Traditional chinese medicine composition for treating membranous nephropathy, preparation method therefor, and use thereof

Abstract

Provided are a traditional Chinese medicine composition for treating membranous nephropathy, a preparation method therefor, and use thereof. The traditional Chinese medicine composition comprises the following traditional Chinese medicine raw materials in parts by weight: 3-9 parts of Taxus chinensis, 9-12 parts of Curcuma zedoaria, 15-30 parts of Astragalus membranaceus, 6-12 parts of Panax notoginseng, and 10-20 parts of Gynostemma pentaphyllum. At the animal level, the traditional Chinese medicine composition can reduce proteinuria, regulate the lipid metabolism disorder, and protect the kidney. Furthermore, it is proved that the traditional Chinese medicine composition can alleviate the vacuolar degeneration of GBM or GBM thickening in experimental rats. Immunohistochemical analysis demonstrates that the formula can reduce the expression level of TGF-β1 in the renal interstitium, reduce the expression of ɑ-SMA and Col-IV collagen, reduce the deposition of ECM in the renal interstitium, and delay glomerulosclerosis and renal interstitial fibrosis.

Description

A traditional Chinese medicine composition for treating membranous nephropathy, its preparation method and application Technical Field

[0001] The present invention relates to the technical field of traditional Chinese medicine, and in particular to a traditional Chinese medicine composition for treating membranous nephropathy, a preparation method thereof and application thereof. Background Art

[0002] Membranous nephropathy (MN) is a common pathological form of nephrotic syndrome in adults. It is a severe immune-mediated renal disease characterized histomorphologically by immune complex deposition in the subglomerular epithelial space, mesangial thickening, and impaired podocyte structural integrity. Its main clinical features are proteinuria, hypoproteinemia, and edema. The pathogenesis of MN is complex, involving numerous factors, including genetics, autoimmune mechanisms, complement activation, and the interplay between cell-specific adaptability and reactivity. It is also associated with the development and progression of a variety of diseases, including systemic autoimmune diseases such as lupus nephropathy type V caused by systemic lupus erythematosus, malignancies such as multiple myeloma, infections such as hepatitis B-related MN, and drug intoxication. Reports abroad suggest that MN accounts for 25%-40% of nephrotic syndrome in adults, while reports in China suggest a 10%-15% incidence. MN responds poorly to steroids and immunosuppressants, and approximately 40% of patients ultimately develop end-stage renal failure (ESRD) and die.

[0003] The pathogenesis of MN mainly includes the following aspects: (1) The presence of HLA alleles causes autoimmune mechanism disorders and low immunity; (2) The polymorphic PLA2R1, due to its unique structure, helps autoantibodies find target antigens; (3) Gene mutations caused by age, genetics or environmental factors cause the body to produce anti-phospholipase A2 receptor autoantibodies that activate the complement pathway; (4) After complement activation, the C5b-9 membrane attack complex is formed, leading to podocyte damage and proteinuria.

[0004] MN is one of the most common causes of nephrotic syndrome in adults. The latest MN prevention and treatment guidelines recommend the use of hormones, immunosuppressants, cytotoxic drugs, etc., either alone or in combination, such as glucocorticoids + cyclophosphamide, glucocorticoids + cyclosporine, glucocorticoids + mycophenolate mofetil, rituximab, tacrolimus, etc. Although these drugs can achieve the purpose of treating MN, they have the disadvantages of many contraindications, serious and complex adverse reactions, large side effects, high recurrence rate, and high price in clinical use. Therefore, more, more advanced, more reasonable, more holistic, and comprehensive drugs are still needed to improve the current treatment strategies, help MN patients delay the pathological process, reduce the occurrence of MN complications, and improve the quality of life.

[0005] Disclosure of the invention

[0006] The purpose of the present invention is to overcome the defect that existing drugs are not effective in treating membranous nephropathy.

[0007] In order to achieve the above-mentioned object, the present invention provides a traditional Chinese medicine composition for treating membranous nephropathy, which comprises the following traditional Chinese medicine raw materials in parts by weight: 3 to 9 parts of Taxus chinensis, 9 to 12 parts of Curcuma zedoaria, 15 to 30 parts of Astragalus membranaceus, 6 to 12 parts of Panax notoginseng, and 10 to 20 parts of Gynostemma pentaphyllum.

[0008] Preferably, 5 to 7 parts of Taxus chinensis, 10 to 11 parts of Curcuma zedoaria, 18 to 25 parts of Astragalus membranaceus, 8 to 10 parts of Panax notoginseng, and 12 to 18 parts of Gynostemma pentaphyllum.

[0009] Preferably, 9 parts of Taxus chinensis, 12 parts of Curcuma zedoaria, 30 parts of Astragalus membranaceus, 12 parts of Panax notoginseng, and 20 parts of Gynostemma pentaphyllum.

[0010] Preferably, the traditional Chinese medicine composition further comprises pharmaceutically acceptable excipients.

[0011] Preferably, the dosage form of the Chinese medicine composition is selected from any one of tablets, ointments, granules, granules, and oral liquids.

[0012] The present invention also provides a method for preparing a traditional Chinese medicine composition for treating membranous nephropathy, comprising drying the various traditional Chinese medicine raw materials, mixing them uniformly, and crushing them into fine powder; or adding water and decocting them for 2 to 4 hours, concentrating the decoction, and drying them to obtain the traditional Chinese medicine composition.

[0013] Preferably, after being crushed into fine powder, a pharmaceutically acceptable carrier and / or excipient is added.

[0014] The present invention also provides a use of the above-mentioned traditional Chinese medicine composition for treating membranous nephropathy, wherein the traditional Chinese medicine composition is used for preparing a medicine for treating membranous nephropathy.

