinese medicine composition is made of the following raw materials by weight: amethyst 29 -31 parts, 14-16 parts of dodder, 14-16 parts of Rehmannia glutinosa, 11-13 parts of Radix japonica, 14-16 parts of mulberry parasite, 8-10 parts of red peony root, 14-16 parts of angelica root, 14-16 parts of white peony root parts, Chuanxiong 8-10 parts, Atractylodes Rhizoma 14-16 parts, Codonopsis 14-16 parts, Poria 14-16 parts, Eucommia 11-13 parts, Continuum 11-13 parts, Chenpi 8-10 parts.

A further technical scheme of the present invention: the traditional Chinese medicine composition is made from the following raw materials by weight: 30 parts of amethyst, 15 parts of dodder, 15 parts of Rehmannia glutinosa, 12 parts of Radix Glycyrrhiza, 15 parts of mulberry and 9 parts of red peony , 15 parts of Angelica, 15 parts of Baishao, 9 parts of Chuanxiong, 15 parts of Atractylodes, 15 parts of Codonopsis Radix, 15 parts of Poria, 12 parts of Eucommia, 12 parts of truncation, 9 parts of tangerine peel

In order to realize the above-mentioned second purpose, the technical scheme adopted in the present invention is: the described disease is the disease of repeated implantation failure of embryos due to kidney deficiency and blood stasis.

Further technical scheme of the present invention: the described disease is the disease of repeated implantation failure of embryos due to yang deficiency and blood stasis.

Further technical scheme of the present invention: the described disease is the disease of repeated implantation failure of embryos due to yang deficiency and blood stasis.

The prescription of the present invention is solved: the whole prescription adds the traditional Chinese medicines of promoting blood circulation and removing blood stasis, strengthening the spleen and nourishing qi, nourishing the kidney and relieving the miscarriage on the basis of warming the kidney yang, which is suitable for infertile patients with kidney deficiency and partial cold, especially suitable for patients with repeated embryo implantation failure. In the prescription, amethyst is sweet and warm, enters the heart and liver meridians, and treats women with blood deficiency and cold infertility. The "Benjing" records: "Insufficient tonic, women's cold in the uterus, sterilized for ten years without children." Amethyst is the king, warms the kidneys and yang, warms the uterus, warms the cold, and nourishes the middle. In addition, amethyst can supplement the central qi, benefit the heart and liver, and unclog blood vessels, so it is an essential medicine for people with restless mind, lack of liver blood and infertility in women with blood sea cold deficiency. Dodder, mulberry parasite, Eucommia ulmoides, and eucommia are the ministerial medicines. Dodder tonifies the kidney and nourishes the essence, and the kidney is prosperous. It can nourish the fetus; Eucommia warms the uterus, soothes the fetus, and combines four herbs to warm the kidney yang, nourish the kidney yin, invigorate the kidney qi, and strengthen the muscles and bones, so that the embryo is stable, and it is not easy to slip the fetus. Chixueteng nourishes blood and activates blood and red peony root for dispelling blood stasis and relieving pain are the main herbs of the prescription. When it is solid to warm the kidney, add an appropriate amount of blood-activating products to the purpose of "removing blood stasis and creating new", and at the same time, it is consistent with the mechanism of improving the body's microcirculation advocated by Western medicine theory. Rehmannia glutinosa, Angelica sinensis, Chuanxiong and Baishao are combined into Siwu Decoction to nourish blood and blood, regulate meridians and remove blood stasis. Traditional Chinese medicine believes that the kidney is the foundation of the innate, the viscera of the essence and blood, and controls reproduction. The deficiency of kidney-yang qi and the depletion of essence and blood make it difficult to conceive. The two sides are combined into Bazhen Decoction as an adjuvant, which makes Qi and blood active, and nourishes the kidney and relieves the miscarriage. In the end, tangerine peel is used as an adjuvant, which has the power of regulating qi and strengthening the spleen. The combination of various medicines can warm the kidney yang, invigorate the spleen and qi, invigorate the kidney and relieve the miscarriage, promote blood circulation and remove blood stasis, and together achieve the power of helping pregnancy.

The advantages of the present invention are: based on the overall concept of traditional Chinese medicine, the present invention gives full play to the advantages of TCM syndrome differentiation and treatment, treats different etiologies from multiple perspectives, combines deficiency and actuality, treats both the symptoms and the symptoms, and selects for the compatibility of various medicines, warming the kidney yang, strengthening the spleen and nourishing qi, and invigorating the kidney. According to the theory of traditional Chinese medicine, the traditional Chinese medicine composition is prepared according to the specific weight proportion. The traditional Chinese medicine composition has fast curative effect, short course of treatment and remarkable effect on diseases of repeated embryo implantation failure. ; In addition, it has no toxic and side effects on the human body.