[0015] Preferably, the Chinese medicine composition is used to prepare a medicine for treating glomerular diseases.

[0016] Preferably, the Chinese medicine composition is used to prepare a medicament for treating nephrotic syndrome.

[0017] Compared with the prior art, the beneficial effects of the present invention include at least:

[0018] In animals, the traditional Chinese medicine composition of the present invention can reduce proteinuria, regulate lipid metabolism disorders, and protect the kidneys. Experimental studies further demonstrated that the composition can alleviate vacuolar degeneration or thickening of GBM in rats. Electron microscopy showed that the composition can alleviate podocyte fusion damage and reduce spike formation. Immunohistochemistry showed that the composition can reduce the expression of TGF-β1 in the renal interstitium, reduce the expression of α-SMA and Col-IV collagen, reduce the deposition of ECM in the renal interstitium, and delay glomerular sclerosis and renal interstitial fibrosis.

[0019] At the clinical level, all patients showed improvement or significant improvement in 24-hour urine protein, plasma albumin, D-dimer, fibrinogen, triglycerides, total cholesterol, blood β2-microglobulin, and urine β2-microglobulin compared to three months prior to treatment. After six months of follow-up, eight patients in the integrated Chinese and Western medicine group gradually discontinued immunotherapy and switched to pure Chinese medicine treatment. The overall effective rate reached 93.33%, with a complete remission rate of 43.33%. The Chinese herbal composition of this invention plays an important role in the treatment of medullary nephropathy.

[0020] BRIEF DESCRIPTION OF THE DRAWINGS

[0021] FIG1 is a diagram showing the HE staining results of rats according to the present invention.

[0022] FIG2 is a diagram showing the Masson staining results of rats according to the present invention.

[0023] FIG3 is an electron microscopic observation diagram of rat kidney tissue according to the present invention.

[0024] FIG4 is a graph showing the immunohistochemical expression results of TGF-β1 in rat kidney tissue according to the present invention.

[0025] FIG5 is a graph showing the immunohistochemical expression results of CoI-IV in rat kidney tissue according to the present invention.

[0026] FIG6 is a graph showing the expression results of α-SMA immunohistochemistry in rat kidney tissue according to the present invention.

[0027] Best Mode for Carrying Out the Invention

[0028] The technical solution of the present invention is further described below with reference to the accompanying drawings.

[0029] Membranous nephropathy (MN) is a difficult type of chronic glomerulonephritis to treat. Currently, Western medicine typically treats severe MN with a combination of glucocorticoids (GC) and nonspecific immunosuppressants, including alkylating agents such as cyclophosphamide (CTX), calcineurin inhibitors such as cyclosporin A (CsA), and tacrolimus (TAC). However, these traditional immunosuppressants have low remission rates and are associated with drug dependency. Clinical relapses are common during dose reduction, and some immunosuppressants have significant adverse reactions, including elevated serum creatinine and accelerated renal function deterioration. Traditional Chinese medicine (TCM), guided by a holistic approach and based on syndrome differentiation and treatment, offers advantages in preventing and treating MN, including fewer adverse reactions, minimal side effects, high safety, good tolerability, multiple targets, low relapse rates after drug discontinuation, and definite efficacy. Therefore, vigorously developing traditional Chinese medicine to prevent and treat MN will effectively avoid the shortcomings and disadvantages of the current commonly used Western medicine treatment options in clinical practice, such as many contraindications, serious and complex adverse reactions, large side effects, high recurrence rate, and high prices, and bring excellent treatment effects to patients while reducing the burden on patients and society. Therefore, the prevention and treatment of MN with traditional Chinese medicine is a long-standing hot topic and key issue in the medical field.

[0030] Traditional Chinese Medicine (TCM) considers the fundamental pathogenesis of membranous nephropathy to be root deficiency and superficial excess. The root deficiency is often due to insufficient endowment or spleen and kidney deficiency, while the superficial excess is often due to the accumulation of dampness and heat, which damages kidney qi. Spleen deficiency prevents the retention of essence, and kidney deficiency prevents the consolidation of essence and qi, leading to turbid semen and its downward flow. Chronic disease damages the kidney meridians, causing blood stasis and water retention, resulting in edema. This disease is often recurrent and difficult to cure. This study utilizes a compound formula, Rhus chinensis, which is known for its heat-clearing, turbidity-reducing, and qi-invigorating effects, based on the patient's underlying syndromes.

[0031] Part I Animal Experiments

[0032] 1 Materials and Methods

[0033] 1.1 Experimental Animals

[0034] Twenty-four male Sprague-Dawley rats, SPF grade, weighing 160–180 g and 6 weeks old, were purchased from Changzhou Cavens Laboratory Animal Co., Ltd. (Laboratory Animal License: SCXK(Su)2021-0013) and housed in the Animal Laboratory of Shanghai Sixth People's Hospital (Laboratory Animal Facility License: SYXK(Shanghai)2021-0028). The rats were maintained in an environment with a temperature of 23–26°C and a humidity of 45–50%, with free access to food and water.

[0035] 1.2 Experimental drugs

[0036] The compound Chinese herbal medicine composition of Taxus chinensis (batch number: 2306201), Panax notoginseng (batch number: 230305), Curcuma zedoaria (batch number: 220101), Astragalus membranaceus (batch number: 210401), and Gynostemma pentaphyllum (batch number: 210601) were purchased from Anhui Heyaopu Chinese Medicine Effective Company. These five herbs were decocted twice with 10 times the amount of water, each time for 2 hours. The filtrates were then filtered and concentrated under reduced pressure to a clear paste with a relative density of 1.23-1.28 (80°C). The mixture was then vacuum-dried and ground into a coarse powder for later use. Losartan tablets (50 mg) were purchased from Merck & Co., Ltd., with approval number: National Medicine Standard J20180053.

[0037] 1.3 Main Reagents

[0038] 1.4 Main instruments

[0039] 1.5 Animal model preparation, experimental grouping, and drug administration

[0040] After one week of adaptive feeding, 24 SD rats were randomly divided into a normal group (N) (n=6) and a model group (n=18). A membranous nephropathy rat model was established using a modified Border method, with a 24-hour total urinary albumin (24hUTP) level ≥20 mg as the criterion for successful model establishment. After successful model establishment, rats were randomly divided into a model group (M), a losartan group (L), and a compound yew group (H), with six rats in each group. Rats were given losartan (10 mg / kg) and yew compound (1.5 g / kg) by oral gavage, calculated based on the body surface area of ​​humans and rats, once daily for eight weeks. At the end of the eighth week, 24-hour urine was collected in metabolic cages, the supernatant was centrifuged, and stored at 4°C until further use. After the experiment, the rats were deprived of food and water for 12 hours and anesthetized with isoflurane inhalation. Serum and renal tissue were collected and stored at -80°C until further use.

[0041] 1.6 Index detection

[0042] 1.6.1 Blood and urine biochemical index detection The rat serum BUN, SCr, TG, TC, ALB, LDL, and HDL levels were detected using an automatic biochemical analyzer according to the kit instructions, and the 24-h urine total albumin (24hUTP) was detected using an enzyme-linked immunosorbent assay.

[0043] 1.6.2 Pathological changes in renal tissue were examined by hematoxylin-eosin (HE) staining, Masson staining, and transmission electron microscopy. Renal tissue fixed in 4% paraformaldehyde was dehydrated, paraffin-embedded, and sectioned. Sections were prepared using the HE and Masson staining methods, respectively, and observed under a light microscope for renal histopathological changes. Renal tissue was fixed in 2.5% glutaraldehyde solution, washed three times in PBS, fixed in 1% osmium hydroxide, and dehydrated in acetone in successive steps before embedding, polymerization, and ultrathin sectioning. Transmission electron microscopy and photography were performed for double staining with uranyl acetate and lead phosphate.

[0044] 1.6.3 Immunohistochemical analysis of renal tissue for TGF-β1, Col-IV, and α-SMA protein expression (SABC method) was performed strictly according to the kit instructions. Sections were routinely sectioned, deparaffinized, and hydrated; washed with PBS for 5 minutes each; blocked with fresh 3% H₂O₂ in distilled water or PBS for 5-10 minutes at room temperature; and washed three times with distilled water. Antigen retrieval was performed; washed with PBS for 5 minutes; normal goat serum blocking solution was added dropwise for 20 minutes at room temperature, and excess liquid was removed. Rabbit anti-rat primary antibody was added dropwise at 4°C overnight; washed three times with PBS for 2 minutes each; goat anti-rabbit secondary antibody was added dropwise at 37°C for 20 minutes; washed three times with PBS for 20 minutes each; SABC reagent was added dropwise at room temperature for 20 minutes; washed four times with PBS for 5 minutes each. DAB color development was performed using a DAB color development kit; washed with distilled water, counterstained with hematoxylin for 2 minutes, and differentiated with hydrochloric acid-alcohol; dehydrated, cleared, mounted, and examined under a microscope. Grayscale values ​​of bands were analyzed using Image J software.

[0045] 1.7 Statistical methods

[0046] SPSS 22.0 statistical software was used for data analysis. All measurement data were tested to be normally distributed with homogeneous variance and expressed as mean ± standard deviation (x ± s). One-way analysis of variance was used for comparison between groups, and LSD-t test was used for multiple comparisons. P < 0.05 indicated statistically significant differences.

[0047] Experimental results:

[0048] 1. Comparison of rat body weight, kidney weight / body weight, and 24hUTP

[0049] As shown in Table 1, there were significant statistical differences in body weight, kidney weight index, and 24-hour urine albumin content between the rats in the normal control group and those in the model control group, compound yew group, and losartan group (P < 0.01). There were also significant statistical differences in body weight, kidney weight index, and 24-hour urine albumin content between the rats in the losartan group and those in the model control group (P < 0.01). There were also significant statistical differences in body weight, kidney weight index, and 24-hour urine albumin content between the rats in the compound yew group and those in the model control group and losartan group (P < 0.01).

[0050] Table 1 Comparison of body weight, renal mass index, and 24-hour urine microalbumin excretion rate of rats in each group at the end of the 8th week ( ±s)

[0051] GroupnBW (g) RW / BW (mg / g) UTP (μg / 24 h) N6584.40±5.592.28±0.0871.53±10.17M6437.92±8.19**4.98±0.06**1473.75±11.15**L6475.68±9. 53**##3.77±0.04**##786.88±74.26**##H6506.77±7.25**##■■3.36±0.05**##■■586.91±64.03**##■■

[0052] Note: *: compared with the normal group (N), P < 0.05, **: compared with the normal group (N), P < 0.01; #: compared with the model control group (M), P < 0.05, ##: compared with the model control group (M), P < 0.01; ■: compared with the losartan group (L), P < 0.05, ■■: compared with the losartan group (L), P < 0.01. (The same below)

[0053] 2. Comparison of blood biochemical parameters in rats of each group

[0054] As shown in Tables 2 and 3, BUN and Scr levels in the normal control group were significantly different from those in the model control group, the compound yew group, and the losartan group (P < 0.01). Alb, TCH, and TG levels in the normal control group were statistically different or significantly different from those in the model control group and the losartan group (P < 0.05 or P < 0.01). LDL and HDL levels in the normal control group were statistically different from those in the model control group (P < 0.05). BUN, Scr, Alb, LDL, and HDL levels in the losartan group were statistically different or significantly different from those in the model control group (P < 0.05 or P < 0.01). BUN, Scr, Alb, TCH, TG, LDL, and HDL levels in the compound yew group were statistically different or significantly different from those in the losartan group and the model control group (P < 0.05 or P < 0.01).