Description of drawings

Figure 1 shows the morphology of endometrial tissue observed by HE staining.

Detailed ways

The present invention will be further described below in conjunction with specific embodiments. It should be understood that these examples are only used to illustrate the present invention and not to limit the scope of the present invention. In addition, it should be understood that those skilled in the art can make various changes or modifications to the present invention after reading the contents recorded in the present invention, and these equivalent forms also fall within the scope defined by the appended claims of the present application.

Embodiment 1 Traditional Chinese medicine composition (1) for treating the disease of repeated embryo implantation failure

According to the following proportions by weight, the raw materials are obtained:

30 parts of amethyst, 15 parts of dodder, 15 parts of Rehmannia glutinosa, 12 parts of Scarlet vine, 15 parts of mulberry parasite, 9 parts of red peony, 15 parts of angelica, 15 parts of white peony, 9 parts of Chuanxiong, 15 parts of Atractylodes, 15 parts of Codonopsis , 15 parts of Poria, 12 parts of Eucommia, 12 parts of continuation and 9 parts of dried tangerine peel.

Embodiment 2 treats the Chinese medicine composition (two) of the disease of repeated embryo implantation failure

According to the following proportions by weight, the raw materials are obtained:

29 parts of amethyst, 16 parts of dodder, 14 parts of Rehmannia glutinosa, 13 parts of Radix Glycyrrhizae, 14 parts of mulberry, 8 parts of red peony, 14 parts of angelica, 16 parts of white peony, 8 parts of Chuanxiong, 14 parts of Atractylodes, 16 parts of Codonopsis , 16 parts of Poria, 11 parts of Eucommia, 11 parts of continuation and 8 parts of dried tangerine peel.

Embodiment 3 Chinese medicine composition (three) for treating the disease of repeated embryo implantation failure

According to the following proportions by weight, the raw materials are obtained:

31 parts of amethyst, 14 parts of dodder, 16 parts of Rehmannia glutinosa, 11 parts of Scutellaria, 16 parts of mulberry, 10 parts of red peony, 16 parts of angelica, 14 parts of white peony, 10 parts of Chuanxiong, 16 parts of Atractylodes, 14 parts of Codonopsis , 14 parts of Poria, 13 parts of Eucommia, 13 parts of continuation and 10 parts of dried tangerine peel.

Embodiment 4 The Chinese medicine composition (four) of the treatment of embryo repeated implantation failure disease

According to the following proportions by weight, the raw materials are obtained:

30 parts of amethyst, 16 parts of dodder, 16 parts of Rehmannia glutinosa, 13 parts of Scutellaria, 15 parts of mulberry, 8 parts of red peony, 14 parts of angelica, 15 parts of white peony root, 9 parts of Chuanxiong, 15 parts of Atractylodes, 14 parts of Codonopsis , 14 parts of Poria, 13 parts of Eucommia, 13 parts of continuation and 8 parts of dried tangerine peel.

Embodiment 5 The traditional Chinese medicine composition (5) of the treatment of embryo repeated implantation failure disease

According to the following proportions by weight, the raw materials are obtained:

29 parts of Amethyst, 14 parts of Cuscuta, 15 parts of Rehmannia glutinosa, 11 parts of Scutellaria, 16 parts of mulberry, 10 parts of red peony, 16 parts of Angelica, 16 parts of white peony, 8 parts of Chuanxiong, 14 parts of Atractylodes, 16 parts of Codonopsis , 15 parts of Poria, 11 parts of Eucommia, 12 parts of continuation and 10 parts of dried tangerine peel.

Embodiment 6 The traditional Chinese medicine composition (six) of the treatment of embryo repeated implantation failure disease

According to the following proportions by weight, the raw materials are obtained:

31 parts of amethyst, 15 parts of dodder, 14 parts of Rehmannia glutinosa, 12 parts of Radix Glycyrrhiza, 14 parts of mulberry, 9 parts of red peony, 15 parts of angelica, 14 parts of white peony, 10 parts of Chuanxiong, 16 parts of Atractylodes, 15 parts of Codonopsis , 16 parts of Poria, 12 parts of Eucommia, 11 parts of continuation and 9 parts of dried tangerine peel.