[0055] Table 2 Comparison of blood urea nitrogen (BUN), serum creatinine (Scr), and plasma albumin (Alb) in rats of each group at the end of the 8th week ( ±s)

[0056] GroupnBUN(mmol / l)Scr(umol / l)Alb(g / L)N65.99±0.5257.81±2.2931.50±3.21M618.30±1.96**96.77±5.33**22.1 7±2.93**L615.20±0.63**##68.27±3.63**##26.00±1.41*#H613.15±0.99**##■■64.30±2.93**##■■29.17±1.72##■

[0057] Table 3 Comparison of total cholesterol (TCH), triglyceride (TG), low-density lipoprotein (LDL), and high-density lipoprotein (HDL) in rats of each group at the end of the 8th week ( ±s)

[0058] GroupnTCH(mmol / l)TG(mmol / l)LDL(mmol / l)HDL(mmol / l)N60.94±0.210.37±0.220.18±0.090.83±0.20M61.86±0.43**0.88±0.21** 0.33±0.10*0.30±0.31*L61.31±0.31*0.64±0.22*0.23±0.08#1.04±0.24##H61.07±0.26#■0.39±0.11##■0.10±0.06##■0.83±0.25#■

[0059] 3. Glomerular pathological damage

[0060] As shown in Figures 1 and 2, the pathological damage of the rats in the model group was more severe than that in the compound yew group and the losartan group. The difference was significant after statistical analysis (P < 0.01). HE staining results showed that the rats in the model control group and the compound yew group had glomerular mesangial cell proliferation and basement membrane thickening. Masson staining showed aneurysmal dilatation of glomerular capillaries, nodular proliferation of mesangial matrix, sclerosis of some glomeruli, vacuolar degeneration of some renal tubular epithelial cells, and varying degrees of inflammatory cell infiltration. The renal tissue damage in the losartan group was alleviated to varying degrees compared with the model control group.

[0061] 4. Electron microscopic observation of renal tissue of rats in each group

[0062] As shown in Figure 3, there were no significant pathological changes in the renal tissue of the normal control group. Compared with the normal multi-irradiation group, the model control group showed glomerular mesangial matrix hyperplasia, homogeneous GBM thickening, and partial glomerulosclerosis. Compared with the model control group, the glomerular mesangial matrix hyperplasia in the compound yew group and the losartan group was reduced, and GBM thickening was alleviated to varying degrees.

[0063] 5. Immunohistochemistry Results

[0064] 5.1 TGF-β1 immunohistochemical expression results

[0065] As shown in Figure 4 , compared with the normal control group, the expression of TGF-β1 in the kidney tissue of the model group rats was significantly increased, and the expression of TGF-β1 in the kidney of the losartan group and the compound yew group was lower than that of the model group (P < 0.05 or P < 0.01).

[0066] 5.2 Col-IV immunohistochemical expression results

[0067] As shown in Figure 5 , compared with the normal control group, the expression of Col-IV in the kidney tissue of the model group rats was significantly increased; the expression of Col-IV in the kidney of the losartan group and the compound yew group was lower than that of the model group (P < 0.05 or P < 0.01).

[0068] 5.3 α-SMA Immunohistochemical Expression Results

[0069] As shown in Figure 6, the expression of α-SMA in the model group was higher than that in the normal group. Compared with the model group, the expression of α-SMA in the kidneys of the losartan group and the compound yew group decreased in turn.

[0070] In summary, animal studies have demonstrated that this Chinese herbal composition can reduce proteinuria, regulate lipid metabolism disorders, and protect the kidneys. Experimental studies have further demonstrated that this Chinese herbal composition can reduce vacuolar degeneration or GBM thickening in rats. Electron microscopy has shown that this Chinese herbal composition can alleviate podocyte fusion damage and reduce spike formation. Immunohistochemistry has shown that this Chinese herbal composition can reduce the expression of TGF-β1 in the renal interstitium, decrease the expression of α-SMA and Col-IV collagen, reduce the deposition of ECM in the renal interstitium, and delay glomerular sclerosis and renal interstitial fibrosis.

[0071] Part II Clinical Research

[0072] General Information

[0073] A total of 60 patients with membranous nephropathy (MN) stages I-III diagnosed by renal biopsy pathology were collected from the Department of Traditional Chinese Medicine Nephrology and wards of our hospital between January 2018 and December 2022. The 60 patients were randomly divided into 40 mild MN patients (24hUTP < 3.5g) and 20 severe MN patients (24hUTP ≥ 3.5g). Among the 40 mild MN patients, 23 were male and 17 were female, with a mean age of 49.75±16.39 years and a mean disease duration of 43.15±29.09 months. Among the 20 severe MN patients, 15 were male and 5 were female, with a mean age of 44.85±14.85 years and a mean disease duration of 35.40±24.11 months. Eighteen patients had hypertension.

[0074] 2. Diagnostic criteria

[0075] 2.1 Western medicine diagnostic criteria

[0076] Refer to the 2011 Chinese Medical Association Clinical Diagnosis and Treatment Guidelines: Nephrology and Wang Haiyan's Nephrology. (1) Clinical manifestations are NS (massive proteinuria, hypoproteinemia, severe edema, and hyperlipidemia), which may be accompanied by a small amount of microscopic hematuria; (2) Some patients have hypertension; (3) Renal biopsy pathology diagnoses idiopathic membranous nephropathy; (4) Blood antiphospholipase A2 receptor antibodies are positive; (5) Secondary factors are excluded.