Embodiment 7 Chinese medicine composition (seven) for the treatment of the disease of repeated embryo implantation failure

According to the following proportions by weight, the raw materials are obtained:

29 parts of Amethyst, 14 parts of Dodder Seed, 14 parts of Rehmannia glutinosa, 13 parts of Scarlet Root, 15 parts of Mulberry, 9 parts of Red Peony, 14 parts of Angelica, 16 parts of White Peony, 8 parts of Chuanxiong, 14 parts of Atractylodes Rhizoma, 16 parts of Codonopsis , 14 parts of Poria, 13 parts of Eucommia, 12 parts of continuation and 8 parts of dried tangerine peel.

Embodiment 8 Traditional Chinese medicine composition (8) for treating the disease of repeated embryo implantation failure

According to the following proportions by weight, the raw materials are obtained:

31 parts of amethyst, 15 parts of dodder, 16 parts of Rehmannia glutinosa, 11 parts of Scutellaria, 16 parts of mulberry, 8 parts of red peony, 16 parts of angelica, 14 parts of white peony root, 10 parts of Chuanxiong, 16 parts of Atractylodes, 14 parts of Codonopsis , 16 parts of Poria, 12 parts of Eucommia, 11 parts of continuation and 10 parts of dried tangerine peel.

Embodiment 9 Chinese medicine composition (nine) for the treatment of the disease of repeated embryo implantation failure

According to the following proportions by weight, the raw materials are obtained:

30 parts of Amethyst, 16 parts of Cuscuta, 15 parts of Rehmannia glutinosa, 13 parts of Radix Glycyrrhiza, 14 parts of mulberry, 10 parts of red peony, 14 parts of Angelica, 15 parts of white peony, 9 parts of Chuanxiong, 15 parts of Atractylodes, 15 parts of Codonopsis , 15 parts of Poria, 11 parts of Eucommia, 13 parts of continuation and 9 parts of dried tangerine peel.

Embodiment 10 The traditional Chinese medicine composition (ten) of treating the disease of repeated embryo implantation failure

According to the following proportions by weight, the raw materials are obtained:

31 parts of amethyst, 15 parts of dodder, 16 parts of Rehmannia glutinosa, 12 parts of Scutellaria, 15 parts of mulberry, 10 parts of red peony, 15 parts of angelica, 16 parts of white peony, 9 parts of Chuanxiong, 14 parts of Atractylodes, 16 parts of Codonopsis , 15 parts of Poria, 13 parts of Eucommia, 13 parts of continuation and 8 parts of dried tangerine peel.

Example 11

1 Research object

All cases were from patients who underwent in vitro fertilization and frozen embryo transfer in the Department of Reproductive Medicine, Shanghai First Maternity and Infant Health Hospital from January 2019 to September 2019.

2 case selection

2.1 Diagnostic criteria

Diagnostic criteria of Western medicine: in line with IVF-ET/ICSI indications:

1) tubal infertility;

2) Endometriosis;

3) Male factor infertility;

4) Abnormal ovulation;

5) Unexplained infertility;

6) Cervical factors, etc.

TCM syndrome differentiation criteria for kidney (yang) deficiency and blood stasis:

1) Main symptoms: prolonged infertility, late menstruation or sparse hair, amenorrhea if the amount is small, pale color, or lumps;

2) Secondary symptoms: dizziness and tinnitus, soreness and coldness of waist and knees, cold lower abdomen, nocturnal urination, excessive vaginal discharge, thin quality, dark eyes, dark spots on the face, apathy of libido, mental fatigue;

3) Tongue pulse: dull tongue, white coating, thin or slow pulse;

4) There are 2 main symptoms and 1-2 secondary symptoms, which can be diagnosed by referring to the tongue and pulse.

2.2 Inclusion criteria

1) Meet the indications for entering the IVF-ET cycle and the TCM dialectical criteria for kidney deficiency and blood stasis syndrome;

2) Age 25-45 years old;

3) Embryo transfer failed for three or more pregnancies;

4) Those who have cryopreserved high-quality embryos (≥2) in our center;

5) Those who have no uterine cavity lesions or have been cured by hysteroscopy and other surgeries;

6) Those who have no hydrosalpinx or have been surgically ligated;

7) Sign the informed consent form.

2.3 Exclusion criteria

1) Diseases of the uterus and uterine cavity that have not been treated or cured by hysteroscopy, such as uterine malformations, uterine fibroids, endometrial polyps, endometritis, intrauterine adhesions, uterine fibrosis, adenomyosis, etc. ;

2) Those with hydrosalpinx and no surgical treatment;

3) Those who cannot cooperate, such as those with neurological and mental disorders;

4) Those with incomplete information;

5) Those who are allergic to the drugs used;

6) Participating in other clinical investigators.