[0077] 2.2 Standards for TCM Syndrome Differentiation

[0078] With reference to the TCM syndrome diagnostic criteria for edema formulated in the Diagnosis, Syndrome Differentiation and Efficacy Evaluation of Chronic Glomerulonephritis (Trial Program) and the 2002 Guiding Principles for Clinical Research of New Chinese Medicines, the syndrome belongs to spleen and kidney deficiency, dampness and turbidity with blood stasis.

[0079] ①Spleen and kidney deficiency syndrome

[0080] Main symptoms: dizziness, tinnitus, turbid urine; fatigue, edema of limbs, or edema of eyelids and feet;

[0081] Secondary symptoms: soreness and weakness in waist and knees, shortness of breath and laziness to speak, abdominal distension and poor appetite, cold body and limbs.

[0082] Tongue and pulse: There are tooth marks on the side of the tongue and the pulse is weak.

[0083] ② Dampness and turbidity syndrome

[0084] Main symptoms: nausea and vomiting, heaviness and drowsiness, loose stools.

[0085] Secondary symptoms: abdominal distension and sticky taste in the mouth.

[0086] Tongue and pulse: There are teeth marks on the edge of the tongue, the tongue coating is white or yellow (essential), and the pulse is weak and rapid.

[0087] ③ Blood stasis syndrome

[0088] Main symptoms: skin and nails, dark purple lips.

[0089] Secondary symptoms: heaviness and soreness in limbs, chest tightness and abdominal distension.

[0090] Tongue and pulse: The tongue is dull or has ecchymosis, and the sublingual veins are purple and distended; the pulse is deep and thin or slow and weak, or deep and astringent.

[0091] Syndrome differentiation suggestions: Dizziness, tinnitus, and turbid urine can be diagnosed as kidney deficiency syndrome; fatigue, weakness, and edema of the limbs can be diagnosed as spleen deficiency syndrome; loose stools, greasy tongue coating, and abdominal distension can be diagnosed as dampness syndrome; and a dull tongue or ecchymosis can be diagnosed as blood stasis syndrome.

[0092] If the patient has two main symptoms and two secondary symptoms, and the tongue and pulse conditions are consistent with the above-mentioned blood stasis and collateral obstruction, the patient can be diagnosed with spleen and kidney yang deficiency and blood stasis and collateral obstruction syndrome.

[0093] 3. Inclusion criteria

[0094] (1) Age 16-75 years; (2) Blood pressure controlled below 130 / 80 mmHg (1 mmHg ≈ 0.133 kPa) through exercise, diet control, and oral antihypertensive drugs; (3) Blood creatinine (SCr) level is basically normal.

[0095] 4. Exclusion criteria

[0096] (1) Those with secondary nephrotic syndrome; (2) Those with infectious diseases, poisoning, infection, mental illness, and serious heart, brain, liver and other diseases; (3) Those who do not cooperate with treatment, do not use medication according to regulations, cannot judge the effect, or have incomplete information that affects the effect.

[0097] 5. Treatment Methods

[0098] 5.1 Basic treatment

[0099] All patients were placed on a low-salt, low-fat, high-quality protein diet, and health education was provided. Treatment was based on the 2020 Kidney Disease: Improving Global Outcomes (KDIGO) guidelines, including antihypertensive, anticoagulant, diuretic, lipid-lowering, and immunosuppressive therapy. Patients with hyperlipidemia were treated with atorvastatin calcium tablets (Beijing Jialin Pharmaceutical Co., Ltd., National Medicine Approval No. H20180208) 20 mg orally once daily for lipid-lowering therapy. Cilostazol 50 mg orally twice daily was used as an anticoagulant. Patients with concomitant essential hypertension were treated with antihypertensive medications other than angiotensin-converting enzyme inhibitors (ACEIs) or angiotensin receptor blockers (ARBs) to maintain blood pressure below 130 / 80 mmHg. For patients with severe edema, furosemide and spironolactone tablets were intermittently used for diuresis and swelling reduction.

[0100] 5.2 Immunotherapy

[0101] For patients with 24hUTP 3.5-6g / L, immunotherapy is selected, and prednisone 0.5mg / (kg.d) is given; for patients with 24hUTP>6g / L, prednisone 0.5mg / (kg.d) combined with cyclophosphamide 0.8-1g / month is given, with a cumulative dose of 6-8g; for patients with contraindications to cyclophosphamide, prednisone 0.5mg / (kg.d) combined with tacrolimus 0.050-0.075mg / (kg.d) is given for treatment. Tacrolimus is administered in two doses with an interval of 12h, and the blood drug concentration is maintained at 5-10ng / ml.

[0102] 5.3 Chinese herbal compound yew

[0103] Astragalus 30g, Taxus 9g, Panax notoginseng 12g, Curcuma 12g, and Gynostemma pentaphyllum 20g. All five medicinal pieces were purchased from Anhui Heyaopu Chinese Medicine Effective Company. The above five herbs represent a single daily dose for adults, prepared based on a 90-day dosage. Add 10 times the amount of water and decoct twice for 2 hours each time. Filter, combine the filtrates, and concentrate under reduced pressure to a clear paste with a relative density of 1.23-1.28 (80°C). Take 20ml each time, warm, in the morning and evening, 30 minutes after meals.

[0104] 5.4 Group treatment

[0105] Forty patients with mild MN were randomly divided into a basic treatment group (20 cases, including 11 males and 9 females with a mean age of 51.65±15.76 years and a mean disease duration of 39.85±21.87 months) and a compound yew group (20 cases, including 12 males and 8 females with a mean age of 47.85±17.19 years and a mean disease duration of 46.45±35.16 months). The basic treatment group received losartan tablets 50 mg / d, once daily; the compound yew group received only traditional Chinese medicine treatment, one dose per day, decocted in water to obtain 300 mL of juice, taken warm twice a day, 30 minutes after meals, in the morning and evening. Twenty patients with severe MN were randomly divided into an immunotherapy group (10 patients) and a combined traditional Chinese and Western medicine group (10 patients). The immunotherapy group included seven males and three females with a mean age of 47.70±15.74 years and a mean disease duration of 40.30±25.97 months. Immunosuppressants were added to baseline treatment. The combined traditional Chinese and Western medicine group included eight males and two females with a mean age of 42.00±14.14 years and a mean disease duration of 30.50±22.34 months. Basic treatment was supplemented with the traditional Chinese medicine compound Rhododendron chinense. All four groups received a 12-week course of treatment. There were no statistically significant differences in mean age and disease duration before treatment among the groups (P>0.05).

[0106] 6. Observation indicators

[0107] (1) Comparison of the changes in syndrome scores before and after treatment in the four groups. Each syndrome was scored as 0, 2, 4, and 6 points according to none, mild, moderate, and severe, respectively. The syndrome efficacy was observed. (2) Observation of clinical efficacy. (3) Comparison of plasma albumin, triglycerides, cholesterol, renal function (serum creatinine, urea nitrogen), fibrinogen, and 24-hour urine protein levels before and after treatment in the four groups. Changes in TCM syndrome scores, 24-hour urine protein (24hUTP), serum creatinine (Scr), urea nitrogen (BUN), uric acid (UA), serum cystatin (Cys-C), serum albumin (Alb), hemoglobin (Hb), red blood cell count (RBC), total cholesterol (TCH), triglycerides (TG), low-density lipoprotein (LDL), D-dimer (DD), fibrin degradation products (FDP) before and after treatment in the four groups were observed, as well as clinical efficacy and residual renal function.

[0108] 7. TCM Syndrome Efficacy Standards

[0109] Markedly effective: Significant improvement in TCM clinical symptoms and signs, with a symptom score reduction of ≥70%; Effective: Improvement in TCM clinical symptoms and signs, with a symptom score reduction of ≥30% (>70%); Ineffective: No significant improvement in TCM clinical symptoms and signs, or even worsening, with a symptom score reduction of <30%. Total effective rate = (markedly effective + effective) / total number of cases × 100%.

[0110] 8. Clinical efficacy standards

[0111] (1) Clinical remission: Symptoms and positive signs completely disappear, and proteinuria achieves complete remission, i.e., urine protein quantification <0.3 g / d, confirmed at least once every week, and plasma albumin and blood creatinine levels return to normal. (2) Effective: Symptoms and positive signs basically disappear, and proteinuria achieves partial remission, i.e., urine protein quantification <3.5 g / d, and decreases by more than 50% compared with the previous level, confirmed at least once every week, and plasma albumin improves or returns to normal compared with the previous level, and blood creatinine levels are stable. (3) Ineffective: No improvement in clinical manifestations and laboratory tests. Total effective rate = (clinical remission + effective) number of cases / total number of cases × 100%.

[0112] 9. Statistical Methods

[0113] SPSS 19.0 statistical software was used for analysis. Measurement data were expressed as (xˉ±s) and tested with the t test. Enumeration data were expressed as percentages (%) and tested with the chi-square test. P < 0.05 was considered statistically significant.

[0114] Experimental results

[0115] 1. Analysis of the efficacy of 3-month treatment in 60 patients with MN

[0116] As shown in Table 4, among the 60 patients with membranous nephropathy, 14 achieved complete remission, 16 achieved basic remission, 18 improved, and 12 had no effect during the 3-month treatment course. The total effective rate of basic treatment for patients with mild MN was 70%, and the total effective rate of the compound yew group was 85%; the total effective rate of immunotherapy for patients with severe MN was 80%, and the total effective rate of combined Chinese and Western medicine treatment was 90%. After Redit analysis, there was a statistical difference between the compound yew group and the basic treatment group (P<0.05), and there was a statistical difference between the combined Chinese and Western medicine group and the immunotherapy group (P<0.05); this shows that the efficacy of using the Chinese medicine compound yew as the main treatment for MN is better than that of the basic treatment group and the immunotherapy group.

[0117]

[0118] Note: Radit: *P<0.05 compared with the basic treatment group; #P<0.05 compared with the immunotherapy group.

[0119] 2. Improvement of blood urea nitrogen (BUN), serum creatinine (Scr), triglycerides (TG), and cholesterol (TCH) in 60 patients with MN after 3 months of treatment

[0120] As shown in Table 5, after treatment, BUN, TG, and TCH levels in the 60 MN patients all improved compared to pre-treatment levels, with statistically significant or significant differences (P < 0.05 or P < 0.01). SCr levels also improved after treatment in the compound yew group and the integrated Chinese and Western medicine group, with statistically significant differences (P < 0.01). Among patients with mild MN, SCr and TG levels improved more significantly in the compound yew group than in the basic treatment group (P < 0.05). Among patients with severe MN, BUN, Scr, TG, and TCH levels improved more significantly in the integrated Chinese and Western medicine group than in the immunotherapy group (P < 0.05).

[0121]

[0122] Note: Compared with before treatment: * P < 0.05; ** P < 0.01; Comparison between groups: # P < 0.05; ## P < 0.01. (Same below)

[0123] 3. Improvement of 24h urine protein (24hUTP), plasma albumin (Alb), blood β2-microglobulin (S-β2-MG), and urine β2-microglobulin (U-β2-MG) in 60 MN patients before and after treatment

[0124] As shown in Table 6, among the 60 patients with membranous nephropathy, 24-hour UTP, Alb, S-β2-MG, and U-β2-MG all improved after treatment compared with those before treatment, with statistically significant differences (P < 0.05 or P < 0.01). Among patients with mild MN, the compound yew group showed significantly better improvements in UTP, Alb, S-β2-MG, and U-β2-MG after treatment than the basic treatment group, with statistically significant differences (P < 0.05). Among patients with severe MN, the combined traditional Chinese and Western medicine group showed significantly better S-β2-MG and U-β2-MG after treatment compared with the immunotherapy group, with statistically significant differences (P < 0.05).