2.4 Elimination and shedding standards

1) Those who cannot continue treatment due to various reasons;

2) Those with adverse drug reactions;

3) Those with concurrent diseases;

4) Those who withdraw from the test process by themselves.

3 treatment options

3.1 Random grouping

80 selected RIF patients underwent endometrial preparation, and SPSS 21.0 statistical software was used to generate random numbers according to the number of cases. According to the order of admission, they were divided into a traditional Chinese medicine group and a control group, with 40 cases in each group.

3.2 Treatment methods

(1) Endometrial preparation: The subjects entered the week on the 2nd-3rd day after the menstrual cycle or drug withdrawal bleeding, and on the same day after the ovarian cyst was excluded by the first B-ultrasound, Fenmotonghong tablets (estradiol 2mg, Abbott, Netherlands, approval number H20110159) 4 mg, twice a day, after oral administration for 12 consecutive days, a second B-ultrasound was performed to monitor the thickness of the intima. (Estradiol/Dydrogesterone 2mg, Abbott, Netherlands, Approval No. H20110159), 4mg, 2 times a day, continuous oral administration for 3 days for endometrial transformation, the day is designated as the day of endometrial transformation, 3 days after transformation Cleavage-stage embryos can be transferred or blastocysts can be transferred 5 days after transformation; if the subject's second B-ultrasound monitoring endometrial thickness is less than 8mm, proganol (estradiol valerate 1mg, Bayer, Germany, Approval number J20130009) 1mg, once a day, vaginal administration. Estrogen is taken for a maximum of 3 weeks, and if the endometrium is still <8cm, the cycle will be cancelled.

(2) Luteal support: after frozen embryo transfer, Fenmotonghuang tablets (estradiol/dydrogesterone) 4mg, 2 times a day, Angelan (progesterone soft capsule 0.1g, Bessen, Belgium, approval number) H20130336) 0.2g, 2 times a day, vaginal medication.

(3) Traditional Chinese medicine group: The subjects started to take the embodiment 1 of the present invention orally on the 2nd to 3rd day after the menstrual cycle or drug withdrawal bleeding on the basis of the routine endometrial preparation and luteal support program. Drug composition: purple Quartz 30g, Dodder Seed 15g, Rehmannia glutinosa 15g, Sophora japonica 12g, Mulberry parasitic 15g, Red peony 9g, Angelica 15g, White peony root 15g, Chuanxiong 9g, Atractylodes 15g, Codonopsis 15g, Poria 15g, Eucommia ulmoides 12g, Interrupted 12g, dried tangerine peel 9g, (granule preparation: the Chinese medicine composition of embodiment 1 is continuously decocted 3 times, and the water addition of each decoction is 10 times the weight of the Chinese medicine composition, and each decoction time is 3h, after collecting 3 times of decoction filtrate; concentrate the filtrate to a relative density of 1.33 at 60°C, dry and pulverize to obtain extract powder; add starch and dextrin to extract powder to make granules). The granules were taken once a day and continued to be taken until 14 days after embryo transfer.

(4) Control group: Frozen embryo transfer was performed only with conventional endometrial preparation + luteal support program.

3.3 Observation indicators and methods

3.3.1 General information: age of the patient, years of infertility, type of infertility, infertility factors, number of previous transplants, basal hormone levels, and basal endometrial status.

3.3.1 Comparison of clinical pregnancy rate and implantation rate

1) Clinical pregnancy rate = (number of clinical pregnancy cycles/total number of transplant cycles) × 100%

2) Implantation rate = (number of implanted embryos/total number of embryos transferred) × 100%

3.3.2 Endometrial vaginal ultrasound indicators:

1) Endometrial thickness on the day of endometrial transformation: instruct the subject to lie supine on the examination bed, take the bladder lithotomy position, cover the surface of the vaginal probe with a disposable condom and slowly put it into the subject's vagina, and turn the vaginal probe until it is clear The sagittal plane of the uterus is shown, and the measurement standard of endometrial thickness is the distance between the interface between the anterior and posterior myometrium of the uterus and the endometrium from about 2cm below the fundus of the uterus.

2) Endometrial type on the day of endometrial transformation: Gonen classification criteria are used: Type A: typical triline, the junction of the endometrium and the myometrium and the central endometrial line are all hyperechoic, and between the outer layer and the central line The area is hypoechoic; Type B: Weak triline type, the midline of uterine cavity echo is not obvious or intermittent; Type C: The midline of uterine cavity is homogeneous and strong echo without strong echo.

3) Endometrial blood flow on the day of endometrial transformation Uterine spiral artery blood flow parameters: vascular pulsatility index (PI) and vascular resistance index (RI).

4) Endometrial blood flow on the day of endometrial transformation: the quantification standard of subendometrial blood flow by Gao Minzhi, etc. was used. Transvaginal ultrasound showed the longitudinal section of the uterus, and the most small blood vessels were selected for counting, ranging from the endometrium and the anterior and posterior muscles. The area between the interfaces of the layers. Subintimal blood flow is divided into 4 grades:

Type I: There is no obvious blood flow signal in the intimal region, and the number of blood flow branches is 0;

Type II: Sparse blood flow signals are seen in the intima area, and the number of blood flow branches is ≤ 2;

Type III: There are many blood flow signals in the intima area, and the number of blood flow branches is > 2 and ≤ 4;

Type IV: rich blood flow signals are seen in the intima area, and the number of blood flow branches is greater than or equal to 5;

4 results

4.1 Baseline Comparison

After statistical analysis, the data between the two groups were balanced and comparable in terms of age, infertility course, infertility factors, infertility type, number of transplants, FET preparation plan, basal endometrial thickness, and basal hormone levels (P> 0.05).

4.2 Comparison of clinical pregnancy rate and miscarriage rate

There was no significant difference in the clinical pregnancy rate between the two groups (p>0.05). After stratifying the patients by age, the pregnancy rate in the traditional Chinese medicine group was higher than that in the control group (44.8% VS 18.5%), and the difference was statistically significant. Significance (p<0.05). The abortion rate in the traditional Chinese medicine group was significantly lower than that in the control group (2.5% VS 17.5%), and the difference was statistically significant (p<0.05).

Comparison of clinical pregnancy rate between two groups [cases (%)]

Comparison of clinical pregnancy rate of patients <35 years old between two groups [cases (%)]

Comparison of miscarriage rates between the two groups [cases (%)]

4.3 Endometrial Vaginal Ultrasound Results

(1) Endometrial thickness

After treatment, the endometrial thickness of the traditional Chinese medicine group was higher than that of the control group, and there was a statistical difference (p<0.05)

Endometrial thickness of each group on the day of endometrial transformation

Note: Compared with the control group, ^★ p<0.05.

(2) Comparison of endometrial types

After treatment, the proportion of type A intima in the traditional Chinese medicine group was the highest, and there was a statistical difference compared with the control group (p<0.05).

Comparison of endometrial types in each group on the day of transplantation [cases (%)]

Note: Compared with the control group, ^★ p<0.05.

(3) Comparison of endometrial blood flow

Comparison of the number of subendometrial blood flow on the day of endometrial transformation after treatment showed that type III and IV endometrium in the traditional Chinese medicine group increased compared with the control group. The difference was statistically significant (p<0.05).

Comparison of the number of endometrial blood flow on the day of transformation [case (%)]

Note: Compared with the control group, ^★ p<0.05.

Example 12

1. Methods and results of animal experiments

1 material

1.1 Animals

Clean-grade sexually mature nulliparous female Sprague Dawley rats weighing 220-250 g and clean-grade sexually mature nulliparous male Sprague Dawley rats weighing 300-330 g. Provided by the Animal Experiment Center of Shanghai University of Traditional Chinese Medicine, license: SCXK (Beijing) 2016-0011.

1.2 Drugs

Embodiment 3 of the present invention. Drug Specifications: Amethyst 31g, Dodder Seed 14g, Rehmannia glutinosa 16g, Chixueteng 11g, Mulberry Parasitic 16g, Red Peony 10g, Angelica 16g, White Peony 14g, Chuanxiong 10g, Atractylodes 16g, Codonopsis 14g, Poria 14g, Eucommia 13g, continued Broken 13g, dried tangerine peel 10g, decocted in water and concentrated into "low-dose decoction" (concentration of crude drug 1.17g/ml, equivalent to 10 times the human dose), "high-dose decoction" (concentration of crude drug 2.33g/ml, equivalent to 20 times the human dose).

Mifepristone Tablets: Shanghai New Hualian Pharmaceutical Co., Ltd. (25 mg C019141008).

Progesterone injection: Shanghai General Pharmaceutical Co., Ltd. (20mg 160706).