[0125]

[0126] 4. Improvement of D-dimer (DD) and fibrin degradation product (FDP) indicators in 60 patients with MN before and after treatment

[0127] As shown in Table 7, after treatment, DD and FDP in the compound yew group of patients with mild MN improved or significantly improved compared with pre-treatment, with statistically significant differences (P < 0.05 or P < 0.01). Compared with the basic treatment group, DD and FDP improved significantly after treatment (P < 0.05). In patients with severe MN, DD and FDP improved significantly after treatment in the combined traditional Chinese and Western medicine group compared with pre-treatment, with statistically significant differences (P < 0.05). Compared with the immunotherapy group, FDP improved significantly after treatment (P < 0.01).

[0128]

[0129] 5. Comparison of TCM clinical syndromes in 60 patients with MN before and after treatment

[0130] As shown in Table 8, the TCM syndrome scores of the compound yew group, immunotherapy group, and integrated Chinese and Western medicine group all decreased after treatment compared with those before treatment, with statistically significant differences (P < 0.05 or P < 0.01). In the basic treatment group, there were no statistically significant differences in turbid urine for kidney deficiency syndrome, fatigue for spleen deficiency syndrome, or purplish tongue for dampness and blood stasis syndrome before and after treatment (P > 0.05). In the compound yew group, the TCM syndrome scores for loose stools for kidney deficiency syndrome, spleen deficiency syndrome, and dampness and turbidity syndrome, and cyanosis of the lips and nails for blood stasis syndrome, were significantly reduced after treatment compared with the basic treatment group in patients with mild MN (P < 0.05 or P < 0.01). In the integrated Chinese and Western medicine group, the TCM syndrome scores for kidney deficiency syndrome, spleen deficiency syndrome, dampness and turbidity syndrome, and purplish tongue for blood stasis syndrome, were significantly reduced after treatment compared with the immunotherapy group in patients with severe MN (P < 0.05 or P < 0.01).

[0131]

[0132] 6. Follow-up results

[0133] After three months of follow-up, 30 patients treated with compound yew as the primary treatment achieved complete remission in 13, basic remission in 8, improvement in 6, ineffectiveness in 2, and relapse in 1, with an overall effective rate of 93.33% and a relapse rate of 3.33%. Of the 30 patients treated with Western medicine, 8 achieved complete remission, basic remission in 11, improvement in 3, ineffectiveness in 3, and relapse in 5, with an overall effective rate of 73.33% and a relapse rate of 16.67%.

[0134] The present invention treats mild MN patients with 24-hour urine protein less than 3.5g only with the traditional Chinese medicine compound yew. The clinical efficacy of the three-month treatment course reaches 85%, and the complete remission rate reaches 25%. For 10 severe MN patients with 24-hour urine protein greater than 3.5g or / and accompanied by nephrotic syndrome, 5 patients received GC combined with CsA treatment, and 1 patient did not get relief; 3 patients received GC combined with TAC treatment and 1 patient did not get relief; 2 patients got better after receiving GC combined with CTX treatment; at the same time, 10 severe MN patients were observed and given the compound yew treatment on the basis of immunotherapy, 9 patients got better and 1 patient did not get relief. In the three-month treatment course, the total efficacy of the traditional Chinese and Western medicine combined medicine group for severe MN patients treated mainly with the compound yew reached 90%, and the complete remission rate was 40%. After 6-month follow-up, the total efficacy reached 93.33%, and the complete remission rate reached 43.33%.

[0135] All patients showed improvement or significant improvement in 24-hour urine protein, plasma albumin, D-dimer, fibrinogen, triglycerides, total cholesterol, blood β2-microglobulin, and urine β2-microglobulin compared to three months before treatment. After six months of follow-up, eight patients in the integrated Chinese and Western medicine group gradually discontinued immunotherapy and switched to pure Chinese medicine treatment. The overall effective rate reached 93.33%, with a complete remission rate of 43.33%.

[0136] Through the above observations, it can be seen that for severe MN patients with 24h urine protein greater than 3.5g, the combination of Chinese and Western medicine treatment with compound yew as the main ingredient can show good therapeutic effects within 3 months, and the effect is significantly improved with the extension of treatment time. For patients with 24h urine protein less than 3.5g, the prescription alone has a good therapeutic effect. The compound yew Chinese medicine composition plays an important role in the treatment of MN disease.