2 methods

2.1 Observation of estrous cycle and cage

Rat vaginal smears were made at 8:00 AM every day, and items were prepared: ophthalmic cotton swabs, glass slides, and normal saline. Place the female rat on the operating table, press the body of the rat with the palm of the hand, hold the tail of the rat with the thumb and index finger and gently lift it up, wipe the sawdust, urine and feces around the vaginal opening with the other hand, and dip a small amount of cotton swab After the physiological saline is placed into the vagina, it sticks to the vaginal wall and turns for 3-4 circles, and then evenly spreads the vaginal secretions on the cotton swab on the surface of the slide glass, and air-dries naturally. The vaginal smear was observed with an optical microscope to judge the estrous cycle of the rats. Proestrus: Microscopically, a large number of oval epithelial cells (with distinct nuclei) and a small number of polygonal keratinocytes (without nuclei) can be seen. Duration: 17-21 hours; Estrus: Nucleated keratinocytes are predominant in number, deciduous stacks, and a few oval epithelial cells are visible. The duration is 9-15 hours; in the post-estrus stage: nucleated epithelial cells, non-nucleated keratinocytes, and leukocytes coexist, and a large number of granular leukocytes gather around polygonal keratinocytes. Duration 10-14 hours; interestrus: the overall number of cells in this phase is low, mostly white blood cells, and occasionally a small number of epithelial cells can be seen. Duration 60-70 hours. Pre-estrus and estrus female mice were selected to be caged with male mice of the same strain at 6:00 PM on the same day, and the cage ratio was 1:1. The vaginal smear was observed again at 8:00 AM the next day. If sperm was seen on the smear, it was recorded as pregnancy. Day 1 (P D1).

2.2 Grouping and modeling

PD1 rats were grouped by completely randomized grouping method. First, the PD1 female rats were weighed and numbered according to their body weight; secondly, a column of random numbers was generated by SPSS22.0 software, and the random numbers were arranged from small to large; finally, From the sorted random numbers, the numbers corresponding to the first 10 random numbers are specified as the traditional Chinese medicine group (group C), the numbers corresponding to the 11th-20th random numbers are the electroacupuncture group (group P), and the numbers corresponding to the 21st-30th random numbers The corresponding numbers are the acupuncture and medicine group (group M), and the numbers corresponding to the 31st to 40th random numbers are the progesterone group (group N).

Modeling:

The Huang Guangying rat embryo implantation disorder model was improved. Models were established by gavage of mifepristone. Mifepristone + saline suspension (1 mg/ml) was prepared 1 day in advance. The PD3 rats were modeled by gavage of mifepristone suspension, and the gavage dose was 8ml/kg.

2.3 Intervention Program

Group C: According to the conversion principle of body shape coefficient, the concentration of crude drug in the traditional Chinese medicine was 0.9g/ml, and the patients were given intragastric administration at 8AM on PD1-7.

Group P: intramuscular injection of progesterone 2 mg after 9 am on day 1-7 of pregnancy.

Group M: 2ml of normal saline was administered by gavage at 8AM on days 1-7 of pregnancy.

Group N: 2ml of normal saline was given by gavage at 8AM on day 1-7 of pregnancy.

Materials: On the 8th day of PD, after anesthesia at 4PM, the bilateral uterus, fallopian tubes, and ovaries of the rats were dissected (pay attention to removing the mesentery as much as possible, and swab the blood stains with gauze), record the number of pregnant mice, and count the number of implantation sites. Preservation of uterine tissue in sections: Take 0.5cm tissue sections and place them in 4% paraformaldehyde to fix for 48 hours for immunohistochemical detection and HE staining; part of the tissue is removed from embryos and then packed in 2ml cryopreservation tubes. Freeze at 80°C for RT-PCR, Western-blot and ELISA. .

2.4 Observation indicators and methods

2.4.1 Average number of implantation sites: Average number of implantation sites=(total number of implantation sites/number of pregnant mice)×100%. 2.4.2 HE staining to observe the morphology of endometrial tissue

3 results

3.1 Average implantation site results

The pregnancy rate of rats in each PD8 group was higher in group C than in group M (50% vs 17%), and there was a statistical difference (p<0.05). The comparison between the implantation groups of PD8 rats in group C was higher than that in group M, and the difference was statistically significant (p<0.05).

3.2 The morphological comparison of endometrial tissue is shown in Figure 1

The specific manifestations of the loss of decidualization in PD8 rats in the model group are that the uterine cavity is not closed, the epithelial cells of the endometrial cavity are still single-layer columnar, the primary decidualization area is missing under the epithelium and in the stromal layer, and the uterine glands are present, but the glands are not. The body number is small, the gland cavity is small and short. Platelets aggregated in some capillaries, suggesting the possibility of thrombosis. The uterine cavity of PD8 rats in the normal group gradually closed, the number of epithelial cells decreased, the primary decidualized area had appeared in the stromal layer, and the secondary decidualized area with abundant blood vessels gradually appeared on the proximal mesentery side. Endometrial stromal cells were composed of spindles. The fibroblasts differentiated into round-like decidual cells with larger nuclei or binuclear or multinucleated cells. The cytoplasm of decidual cells was darkened due to accumulation of glycogen, and extensive decidualization of the endometrial layer resulted in the extrusion of the uterine glands to the endometrial base. The results of this study showed that the morphology of PD8 endometrium of rats in group P was similar to that in group M, the uterine cavity was not closed, the decidualization was insufficient, and some thrombosis was formed. The morphology of PD8 endometrium in group C was similar to that in group N, and extensive decidualization had appeared.

Example 13

Comparative Example 1

This comparative example is a traditional Chinese medicine composition, which is basically the same as the traditional Chinese medicinal composition in Example 4, except that the amethyst is replaced with Morinda officinalis of the same fraction.

Comparative Example 2

This control example is a traditional Chinese medicine composition, which is basically the same as the traditional Chinese medicine composition in Example 4, the difference is only that the saffron of the same fraction is replaced by S. chinensis.

1 Research object

All cases were from patients who underwent in vitro fertilization and frozen embryo transfer in the Department of Reproductive Medicine, Shanghai First Maternity and Infant Health Hospital from January 2018 to September 2018.

2 case selection

2.1 Diagnostic criteria

Diagnostic criteria of Western medicine: in line with IVF-ET/ICSI indications:

7) tubal infertility;

8) Endometriosis;

9) Male factor infertility;

10) Abnormal ovulation;

11) Unexplained infertility;

12) Cervical factors, etc.

TCM syndrome differentiation criteria for kidney (yang) deficiency and blood stasis:

5) Main symptoms: long-term infertility, late menstruation or sparse hair, amenorrhea if the amount is very small, pale color, or lumps;

6) Secondary symptoms: dizziness and tinnitus, soreness and coldness of waist and knees, cold lower abdomen, nocturnal urination, excessive vaginal discharge, thin quality, dark eyes, dark spots on the face, apathy of libido, mental fatigue;

7) Tongue pulse: dull tongue, white coating, thin or slow pulse;

8) There are 2 main symptoms and 1-2 secondary symptoms, which can be diagnosed by referring to the tongue and pulse.

2.2 Inclusion criteria

8) Meet the indications for entering the IVF-ET cycle and the TCM dialectical criteria for kidney deficiency and blood stasis syndrome;

9) Age 25-45 years old;

10) Embryo transfer failed for three or more pregnancies;

11) Those who have cryopreserved high-quality embryos (≥2 embryos) in our center;

12) Those who have no uterine cavity lesions or have been cured by hysteroscopy and other operations;

13) Those who have no hydrosalpinx or have been surgically ligated;

14) Sign the informed consent.

2.3 Exclusion criteria

7) Diseases of the uterus and uterine cavity that have not been treated or cured by hysteroscopy, such as uterine malformation, uterine fibroids, endometrial polyps, endometritis, intrauterine adhesions, uterine fibrosis, adenomyosis, etc. ;

8) Those with hydrosalpinx and no surgical treatment;

9) Those who cannot cooperate, such as those with neurological and mental disorders;

10) Those with incomplete information;

11) Those who are allergic to the drugs used;

12) Participating in other clinical investigators.

2.4 Elimination and shedding standards

5) Those who cannot continue treatment due to various reasons;

6) Those with adverse drug reactions;

7) Those with concurrent diseases;

8) Those who withdraw from the test process by themselves.

3 treatment options

3.1 Random grouping

60 selected RIF patients underwent endometrial preparation, and SPSS 21.0 statistical software was used to generate random numbers according to the number of cases. According to the order of admission, they were divided into Chinese medicine group and control group 1 and control group 2, with 20 cases in each group.