[0137] The present invention has grasped the basic pathogenesis of MN with deficiency of the root and excess of the superficial through many years of follow-up treatment. The deficiency of the root is insufficient endowment or deficiency of spleen and kidney, and excess of the superficial is dampness and blood stasis. The prescription focuses on invigorating qi and activating blood circulation, clearing heat and reducing turbidity. The whole prescription contains 5 Chinese herbs. Taxus chinensis is pungent, bitter, cold, enters the liver, kidney and spleen meridians, and has the effects of clearing heat and dispersing stagnation, activating blood circulation and removing blood stasis. Modern pharmacology proves that it has the effects of antioxidant, hypoglycemic, anti-inflammatory, diuretic and detumescent. Curcuma zedoaria is pungent, bitter, warm, enters the liver and spleen meridians, and has the effects of breaking blood circulation and promoting qi, eliminating accumulation and relieving pain. This medicine is pungent, bitter, warm, and can enter the blood and qi, and can break blood circulation and remove blood stasis, and can promote qi circulation and relieve pain. It has a strong ability to activate blood circulation and remove blood stasis, and is an important medicine for breaking blood and eliminating symptoms. Panax notoginseng is sweet, slightly bitter, warm, enters the liver and stomach meridians, and has the effects of dispersing blood stasis and stopping bleeding, reducing swelling and relieving pain. Panax notoginseng combined with Curcuma zedoaria can enhance the effects of promoting blood circulation. The effect of dispersing blood stasis, both of them have the functions of breaking and dispersing, moving and collecting, increasing the efficacy of promoting blood circulation and dispersing blood stasis. Modern pharmacology has found that the two medicines have the effects of improving blood circulation and regulating blood lipid metabolism; Astragalus is sweet and warm, enters the spleen and lung meridians, has the effects of replenishing qi and raising yang, benefiting the defense and strengthening the exterior, promoting diuresis and reducing swelling, and is used to treat various types of qi deficiency syndrome. Modern pharmacology has proved that Astragalus contains saponins, sucrose, polysaccharides, various amino acids, folic acid and various trace elements such as selenium, zinc, copper, etc., which can enhance the body's immune function, protect the liver, diuresis, anti-aging, anti-stress, lower blood pressure and a wide range of antibacterial effects, and have a good effect on chronic nephritis and diabetes. Astragalus is combined with Panax notoginseng and Curcuma zedoaria to attack and tonify, and can treat patients with weak constitution accompanied by blood stasis and dampness; Gynostemma pentaphyllum is sweet, bitter and slightly cold in nature, enters the spleen and lung meridians, and has the effects of invigorating qi and strengthening the spleen, clearing away heat and detoxifying, resolving turbidity and lowering lipids. Modern research shows that the main chemical components of Gynostemma pentaphyllum include tetracyclic triterpenoid saponins, such as gypenosapogenin, gypenosapogenin, and flavonoids. Pharmacological research also shows that this product has the effects of regulating blood lipids, enhancing immunity, resisting stress, and resisting liver damage. In short, the whole prescription uses both cold and warm methods, has both movement and astringency, both breaking and dispersing, both tonification and purgation, and the combination is refined and complements each other.

[0138] In summary, the Chinese medicine composition of the present invention comprises 3 to 9 parts of Taxus chinensis, 9 to 12 parts of Curcuma zedoaria, 15 to 30 parts of Astragalus membranaceus, 6 to 12 parts of Panax notoginseng, and 10 to 20 parts of Gynostemma pentaphyllum. In animal models, the Chinese medicine composition can reduce proteinuria, regulate lipid metabolism disorders, and protect the kidneys. Clinically, after using the Chinese medicine composition, all patients' 24-hour urine protein, plasma albumin, D-dimer, fibrinogen, triglycerides, total cholesterol, blood β2-microglobulin, and urine β2-microglobulin were improved or significantly improved compared to before 3 months of treatment. For severe MN patients with 24-hour urine protein greater than 3.5g, a combination of Chinese and Western medicine treatment using the Chinese medicine composition as the main component showed good therapeutic effects within 3 months. For patients with 24-hour urine protein less than 3.5g, the Chinese medicine composition alone showed good therapeutic effects. The Chinese medicine composition plays an important role in the treatment of MN.

[0139] Although the present invention has been described in detail through the above preferred embodiments, it should be understood that the above description is not intended to limit the present invention. After reading the above description, various modifications and substitutions of the present invention will become apparent to those skilled in the art. Therefore, the scope of protection of the present invention should be defined by the appended claims.

Claims

1. A Chinese medicine composition for treating membranous nephropathy, characterized in that: The traditional Chinese medicine composition comprises the following traditional Chinese medicine raw materials in parts by weight: 3 to 9 parts of Taxus chinensis, 9 to 12 parts of Curcuma zedoariae, 15 to 30 parts of Astragalus membranaceus, 6 to 12 parts of Panax notoginseng, and 10 to 20 parts of Gynostemma pentaphyllum.

2. The Chinese medicine composition for treating membranous nephropathy according to claim 1, wherein 5-7 parts of Taxus chinensis, 10-11 parts of Curcuma zedoaria, 18-25 parts of Astragalus membranaceus, 8-10 parts of Panax notoginseng, and 12-18 parts of Gynostemma pentaphyllum.

3. The Chinese medicine composition for treating membranous nephropathy according to claim 1, wherein 9 parts of Taxus chinensis, 12 parts of Rhizoma Curcumae, 30 parts of Radix Astragali, 12 parts of Radix Notoginseng, 20 parts of Gynostemma pentaphyllum.

4. The Chinese medicine composition for treating membranous nephropathy according to claim 1, wherein The traditional Chinese medicine composition also includes pharmaceutically acceptable excipients.

5. The Chinese medicine composition for treating membranous nephropathy according to any one of claims 1 to 4, characterized in that The dosage form of the Chinese medicine composition is selected from any one of tablets, ointments, granules, granules and oral liquids.

6. A method for preparing the Chinese medicine composition for treating membranous nephropathy according to any one of claims 1 to 5, characterized in that: The Chinese medicinal raw materials are dried, mixed evenly, and crushed into fine powder; or boiled in water for 2 to 4 hours, the decoction is concentrated, and dried to obtain the Chinese medicinal composition.

7. The preparation method according to claim 6, wherein After being crushed into fine powder, pharmaceutically acceptable carriers and / or excipients are added.

8. Use of the traditional Chinese medicine composition for treating membranous nephropathy according to any one of claims 1 to 5, characterized in that: The traditional Chinese medicine composition is used for preparing medicine for treating membranous nephropathy.

9. The use according to claim 8, characterized in that The traditional Chinese medicine composition is used for preparing medicine for treating glomerular diseases.

10. The use according to claim 8, characterized in that The traditional Chinese medicine composition is used for preparing medicine for treating nephrotic syndrome.

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