3.2 Treatment methods

(1) Endometrial preparation: The subjects entered the week on the 2-3rd day after the menstrual cycle or drug withdrawal bleeding, and on the same day after the ovarian cyst was excluded by the first B-ultrasound, Fenmotonghong tablets (estradiol 2mg, Abbott, Netherlands, approval number H20110159) 4mg, 2 times a day, after oral administration for 12 consecutive days, a second B-ultrasound was performed to monitor the thickness of the intima. (Estradiol/Dydrogesterone 2mg, Abbott, Netherlands, Approval No. H20110159), 4mg, 2 times a day, continuous oral administration for 3 days for endometrial transformation, the day is designated as the day of endometrial transformation, 3 days after transformation Cleavage-stage embryos can be transferred or blastocysts can be transferred 5 days after transformation; if the subject's second B-ultrasound monitoring endometrial thickness is less than 8 mm, proganol (estradiol valerate 1 mg, Bayer, Germany, Approval number J20130009) 1mg, once a day, vaginal administration. Estrogen is taken for a maximum of 3 weeks, and if the intima is still <8cm, the cycle will be canceled.

(2) Luteal support: after frozen embryo transfer, Fenmotonghuang tablets (estradiol/dydrogesterone) 4mg, 2 times a day, Angelan (progesterone soft capsule 0.1g, Bessen, Belgium, approval number) H20130336) 0.2g, 2 times a day, vaginal medication.

(3) Traditional Chinese medicine group: the subjects started oral administration of Example 4 of the present invention on the basis of the routine endometrial preparation and luteal support program on the 2nd to 3rd day after the menstrual cycle or drug withdrawal bleeding, (granule preparation : The traditional Chinese medicine composition of embodiment 4 is continuously decocted 3 times, the water addition amount of each decocting is 10 times the weight of the traditional Chinese medicine composition, and each decocting time is 3h, and the filtrate after 3 times of decocting is collected; the filtrate is concentrated To a relative density of 1.33 at 60° C., drying and pulverizing to obtain extract powder; adding starch and dextrin to extract powder to make granules). The granules were taken once a day and continued to be taken until 14 days after embryo transfer. In control group 1, subjects started oral administration of control group 1 of the present invention on the basis of routine endometrial preparation and luteal support program on the 2nd to 3rd day after menstrual cycle or drug withdrawal bleeding (preparation of granules is the same as above). In control group 2, the subjects started oral administration of control group 2 of the present invention on the basis of routine endometrial preparation and luteal support program on the 2nd to 3rd day after menstrual cycle or drug withdrawal bleeding (preparation of granules is the same as above).

3.3 Observation indicators and methods

3.3.1 General information: age of the patient, years of infertility, type of infertility, infertility factors, number of previous transplants, basal hormone levels, and basal endometrial status.

3.3.1 Comparison of clinical pregnancy rates

Clinical pregnancy rate = (number of clinical pregnancy cycles/total number of transplant cycles) × 100%

4 results

4.1 Comparison of clinical pregnancy rates

The pregnancy rate in the Chinese medicine group was higher than that in the control group (35.00% VS 20.00% VS 25.00%), and the difference was statistically significant (p<0.05).

Comparison of clinical pregnancy rates among the three groups [cases (%)]

Claims (5)

1. The traditional Chinese medicine composition for treating the diseases of repeated embryo planting failure is characterized by being prepared from the following raw material medicines in parts by weight: 29-31 parts of amethyst, 14-16 parts of semen cuscutae, 14-16 parts of prepared rehmannia root, 11-13 parts of caulis spatholobi, 14-16 parts of parasitic loranthus, 8-10 parts of red paeony root, 14-16 parts of angelica sinensis, 14-16 parts of radix paeoniae alba, 8-10 parts of ligusticum wallichii, 14-16 parts of bighead atractylodes rhizome, 14-16 parts of codonopsis pilosula, 14-16 parts of poria cocos, 11-13 parts of eucommia ulmoides, 11-13 parts of teasel root and 8-10 parts of dried orange peel.

2. The traditional Chinese medicine composition according to claim 1, which is prepared from the following raw materials in parts by weight: 30 parts of amethyst, 15 parts of dodder, 15 parts of prepared rehmannia root, 12 parts of suberect spatholobus stem, 15 parts of Chinese taxillus twig, 9 parts of red paeony root, 15 parts of Chinese angelica, 15 parts of white paeony root, 9 parts of szechuan lovage rhizome, 15 parts of largehead atractylodes rhizome, 15 parts of pilose asiabell root, 15 parts of Indian buead, 12 parts of eucommia bark, 12 parts of himalayan teasel root and 9 parts of tangerine peel.

3. The use of the Chinese medicinal composition according to claim 1 or 2 in the preparation of a medicament for the treatment of embryo repeat planting failure diseases.

4. The use according to claim 3, wherein the disease is kidney deficiency and blood stasis embryo repeated planting failure disease.

5. The use according to claim 3, wherein the disease is a disease of repeated implantation failure of embryos due to yang deficiency and blood stasis.

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